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SYSTEMATIC REVIEW published 17 March 2022 doi 10 3389 fneur 2022 815056 Systematic Review and Dosage Analysis Hyperbaric Oxygen Therapy Efficacy in Mild Traumatic Brain Injury Persistent Postconcussion Syndrome Paul G Harch Hyperbaric Medicine Unit Section of Emergency Medicine Department of Medicine Louisiana State University Health Sciences Center New Orleans LA United States Edited by Eric Peter Thelin Karolinska Institutet KI Sweden Reviewed by Ole Hyldegaard Rigshospitalet Denmark B Robert Mozayeni Translational Medicine Group PC Gaithersburg United States Background Mild traumatic brain injury results in over 15 of patients progressing to Persistent Postconcussion Syndrome a condition with significant consequences and limited treatment options Hyperbaric oxygen therapy has been applied to Persistent Postconcussion Syndrome with conflicting results based on its historical understanding definition as a disease specific therapy This is a systematic review of the evidence for hyperbaric oxygen therapy HBOT in Persistent Postconcussion Syndrome using a dose analysis that is based on the scientific definition of hyperbaric oxygen therapy as a dual component drug composed of increased barometric pressure and hyperoxia Received 15 November 2021 Accepted 18 January 2022 Published 17 March 2022 Methods In this review PubMed CINAHL and the Cochrane Systematic Review Database were searched from August 8 22 2021 for all adult clinical studies published in English on hyperbaric oxygen therapy in mild traumatic brain injury Persistent Postconcussion Syndrome symptoms present at least 3 months Randomized trials and studies with symptomatic and or cognitive outcomes were selected for final analysis Randomized trials included those with no treatment control groups or control groups defined by either the historical or scientific definition Studies were analyzed according to the dose of oxygen and barometric pressure and classified as Levels 1 5 based on significant immediate post treatment symptoms or cognitive outcomes compared to control groups Levels of evidence classifications were made according to the Centre for Evidence Based Medicine and a practice recommendation according to the American Society of Plastic Surgeons Methodologic quality and bias were assessed according to the PEDro Scale Citation Harch PG 2022 Systematic Review and Dosage Analysis Hyperbaric Oxygen Therapy Efficacy in Mild Traumatic Brain Injury Persistent Postconcussion Syndrome Front Neurol 13 815056 doi 10 3389 fneur 2022 815056 Results Eleven studies were included six randomized trials one case controlled study one case series and three case reports Whether analyzed by oxygen pressure or composite oxygen and pressure dose of hyperbaric therapy statistically significant symptomatic and cognitive improvements or cognitive improvements alone were achieved for patients treated with 40 HBOTS at 1 5 atmospheres absolute ATA four randomized trials Symptoms were also improved with 30 treatments at 1 3 Correspondence Paul G Harch pharch lsuhsc edu paulharchmd gmail com Specialty section This article was submitted to Neurotrauma a section of the journal Frontiers in Neurology Frontiers in Neurology www frontiersin org 1 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury ATA air one study positive and negative results were obtained at 1 2 ATA air one positive and one negative study and negative results in one study at 2 4 ATA oxygen All studies involved

Harch Hyperbaric Oxygen Traumatic Brain Injury Classification of Evidence Analysis of Studies analysis demonstrates Level I evidence for the efficacy of 40 HBOTs at 1 5 ATA of oxygen 46 75 in the treatment of blast or blunt mTBI PPCS Studies were classified as Level 1 5 according to CEBM guidelines 3 Classifications were based on significant immediate post treatment symptom and or cognitive outcomes in treatment groups vs no treatment control groups or lower doses of HBOT and compared according to the individual pressure total oxygen combined hyperbaric pressure and total oxygen doses of HBOT The pressure was measured in atmospheres absolute ATA and oxygen in atmosphere minutes AMs AM oxygen dose was the total cumulative oxygen dose over the entire treatment course per group for oxygen in excess of room air It was the sum of the product of the atmospheric pressure times the FiO2 times the number of minutes for each phase of every hyperbaric treatment compression at depth and decompression phase multiplied times the total number of treatments Constant compression and decompression rates were assumed The average pressure from the surface to depth compression and depth to the surface decompression was multiplied by the FiO2 of the breathing gas for these phases A linear increase in FiO2 and 90 FiO2 was assumed by 8 min of compression 98 for protocols in monoplace single person chambers that compressed with 100 oxygen Practice Grade Recommendation was determined from the levels of evidence according to the American Society of Plastic Surgeons grading system in Burns et al 4 METHODS This systematic review was reported in accordance with the Preferred Reporting Items for Systematic Reviews and MetaAnalyses PRISMA guidelines 108 The PRISMA checklist is attached as Supplementary Material Search Method In this review PubMed CINAHL and the Cochrane Systematic Review Database were searched without filters or time limits on August 18 8 and 8 2021 respectively for English language clinical articles using the terms hyperbaric oxygen as well as traumatic brain injury mild traumatic brain injury postconcussion syndrome or persistent postconcussion syndrome Reference lists were reviewed for additional studies Inclusion criteria consisted of studies with immediate post treatment symptom or cognitive outcomes and adult subjects 18 65 years old persistent postconcussion syndrome postconcussion symptoms lasting at least 3 months one or more blast or blunt mTBIs and treatment with HBOT Symptomatic outcomes were those employing TBI symptom questionnaires cognitive outcomes were computerized or manual cognitive tests Affective measures imaging balance eye tracking and sleep measures used as stand alone outcomes were not included The search process for inclusion and exclusion criteria was first search title screen second search abstract review of first search titles third search full text articles of abstracts fourth search detailed review of full text articles Manual data extraction was performed on the final selected articles from the abstracts results and conclusion sections of the articles and entered in Table 1 Numerical data included study publication year design number of subjects gender civilian or military status active duty or veteran education level time from TBI to treatment number of total lifetime TBIs type of TBI blast or blunt percent of subjects with comorbid post traumatic stress disorder PTSD traditional pressure oxygen dose parameters the total number of prescribed HBOTs in the protocol and outcome instruments Symptomatic and cognitive results of the studies were as stated in the abstracts and conclusions without numerical figures their numerical data and calculated oxygen dose study were entered in the text of the Section Results Symptom outcomes for the final selected studies were averaged and analyzed by treatment pressure and total oxygen dose All searches screening selection data extraction and analyses were performed by a single author No automation tools were used for any part of this study Missing data from some articles were obtained from companion articles of the same study on the same subjects If not available from companion articles it was noted as unidentified or unstated Studies were grouped by design for classification of evidence randomized trials cohort studies case controlled studies case series and case reports Reviews and pooled data analyses were excluded Frontiers in Neurology www frontiersin org Methodologic Quality and Risk of Bias Assessment The physiotherapy evidence database PEDro scale 109 was used to assess the methodologic quality risk of bias of the randomized trials included in the final analysis Randomized trials are given a cumulative score of 0 10 based on individual scoring of 10 11 items the first eligibility criteria is omitted from the score because it is an external validity item using 1 point for present and 0 points for absent The 10 items beginning with 2 are 2 random allocation 3 concealed allocation 4 groups similar at baseline 5 subject blinding 6 therapist blinding 7 assessor blinding 8 1 key outcome for 85 of subjects 9 1 key outcome intention totreat analysis 10 1 key outcome between group statistical comparison and 11 1 key outcome point measurements and variability Scoring was performed by the single author based on stated scoring items in the text of each article Additional reporting bias and conflict of interest were noted for investigators who were employees of the funding source All items were scored as 0 if they were not mentioned Attempts were made to contact the authors of the studies to resolve the scoring on allocation concealment Quality of studies poor fair good excellent was judged according to the scoring legend of Cashin et al 110 RESULTS In total 681 articles were searched 11 studies were included in the final analysis Figure 1 The largest exclusion 57 was for 3 March 2022 Volume 13 Article 815056

Time TBIs from TBI to Rx mos PTSD Pressure Dose of HBOT 7 Bla 100 1 5 ATA O2 60 min 39 twice day bid total dive time TDT 5d week 1 month monoplace chamber Sx PEx and SPECT Improvement Sx pre and within one PEx SPECT week post HBOT 1 1 Bla Bla U 1 5 ATA O2 60 min at 40 once day qd depth monoplace 5d week 80 in two 40 treatment blocks chamber separated by 1 month break Improvement Sx Sx ANAM mood and sleep pre and ANAM mood sleep within 4 6 weeks post HBOT time deduced from testing dates 33 6 2 7 Bla 100 Avg PCL 67 1 5 ATA O2 60 min TDT monoplace chamber 21 neuropsychological affective Quality of Life self assess tests Sx PEx SPECT pre and within one week after HBOT with phone questionnaire followup at 6 months post HBOT Significant improvement Sx PEx 15 21 outcome instruments and SPECT Significant worsening 1 21 instruments 3 71 3 4 Bla 33 Blu 8 both 9 U 1 3 ATA air vs 2 4 30 qd 5d week for ATA O2 both for 6 weeks 90 min at depth with two 10 min air breaks for the O2 group The 1 3 Air group slowly drifted to 1 2 ATA over the course of the treatment multiplace chamber PCL M and ImPACT Sx questionnaires pre weekly and 6 weeks after intervention Significant within group improvement on PCL M and ImPACT but no significant differences between groups 33 1 Blu U 1 5 ATA O2 for 60 min 40 qd 5d week for 8 Mindstreams at depth multiplace weeks computerized chamber cognitive test battery Quality of Life questionnaire SPECT pre and 1 3 weeks after the HBOT protocol Significant improvement in cognitive function and Quality of Life in both groups post HBOT none during control Increased brain activity on SPECT after HBOT compared to controls in agreement with cognitive improve 8 5 1 75 1 25 Bla U 0 21 1 5 and 2 0 ATA O2 at 2 ATA pressure 60 min at depth multiplace chamber No between group differences Rivermead and PCL Significant improvement PCL in 2 0 ATA O2 group Design Number of Age yrs subjects Male M Female F Unidentified or Unstated U Status Mi Educa Military AD tion Active Duty yrs V Veteran C Civilian Harch et al 66 Case Report 1M 25 Mi V 15 36 Wright et al 67 Case Report 2M 23 22 Mi AD U 8 Harch et al 68 Prospective 16 M Case Series included in Harch et al 80 30 Mi 8 AD 8 V 12 9 Wolf et al 69 Air Force funded RCT double blind 28 3 Mi AD Slightly 12 44 C 15 3 23 2 Mi AD U 4 Boussi Gross et al RCT cross over 56 24 M 32 F single blind 70 March 2022 Volume 13 Article 815056 Cifu et al 71 Defense Advanced Research Projects Agency DARPA funded RCT double blind 61 M Number of HBOTs 40 bid 5d week 1 month 40 qd 5d week within 10 weeks Outcome instruments Sx symptoms PEx physical exam PCS sand PTSD symptom questionnaires Rivermead and PCL M pre and immediately post intervention Results Continued Hyperbaric Oxygen Traumatic Brain Injury TBI Blast Bla Blunt Blu References 50 48 M 2 F Harch Frontiers in Neurology www frontiersin org TABLE 1 Retrieved analyzed studies and companion articles

Harch Frontiers in Neurology www frontiersin org TABLE 1 Continued Results 5 PTSD Pressure Dose of HBOT Number of HBOTs Outcome instruments Sx symptoms PEx physical exam 1 75 1 25 Bla U 0 21 1 5 and 2 0 ATA O2 at 2 ATA pressure 60 min at depth multiplace chamber 40 qd 5d week within 10 weeks No beneficial effect Battery of 55 cognitive 1 5 or 2 0 ATA O2 psycho motor and compared to sham balance tests pre and within 1 week post intervention 8 5 1 75 1 25 Bla U 0 21 1 5 and 2 0 ATA O2 at 2 ATA pressure 60 min at depth multiplace chamber 40 qd 5d week within 10 weeks 17 cognitive psychomotor functional Quality of Life tests pre within 1 week and 3 months post last HBOT No significant time by intervention interaction for any outcome measure U 8 5 1 75 1 25 Bla U 0 21 1 5 and 2 0 ATA O2 at 2 ATA pressure 60 min at depth multiplace chamber 40 qd 5d week within 10 weeks Computerized eye tracking measurement of fixation saccades and smooth pursuit pre immediately and 3 months post intervention No significant between group effects or 1 5 or 2 0 ATA O2 effects vs sham Mi AD 2 3rds with 12 years 22 9 3 1 U 65 47 72 Avg PCL C 51 3 1 2 ATA air or 1 5 ATA 40 qd within 10 O2 50 min at depth weeks vs No chamber treatment multiplace chamber RPQ 3 13 and 16 NSI ANAM pre after 20 and 40 treatments or 10 weeks PCL C depression anxiety pain sleep Quality of Life pre and after 40 treatments or 10 weeks No difference between air and O2 treatments both groups Significantly improved compared to no hyperbaric treatment 28 3 Mi AD Slightly 12 3 71 3 4 Bla 33 Blu 8 both 9 U 1 3 ATA air vs 2 4 30 qd 5d week for 6 ATA O2 both for weeks 90 min at depth with two 10 min air breaks The 1 3 Air group drifted to 1 2 ATA slowly over the course of the treatment multiplace chamber ImPACT ANAM ToVA PCL M pre weekly and 6 weeks after intervention Significant improve ImPACT visual memory and time processing ANAM code substitution recall match to sample and simple reaction PCL M within both groups no significant between group findings Sub groups identified 33 Mi AD Not stated for the 64 2 3rds with 12 years for the 72 subjects 24 4 U U for the 64 but 65 47 72 Avg PCL C 51 3 1 2 ATA air or 1 5 ATA 40 qd within 10 O2 60 min at depth weeks vs No chamber treatment multiplace chamber Simple Reaction Time SRT and Procedural Reaction Time PRT of ANAM pre at midpoint 6 weeks and within 1 month post intervention 13 weeks SRT worsened for local care group No significant differences between 3 groups over time Design Number of Age yrs subjects Male M Female F Unidentified or Unstated U EducaStatus Mi Military AD tion Active Duty yrs V Veteran C Civilian Walker et al 72 DARPA funded RCT part of Cifu et al 71 61 M 23 2 Mi AD U 8 5 Cifu et al 73 DARPA RCT part of funded Cifu et al 71 61 M 23 3 Mi AD U Cifu et al 74 DARPA RCT part of funded Cifu et al 71 61 M 23 3 Mi AD RCT Miller et al double blind 75 HOPPS Hyperbaric Oxygen Therapy for Persistent Post concussive Symptoms Army funded 72 69 M 3 F 31 4 Wolf et al 76 Air Force funded RCT part of Wolf et al 69 Air Force 50 48 M 2 F Churchill et al 77 HOPPS RCT Part of Miller et al 75 HOPPS 64 of 72 61 M 3 F Continued Hyperbaric Oxygen Traumatic Brain Injury March 2022 Volume 13 Article 815056 TBI Blast Bla Blunt Blu Time TBIs from TBI to Rx mos References

References Design Skipper et al 78 DARPA and HOPPS RCT 2 40 All M combined studies Cifu et al 71 DARPA and Miller et al 75 HOPPS RCT part of Shandley et al 79 Air Force funded Wolf et al 69 Air Force Harch et al 80 Number of Age yrs subjects Male M Female F Unidentified or Unstated U 28 U M F 6 Case Controlled 30 28 M 2 F Partial imaging control group of population normals 71 70 M 1 F Time TBIs from TBI to Rx mos TBI Blast Bla Blunt Blu PTSD Pressure Dose of HBOT Number of HBOTs Outcome instruments Sx symptoms PEx physical exam Results 28 1 Mi AD 57 5 with some college or more U 87 5 with multiple U U for the 40 as a group 0 21 1 5 and 2 0 40 qd within 10 ATA O2 at 2 ATA weeks both studies pressure 60 min at depth and 1 2 ATA air or 1 5 ATA O2 60 min at depth vs No chamber treatment multiplace chambers PCS PTSD anxiety DARPA PCS scores depression and worse 1 5 ATA O2 Quality of Life average improved 0 21 and 39 months post 2 0 ATA O2 groups treatment HOPPS PCS scores worsened in all groups U Mi AD U U U U U 1 3 ATA air vs 2 4 30 qd 5d week for ATA O2 both for 6 weeks 90 min at depth with two 10 min air breaks The 1 3 Air group drifted to 1 2 ATA slowly over the course of the treatment multiplace chamber ImPACT BrainCheckers PCL M pre and post 5 10 15 20 25 30 HBOTs and at 6 week followup Stem cells from peripheral blood flow cytometry pre post 15 and 30 HBOTs and at 6 week followup HBOT at 2 4 ATA correlated with stem cell mobilization and increased cognitive performance 30 3 Mi 11 AD 19 V 13 1 40 2 3 5 Bla 77 23 30 1 5 ATA O2 60 min Avg PCL 63 4 TDT monoplace chamber 21 neuropsych affective Quality of Life self assessment tests Sx PEx pre and within 1 week after HBOT with phone questionnaire followup at 6 months post HBOT SPECT pre after 1st and 40th HBOTs Significant improvement Sx neurol PEx IQ memoy attention dominant hand motor speed and dexterity Quality of Life anxiety PTSD depression suicidal ideation psychoactive medications Further Sx improve at 6 months Significant SPECT improvement after 1 and 40 HBOTs 32 8 Mi 68 AD 3V 25 6 3 6 Bla 32 49 Blu 20 avg 44 9 Combination of Bla and Blu 48 1 2 ATA air or 1 5 ATA 40 qd Monday to Sx Quality of Life O2 60 min at depth Friday over 12 weeks neuropsychological multiplace chamber neurological EEG sleep auditory vestibular autonomnic visual neuroimaging and laboratory testing pre at 13 weeks and 6 months post randomization 12 month post randomization online phone questionnaires Significant improvement NSI and PTSD Sx in HBOT group more pronounced for those with PTSD with regression at 3 and 9 months post treatment HBOT improved some cog processing speed and sleep measures Pts With PTSD and HBOT improved functional balance and reduced vestibular complaints at 13 weeks 82 some college or more 40 bid 5d week 1 month Continued Hyperbaric Oxygen Traumatic Brain Injury March 2022 Volume 13 Article 815056 Weaver et al RCT 81 BIMA Brain double blind Injury Mechanisms of Action DoD funded Status Mi Educa Military AD tion Active Duty yrs V Veteran C Civilian Harch Frontiers in Neurology www frontiersin org TABLE 1 Continued

Harch Frontiers in Neurology www frontiersin org TABLE 1 Continued Time TBIs from TBI to Rx mos TBI Blast Bla Blunt Blu PTSD Pressure Dose of HBOT Number of HBOTs Outcome instruments Sx symptoms PEx physical exam Results Design Number of Age yrs subjects Male M Female F Unidentified or Unstated U EducaStatus Mi Military AD tion Active Duty yrs V Veteran C Civilian Walker et al 82 BIMA DoD funded RCT Part of Weaver et al 81 BIMA 71 70 M 1 F 75 healthy volunteer 58 M 17 F 32 8 39 3Volunteers Mi 68 AD 3V 82 92 Mi 1 AD 21 V some 53 C college or more 25 6 3 6 Bla 32 49 Blu 20 avg 44 9 Combination of Bla and Blu 48 Sleep quality by Pittsburgh Sleep 1 2 ATA air or 1 5 ATA 40 qd Monday to O2 60 min at depth Friday over 12 weeks Quality Index PSQI self reports multiplace chamber sleep diary screen for signifcantly abnormal compared to normals obstructive sleep 70 obstructive apnea restless leg sleep apnea risk syndrome and cataplexy objective Significant actigraphic measures improvement PSQI of sleep wake All pre 5 8 measures at 13 weeks and 2 8 at 6 13 weeks and 6 months in HBO months post group compared to randomization air group No changes in other measures Meehan et al 83 BIMA DoD funded RCT Part of Weaver et al 81 BIMA 71 70 M 1 F 75 healthy volunteer 58 M 17 F 32 8 39 3Volunteers Mi 68 AD 3V 82 92 Mi 1 AD 21 V some 53 C college or more 25 6 3 6 Bla 32 49 Blu 20 avg 44 9 Combination of Bla and Blu 48 Dynamic 1 2 ATA air or 1 5 ATA 40 qd Monday to O2 60 min at depth Friday over 12 weeks posturography vestibular evoked multiplace chamber myo genic potentials tandem gait Romberg Sharpened Romberg Berg Balance Scale Beck Anxiety Inventory PTSD Checklist Civilian DSM IV PTSD Module Center for Epidemiologic Study Depression Scale all at baseline 13 weeks and 6 months post randomization 32 29 M 3 F 30 5 Mi 7 AD 12 V U C 13 114 U Bla 47 22 7 32 Blu 53 1 5 ATA O2 for 45 min 40 qd 5d wk with monoplace or 50 min option for 20 or 40 multiplace at depth additional HBOTs for residual symptoms 7 References Mozayeni et al 84 Prospective Case Series mTBI cohort worse than healthy volunteers on balance and gait measures and affective symptoms Significant improvement postural control favored HBOT but were minimal Those with affective Sx especially PTSD had the most improvement in postural control and otolith function after HBOT Continued Hyperbaric Oxygen Traumatic Brain Injury March 2022 Volume 13 Article 815056 ANAM and CNS Vital Improvement in 13 17 Signs computerized neurocognitive and 8 8 mood measures cognitive test batteries pre post 40 More than 40 HBOTs blast TBI less delay 60 and 80 to treatment and treatments patients with PTSD showed greater improvement

Harch Frontiers in Neurology www frontiersin org TABLE 1 Continued Design Number of Age yrs subjects Male M Female F Unidentified or Unstated U Status Mi Educa Military AD tion Active Duty yrs V Veteran C Civilian Hart et al 86 BIMA RCT Part of Weaver et al 81 BIMA 42 41 M 1 F Mi 40 AD 2 V U Hart et al 87 Air Force DARPA HOPPS BIMA RCT 4 pooled 254 DoD 248 M 6 F 29 3 135 3 other studies DoD studies Wolf et al 69 Air Force Cifu et al 71 DARPA Miller et al 75 HOPPS and Weaver et al 81 BIMA Mi AD Wetzel et al 88 BIMA RCT Part of Weaver et al 81 BIMA 71 70 M 1 F 75 healthy volunteer 58 M 17 F 32 8 39 Mi 68 AD 3V 82 92 Mi 1 AD 21 V some 53 C college or more Shytle et al 89 Case Report 3 2 M 1 F 27 3 Mi 2 V 1 U 1 college graduate 2 U 48 1 U 2 Harch et al 46 RCT cross over 50 21 M 29 F single blind 42 5 C 41 Mi 9 1 AD 8 V 14 0 HBOT 15 6 Control 55 2 33 8 Time TBIs from TBI to Rx mos 26 1 Some 21 7 college or more 58 8 high school grad 41 2 25 6 PTSD Pressure Dose of HBOT Number of HBOTs Outcome instruments Sx symptoms PEx physical exam Results 3 7 Bla 31 52 Blu 21 comination 48 1 2 ATA air or 1 5 ATA 40 qd Monday to PPCS PTSD Quality O2 60 min at depth Friday over 12 weeks of Life pain multiplace chamber depression anxiety alcohol use at 24 months 40 subjects 36 months 14 subjects post study Avg for 3 studies 3 4 4th study Cifu 75 with 1 TBI 43 by PCL For 3 58 by Clin studies 116 blast Inter view 22 blunt 43 both not reporter for Miller et al 75 Above Wolf 69 Cifu et al 71 Miller et al 75 Weaver et al 81 125 HBOT 106 sham 23 local care 3 6 Bla 32 49 Blu 20 avg 44 9 Combination of Bla and Blu 48 1 2 ATA air or 1 5 ATA 40 qd Monday to Eye movement O2 60 min at depth Friday over 12 weeks tracking for saccadic multiplace chamber and smooth pursuit pre at 13 weeks and 6 months post randomization No between group but within group improvement at all time points Normals and all BIMA subjects no longer significantly different at 13 weeks and 6 months 20 qd 5d week 30 bid 5d week 35 bid 5d week x 25 qd 5d week x 10 Computer assisted assessments type not specified pre post for all 3 1 patient also had pre post CNS Vital Signs Inc and 1 had pre post NeuroPsychTM Improvements on parameters within neuro cognitive domains symptoms reduction in suicidal related symptoms Symptom neuro psychological and psychological testing pre post and 2 month after last HBOT Weekly NSI during treatment for both groups Significant improvement PPCS and PTSD symptoms ANAM memory depression anxiety sleep quality of life vs control Crossover experienced same improvement after HBOT 8 March 2022 Volume 13 Article 815056 Final analysis studies in bold TBI Blast Bla Blunt Blu DoD 40 three studies 71 75 81 and 30 Air Force study 69 1 daily IEDs Bla 2 burn pit Blu 1 numerous Bla 1 IEDs 1 Blu at least 3 4 Bla 100 1 75 ATA O2 1 5 ATA O2 1 5 ATA O2 All at 60 min TDT monoplace chamber 3 9 0 Control no Rx vs 1 5 40 qd 5d week ATA O2 60 min TDT monoplace chamber Blu 45 Bla 5 PCS PTSD and neuropsychological measures for all studies No significant differences between groups at 24 and 36 months and mean scores near pre intervention values Trend of improvement HBOT for PCS and PTSD Sx verbal memory Dose response to increasing O2 pressure with greater effect with PTSD Direction of results consistent with other studies Hyperbaric Oxygen Traumatic Brain Injury References

Harch Hyperbaric Oxygen Traumatic Brain Injury FIGURE 1 PRISMA flow chart of article search and retrieval 108 outcomes The 11 studies are three case reports 66 67 89 a prospective case controlled series reported in two publications 68 80 a prospective case series 84 and six randomized trials 46 69 71 75 81 reported in 19 publications 46 69 79 81 83 85 88 Three of the randomized trials contained control groups 46 70 75 two with a crossover design 46 70 and the other three were comparative dosing studies 69 71 81 Four of the randomized trials were performed by the U S Department duplicate articles and the second largest exclusion was by title 38 An example of the third search was the Ma et al study 111 on HBOT for firefighters with lifetime TBI and chronic emotional distress The study was excluded due to the primary outcome of regional cerebral blood flow and the absence of symptomatic or cognitive outcomes The 11 studies that met the criteria for inclusion are listed in Table 1 along with companion articles on non primary Frontiers in Neurology www frontiersin org 9 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury cognitive and SPECT improvements could not be explained by placebo effects The study design was changed at the midpoint of Harch et al 68 to include a matched group of imaging controls for all 30 subjects and the full cohort of 30 subjects 27 with comorbid PTSD was reported as a case controlled study in Harch et al 80 Subjects had sustained 3 5 previous TBIs and were treated with 40 HBOTs at 1 5 ATA oxygen 3 420 AMs oxygen 3 35 years post index TBI Their PTSD symptom scores were 32 higher 63 4 than subjects in the four DoD studies 49 5 69 46 5 71 51 3 75 44 9 81 average 48 1 The 30 subjects experienced near identical improvements to the first 16 subjects with statistically stronger p values RPQ total post treatment minus pre treatment 13 5 10 4 CI 17 4 to 9 6 p 0 001 actual p value was less but the cutoff of 0 001 was used for this publication a significant reduction in suicidal ideation and reduction in psychoactive medication use SPECT brain imaging was abnormal in subjects compared to a matched cohort of population normals and normalized after HBOT in 75 of abnormal regions The prospective case series Mozayeni et al 84 reported outcomes on 32 military and civilian subjects treated 9 5 years after mTBI with 40 82 treatments average 55 8 at 1 5 ATA oxygen 4 326 AMs oxygen Mood changes and two computerized neurocognitive test batteries ANAM4 and CNS Vital Signs were administered TBI symptom questionnaires were not used Significant improvement was demonstrated on 8 8 Automated Neuropsychological Assessment Metrics ANAM mood 6 7 ANAM neurocognitive and 7 10 CNS Vital Signs neurocognitive measures Improved outcomes were associated with shorter delays to treatment younger age military status and increased numbers of HBOTs In summary both the prospective case controlled and case series demonstrated multi domain statistically significant outcomes benefits of HBOT at a dose of 1 5 ATA oxygen with one study 80 showing simultaneous and correlative improvements in brain blood flow deficits compared to a group of untreated population normals The six randomized trials 46 69 71 75 81 are separated into three comparative dosing studies 69 71 81 and three randomized controlled trials 46 70 75 based on the understanding of HBOT as a dual component drug therapy consisting of increased barometric pressure and hyperoxia 46 57 58 80 99 101 112 In each of the comparative dosing studies 69 71 81 and one of the controlled studies 75 the lower pressure compressed air groups 69 75 81 or normoxic oxygen group 71 were erroneously characterized as sham controls because the pressure dose and FiO2 of HBOT were less than the historically defined arbitrary limit of 1 4 ATA 100 oxygen 55 56 The inclusion of increased pressure and hyperoxia in these sham controls the two bioactive components of hyperbaric oxygen treatment precludes their characterization as shams they are additional doses of hyperbaric therapy and hence classified as comparative dosing studies 46 80 99 101 112 The first of the comparative dosing studies was conducted by Wolf et al 69 and randomized 50 military subjects 3 71 months after an average of 3 4 mTBIs to 30 1 3 ATA air or 2 4 ATA oxygen HBOTs 1 002 and 6 900 AMs oxygen respectively of Defense DoD 69 71 75 81 and were reported in multiple publications featuring different outcome instruments or followup periods Wolf et al 69 in Wolf et al 76 and Shandley et al 79 Cifu et al 71 in Walker et al 72 and Cifu et al 73 74 Miller et al 75 in Churchill et al 77 Weaver et al 81 in Walker et al 82 Meehan et al 83 Hart et al 86 and Wetzel et al 88 pooled data analyses Skipper et al 78 Hart et al 87 and adverse events and blinding in Churchill et al 85 Analysis was conducted on the three case reports 66 67 89 the case controlled series 80 the case series 84 and six randomized trials 46 69 71 75 81 The case reports consist of Harch et al 66 Wright et al 67 and Shytle et al 89 Harch et al 66 reported symptomatic neurological physical exam and brain blood flow imaging improvements in an mTBI PPCS veteran 3 years after the last of seven blast induced combat mTBIs using forty 1 5 ATA oxygen HBOTs in 1 month 3 420 AMs oxygen Wright et al 67 demonstrated improvement of post injury ANAM deficits symptoms mood and sleep complaints in two military members with 40 and 80 1 5 ATA oxygen HBOTs 3 990 and 7 980 AMs oxygen delivered 8 months after single blastinduced mTBIs Shytle et al 89 reported three cases with limited documentation of PPCS In the first case mTBI PPCS PTSD symptoms and computerized cognitive and mood test deficits abnormalities improved with 20 HBOTs at 1 75 ATA oxygen 1 992 AMs oxygen The second case experienced an improvement in symptoms and computerized affective and cognitive measures with 30 HBOTs at 1 5 ATA oxygen 2 544 AMs oxygen and the third patient improved symptoms and computerized cognitive assessments with 35 HBOTs at 1 5 ATA oxygen 2 964 AMs oxygen In summary six out of six cases demonstrated symptomatic improvement and five out of five demonstrated computerized cognitive testing improvement with HBOT treatment at 1 5 ATA or 1 75 ATA oxygen one patient Statistics were not reported for any patients in any of the case reports In the prospective case controlled series Harch et al 68 80 reported the first 16 68 and then all 30 80 military subjects with mTBI PPCS and PTSD The first 16 subjects 68 which were not case controlled experienced significant improvement in selfreported symptoms PPCS and PTSD symptom questionnaires RPQ total post treatment minus pre treatment 15 6 12 8 CI 22 7 to 8 5 p 0 0002 abnormal neurological exam 7 13 neuropsychologist administered cognitive tests two neuropsychologist administered affective instruments and two quality of life measures after 40 HBOTs and quantitative texture and statistical parametric mapping analysis of SPECT brain blood flow imaging after the first and 40th HBOT at 1 5 ATA oxygen 3 335 AMs oxygen Statistical parametric mapping analysis revealed significant increases in brain blood flow in 85 regions of the brain almost exclusively white matter and in the hippocampi that were consistent with the statistically and clinically significant increases in neuropsychologically measured working and delayed memory The SPECT findings were different from and inconsistent with placebo SPECT brain blood flow studies This non controlled portion of the casecontrolled study concluded that the significant symptomatic Frontiers in Neurology www frontiersin org 10 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury responsible for cognitive improvements in subsets of this group This claim and implication are inconsistent with the symptom data analysis of Scorza et al 113 and the 60 40 air group subset cognitive dominance and lack evidence of stem cell migration implantation in the brain Wolf et al 76 acknowledge the finding of Scorza et al 113 of significant improvement of subjects with co morbid PTSD but omit the major finding that the 2 4 ATA oxygen dose demonstrated a trend toward harm in PPCS without PTSD 113 At the same time it is known that stem cells are mobilized into the circulation of humans at 2 0 and 2 5 ATA oxygen 114 in the absence of PPCS and PTSD and that stem cells are stimulated to proliferate and differentiate at sites of injury in the brain due to HBOT 115 117 but there is no evidence of deposition of circulating stem cells in brain tissue in the Wolf et al study 69 76 79 More likely the circulating stem cells had nothing to do with the improvement in cognition in the PPCS PTSD group since cognitive and symptom improvements were favored in the air pressurization group that had no evidence of stem cell mobilization In summary these cognitive outcome reports 76 79 do not report actual data appear to be misleading and the conclusions of the study cognitive improvements in the 2 4 ATA oxygen group vs the air group are internally inconsistent and inconsistent with their own analysis 113 and the PPCS symptom data in Wolf et al 69 The second comparative dosing study 71 was reported in four publications 71 74 that randomized 61 military subjects 8 5 months post mTBI to one of three 40 treatment 2 0 ATA pressure groups with either 0 21 1 5 or 2 0 ATA oxygen 76 3 720 and 4 860 AMs oxygen In the first publication 71 there were no significant within group improvements post minus pre mean change scores sham 0 05 p 0 98 1 5 ATA oxygen 1 24 p 0 61 2 ATA oxygen 3 77 p 0 19 on the TBI Rivermead Postconcussion Questionnaire RPQ at 10 weeks post first HBOT immediate one week post treatment Similar to Wolf et al 69 no between group change scores were statistically analyzed Rather one way ANOVAs were performed on the pre treatment mean RPQs for the three groups and then on the post treatment mean RPQs Identical analyses were performed on the PCL M for PTSD symptoms the 2 ATA oxygen group experienced the only significant finding in the study a significant decrease in PTSD symptoms The second publication 72 reported cognitive and psychomotor outcomes 1 week posttreatment and found no immediate postintervention beneficial effect of exposure to either 1 5 ATA or 2 0 ATA oxygen compared with the Sham Air intervention Scrutiny of the data suggests varying effects of the different doses of HBOT on the 55 outcomes especially for four cognitive tests California Verbal Learning Test Long Delay Free Recall Index for Recognition Short Delay Cue Recall and Recognition Total Hits but large standard deviations of the mean scores and separate one way ANOVAs of the baseline and post treatment outcomes make exact comparisons between the groups difficult Identical to the first publication on this study no change score analysis was performed on any outcome The third publication 73 reported 1 week and 3 month symptom functional cognitive and psychomotor outcomes and found No significant time by intervention interaction The authors found 6 week post treatment symptom reductions improvements of 13 in the HBOT group and 41 in the 1 3 ATA air group with 10 subtests improved on the Immediate Post Concussion Assessment and Cognitive Testing ImPACT symptom testing for the 1 3 ATA air group vs two subsets in the 2 4 ATA oxygen group but no within or between group change score statistics were performed on this data For the combined ImPACT symptom and cognitive total score 6 weeks post treatment there were statistically significant reductions improvements in both groups 12 in the HBO2 group and 31 in the air group control group t 3 76 p 0 001 and the HBO2 group t 3 9 p 0 001 but no statistical analysis was performed on the between group improvements Similarly there were statistically significant within group improvements in 6 weeks post treatment PTSD symptoms but no change score comparison between the two groups Rather it was stated that Difference testing between the sham control and HBO2 groups did not reveal any significant differences on the PCL M composite mean score t 0 205 p 0 84 or on the ImPACT total mean score t 0 943 p 0 35 at any time Figures 1 2 including at 6 weeks post exposure Table 1 In a subset analysis of this study Scorza et al 113 reported that the mTBI PPCS subjects without PTSD demonstrated a trend toward harm worsening of scores on the ImPACT symptom score in the 2 4 ATA oxygen group This indicates a larger positive effect on 2 4 ATA subjects with comorbid PTSD and a negative effect on PPCS the target of the study without PTSD Amplification of HBOTinduced PPCS benefit by comorbid PTSD was found in multiple studies 68 80 81 87 The negative finding of Scorza 113 and corresponding 2 4 ATA oxygen group composite symptoms and cognition ImPACT score trajectory Figure 2 in Wolf et al 69 and Figure 4 in Harch et al 46 have been argued as a toxic overdose effect of oxygen in the 2 4 ATA oxygen group 46 80 99 The resulting cognitive outcomes in the study of Wolf et al 69 were published in Wolf et al 76 and showed significant within group improvement for both groups but no statistically significant between group improvements for cognitive measures as well as PTSD symptoms Curiously there are no data presented in the paper Rather a derived metric the relative risk of improvement RROI on subset historical features e g 3 concussions single event

Harch Hyperbaric Oxygen Traumatic Brain Injury generally favoring HBO2 Those with affective symptoms particularly PTSD who were treated with HBOT had the most improvement in postural control and otolith function Hart et al 86 was an extended outcome study and not pertinent to this review Lastly Wetzel et al 88 demonstrated significant abnormalities in ocular metrics in the overall subject population compared to population normals Multiple abnormalities improved in both treatment arms pressurized air and HBOT that persisted by 3 months post treatment at which time the normals and study subjects were statistically indistinguishable In summary the comparative dosing studies 69 71 81 demonstrated significant improvements in symptoms of mTBI PPCS at 1 5 ATA oxygen and 1 3 ATA air deterioration at 1 2 ATA air and 2 4 ATA oxygen and no significant change at 2 ATA of pressure with 0 21 1 5 or 2 0 ATA oxygen In addition significant improvements in cognition balance and gait measures were obtained with the 1 5 ATA oxygen dose The first randomized controlled trial RCT 70 randomized 67 civilians 33 months after a single mTBI to 40 HBOTs at 1 5 ATA oxygen or no treatment control 3 720 vs 0 AMs oxygen in 2 months Mindstreams computerized testing organized into four cognitive domains single photon emission computed tomography SPECT brain blood flow imaging and a quality of life evaluation were performed pre within 1 3 weeks after control and treatment periods for the control crossover group and after treatment for the HBOT group Significant improvements were demonstrated in cognitive function and quality of life in both groups following HBOT but no significant improvement was observed following the control period SPECT imaging revealed elevated brain activity in good agreement with the cognitive improvements Cognitive function improvements in the HBOT group consisted of Information Processing Speed t 31 4 20 p 0 0001 Attention t 31 3 26 p 0 005 Memory t 31 4 13 p 0 0005 and Executive Functions t 31 3 72 p 0 0005 Similar to the Cifu et al studies 71 73 the outcome instrument change scores after the control period and HBOT were not statistically compared Rather the groups were compared at baseline and after both groups had completed HBOT The groups were statistically indistinguishable at both comparisons and both groups had experienced significant within in group improvements after HBOT on all instruments with medium to large effect sizes The second RCT 75 randomized 72 military subjects 23 months after an average of 3 1 mTBIs to one of three groups 40 HBOTs at 1 5 ATA oxygen 1 2 ATA air or no HBOT 3 120 600 or 0 AMs oxygen during 10 weeks Symptom questionnaires computerized neuropsychological testing and traditional neuropsychological testing were administered pre at the midpoint of treatment and after treatment for symptom questionnaires and the computerized cognitive testing and at two time points pre post full treatment for the traditional cognitive tests There were no significant two point change score differences between the three groups on the abbreviated RPQ p 0 24 yet a seemingly large difference between the HBOT and no treatment group 25 of the no treatment group 52 of the HBOT group and 33 of the air pressure group it does not appear that either treatment group was compared F 4 63 7 1 p 0 41 however a secondary F ratio test demonstrated significant improvements regardless of treatment group on five cognitive tests Trails B California Verbal Learning Test Paced Auditory Serial Addition Test Benton Visual Memory Test and Controlled Oral Word Association Test with worsening of the Wechsler Adult Intelligence ScaleIII Working Memory at 2 weeks post treatment Similar to the second publication 72 strict pre post change scores treatment effects for each group were not calculated and similar to both the first 71 and second 72 publications the pre post means of these differences were not compared between groups Regardless the treatment effects for each individual HBOT does group appear small and insignificant The net result is a suggestion of small differing effects of the three different doses of HBOT with statistically significant change only seen for reduction of PTSD symptoms in the 2 ATA oxygen group and some cognitive domains in the combined three groups The third comparative dosing study 81 randomized 71 military subjects 25 6 months after an average of 3 6 TBIs to receive either forty 1 2 ATA air or 1 5 ATA oxygen treatments 600 and 3120 AMs oxygen in a 12 week period Symptom quality of life neuropsychological neurological electroencephalography sleep auditory vestibular autonomic visual neuroimaging and laboratory testing were performed before and immediately 3 and 9 months post treatment Baseline characteristics suggested worse brain injury in the HBOT group due to significantly more frequent diffuse traumatic axonal injury on MRIs greater TBI symptoms RPQ 13 and RPQtotal PTSD symptoms PCL C hyperarousal score number of combat deployments and worse anger control Despite this difference in injury the compressed air group showed a deterioration in postconcussive NSI and RPQ and PTSD symptoms PCL C during the treatment period while the HBOT group experienced significant improvement on the NSI RPQ 3 and PCL C compared to the compressed air group difference in mean change score between groups NSI 7 6 CI 14 4 to 0 7 p 0 03 RPQ 3 1 5 CI 2 7 to 0 3 p 0 01 RPQ 13 5 CI 10 7 to 0 6 p 0 08 PCL C 7 3 CI 13 5 to 1 p 0 02 statistically moreso on the NSI and PCL C for those subjects with PTSD In addition the HBOT group experienced significant improvement in some cognitive processing speed and sleep measures and in those with PTSD improved functional balance and reduced vestibular complaints all compared to the pressurized air treatment Companion publications 82 83 86 88 presented components of Weaver et al 81 in greater detail Walker et al 82 reported markedly disrupted sleep quality in study subjects and significant improvement in self reports of Pittsburgh Sleep Quality Index measures five out of eight at 13 weeks and two out of eight at 6 months in the oxygen group compared to the air pressure group Meehan et al 83 found worse balance gait measures and more affective symptoms in mTBI subjects compared to healthy controls some within group improvements in these domains favoring the HBOT group at 13 weeks and 6 months post treatment significant but minimal improvements in the HBOT group compared to the air group on balance measures and improvements on postural control Frontiers in Neurology www frontiersin org 12 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury Non significant improvement was obtained in the other coprimary outcome Working Memory mean difference in change scores between groups 1 5 6 5 CI 2 23 to 5 13 p 0 431 Overall the HBOT group achieved significant improvement in 11 14 outcome instruments compared to 13 14 after HBOT in the crossed over Control Group Twelve of the fourteen outcome instruments were administered by the blinded neuropsychologist at the non treatment testing site The other two outcomes TBI and PTSD symptom questionnaires were collected at the treatment site by the unblinded hyperbaric technician individually to the no treatment group For the total RPQ score both chamber groups demonstrated significant improvement compared to the no treatment control group 5 4 95 CI 0 5 to 11 3 P 0 008 HBO group 7 95 CI 1 0 to 12 9 P 0 02 air group and no difference between the two chamber groups P 0 70 Significant improvements were achieved despite many subjects not receiving the full 40 treatments 48 of the HBOT group and 59 of the sham group received 40 HBOTs while symptom improvement was greater for those who completed all 40 chamber sessions For the secondary symptom outcome score Neurobehavioral Symptom Inventory NSI the no treatment group showed a slight worsening while both treatment groups showed improvement Pre minus post scores 1 1 CI 7 3 to 5 2 no treatment vs 3 7 CI 3 7 to 11 2 HBOT and 6 9 CI 1 4 to 12 4 sham air Positive scores are improvement Similar to the RPQ two point change metric it appears that neither treatment group was statistically compared to the no treatment group Rather it was stated that these change scores were not statistically different For PTSD symptoms depression generalized anxiety pain sleep quality of life and computerized cognitive testing improvements occurred in both groups generally favoring the air treatment but there was no statistical difference between treatment groups Again none of these data appear to be compared to the no treatment control The third RCT 46 randomized 60 civilian and military subjects 4 6 years after an average of 3 9 mTBIs to 40 HBOTs at 1 5 ATA oxygen 3 420 AM s oxygen in 8 9 weeks vs no treatment 0 AM s The no treatment control group was then crossed over to 40 HBOTs similar to Boussi Gross et al 70 Subjects completed symptom and quality of life questionnaires and neuropsychologist administered neuropsychological psychological and sleep testing or questionnaires The HBOT group experienced significant improvements compared to the control group in one of the two co primary outcomes NSI mean difference in change scores between groups 23 9 9 2 CI 29 2 to 18 6 p 0 0001 with the greatest improvement in the cognitive domain This was the largest percent improvement 52 in symptomatic outcome for a treatment group in all of the studies reviewed The authors noted that this outcome was likely positively biased by the daily interaction of the P I with the subjects during treatment an IRB safety requirement and completion of the symptom questionnaire at the treatment site The 52 figure however was in the range of one other study outcome figure 37 for the same symptom questionnaire and same treatment dose 75 where the P I and subjects were blinded to treatment Analysis of the eight Diagnostic and Statistical Manual IV Text Revised PPCS symptoms 2 demonstrated that the Treatment Group subjects experienced significant improvement in all eight of the PPCS definition symptoms while the Control Group experienced worsening on six of eight symptoms similar to the Weaver et al 81 1 2 ATA air treatment and the Miller et al 75 no treatment groups The HBOT group also experienced significant improvements in Memory Index ANAM Hamilton Depression Scale Hamilton Anxiety Scale Post Traumatic Stress Disorder Checklist Pittsburgh Sleep Quality Index and Quality of Life after Brain Injury compared to the Control Group Frontiers in Neurology www frontiersin org Dose Analysis of Studies Effects of composite pressure and oxygen doses of HBOT on immediate post treatment symptoms are presented in Table 2 for Table 1 randomized trials and case controlled series with symptom outcomes data abstracted from Table 7 in Harch et al 46 The Wolf et al 69 symptom data in Table 7 was recalculated to the immediate post treatment period using the ImPACT symptom data in Table 2 of Wolf et al 69 and the total ImPACT score symptoms and cognition from Exposure Interval 6 in Figure 2 of Wolf et al 69 The data show symptom improvements at 1 5 ATA oxygen and 1 3 ATA air improvements and deteriorations at 1 2 ATA air no effect at the normoxic oxygen control 0 21 ATA oxygen and one dose of oxygen 1 5 ATA oxygen both at 2 ATA pressure an improvement at the second oxygen dose 2 0 ATA and no change at 2 4 ATA oxygen According to the Scorza subset analysis 113 however and trend toward harm of the 2 4 ATA oxygen dose for subjects with mTBI PPCS without PTSD the actual effect of 2 4 ATA oxygen on mTBI PPCS without PTSD is negative The positive 75 and negative 81 outcomes at 1 2 ATA air may be explained by the 50 greater number of patients with comorbid PTSD in Miller et al 75 a subject demographic associated with larger treatment effects in multiple studies 68 80 81 87 and the 70 of subjects at risk for sleep apnea and post treatment testing at altitude in Weaver et al 81 phenomena that would favor negative treatment effects Pressure dose effects on immediate post treatment symptom outcomes for the studies in Table 2 are presented in Figure 2 An asymmetric bell shaped dose response curve shows maximum symptom improvement in the 1 3 1 5 ATA pressure range with much less improvement at lower and higher pressures Oxygen dose effects on symptom outcomes for Table 2 studies are in Figure 3 and demonstrate a similar asymmetric bell shaped doseresponse curve with maximal results in a broad range from 1 002 to 4 860 atmosphere minutes of oxygen and decreased responses at the lowest and highest doses of oxygen The only aberrant value 3 720 is possibly due to the 2 ATA pressure component of the 1 5 ATA O2 2 ATA dose in the Cifu et al study 71 the identical oxygen dose was delivered in Boussi Gross et al 70 at 1 5 ATA pressure with positive cognitive mood and functional imaging changes Analysis of Figures 2 and 3 suggests that barometric pressure in the narrow range of 1 3 1 5 ATA has a more important effect on mTBI PPCS symptoms than oxygen pressure where a near equal oxygen effect occurs from 1 002 to 3 420 4 860 AMs 13 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury TABLE 2 Immediate post treatment symptom changes according to pressure and oxygen dose in mTBI PPCS studies abstracted from Table 7 in Harch et al 46 with change in polarity Pressure dose Oxygen dose atmosphereminutes No chamber treatment 1 2 ATA air 1 3 ATA air 1 5 ATA O2 2 0 21 ATA press O2 2 0 1 5 ATA press O2 2 0 2 0 ATA press O2 2 4 ATA O2 0 46 75 600 75 81 1 002 69 3 120 75 81 3 420 46 80 76 71 3 720 71 4 860 71 6 900 69 31b Wolf et al 69 2 8b Cifu et al 71 Miller et al 75 1 2a 3c 35a 21c 4 a 12 a 37a 11c 36a Harch et al 80 21a 13c Weaver et al 81 Harch et al 46 a 5 6c 2a 10c 52c Scores are percent change Post minus Pre Pre with polarity reversed so that positive represents symptom improvement a Rivermead Postconcussion Questionnaire b Immediate Post Concussion Assessment and Cognitive Testing ImPACT c Neurobehavioral Symptom Inventory NSI Values for both figures are symptom score improvements from Table 2 ImPACT symptom data in Wolf et al 69 that were interpolated to the immediate post treatment time period using the ImPACT total scores symptoms plus cognition from Figure 2 Wolf et al 69 exposure Interval 6 and applying the percent reduction in composite score from Pre exposure to Exposure Interval 6 in Figure 2 Wolf et al 69 to the Pre exposure ImPACT symptom scores in Table 2 Wolf et al 69 FIGURE 2 Symptom Improvements from Table 2 vs pressure dose for HBOT mTBI PPCS studies Symptom percent values represent average percent improvement in symptoms at a given pressure in Table 2 averaging the three different instrument ImPACT NSI and RPQ symptom outcomes Classification of Evidence Methodologic Quality and Risk of Bias Assessment 4 are presented in Tables 3 4 The Level of Evidence and PEDro methodologic quality bias assignments along with PEDro scoring are in Tables 5 6 Miller et al 75 and Harch et al 46 meet Level 1B criteria with a minus sign to designate The CEBM Levels of Evidence 3 and American Society of Plastic Surgeons Grade Practice Recommendation grading systems Frontiers in Neurology www frontiersin org 14 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury FIGURE 3 Symptom Improvements from Table 2 vs oxygen dose for HBOT mTBI PPCS studies Symptom percent values represent the average percent improvement in symptoms for each oxygen dose in Table 2 averaging the three different instrument ImPACT NSI and RPQ symptom outcomes Level 2 studies Boussi Gross et al 70 is a randomized and true controlled crossover trial but did not have published confidence intervals a CEBM 3 requirement for Level 1 classification All four of these studies are given a 2B designation with a plus sign to denote a higher level than true Level 2 cohort studies There is one Level 3 study 80 one Level 4 case series 84 and three Level 5 case reports 66 67 89 Harch et al 68 was omitted from Table 5 due to the inclusion of its 16 subjects in Harch et al 80 The two randomized controlled Level 1B trials of Miller et al 75 and Harch et al 46 demonstrated the benefit of the treatment groups at 1 5 ATA oxygen and one at 1 2 ATA air vs control groups The additional comparative dosing study of Weaver et al 81 Level 2B demonstrated additional proof of the efficacy of the 1 5 ATA oxygen dose compared to a 1 2 ATA air group along with the Level 2B randomized controlled crossover study of Boussi Gross et al 70 the case controlled study of Harch et al 80 Level 3 the case series of Mozayeni et al 84 Level 4 and the 3 case reports 66 67 89 Level 5 PEDro analysis of methodological quality bias is presented in Table 6 According to PEDro scoring statistics 119 studies scoring 6 10 on the PEDro scale are considered moderate to high quality Using the qualitative assessment of Cashin et al 110 for the entire scale of PEDro Scale scores studies were rated as poor

Harch Hyperbaric Oxygen Traumatic Brain Injury TABLE 4 Grade practice recommendations from the American Society of Plastic Surgeons in Burns et al 4 Grade Descriptor Qualifying evidence Implications for practice A Strong recommendation Level I evidence or consistent findings from multiple studies of levels II III or IV Clinicians should follow a strong recommendation unless a clear and compelling rationale for an alternative approach is present B Recommendation Levels II III or IV evidence and findings are generally consistent Generally clinicians should follow a recommendation but should remain alert to new information and sensitive to patient preferences C Option Levels II III or IV evidence but findings are inconsistent Clinicians should be flexible in their decision making regarding appropriate practice although they may set bounds on alternatives patient preference should have a substantial influencing role D Option Level V evidence little or no systematic empirical evidence Clinicians should consider all options in their decision making and be alert to new published evidence that clarifies the balance of benefit vs harm patient preference should have a substantial influencing role From the American Society of Plastic Surgeons Evidence based clinical practice guidelines Borrowed from Swanson et al 118 HBOT group in Harch et al 46 dropped out 81 of allocated subjects retained for personal financial hardship substance abuse an intercurrent new diagnosis of cancer and work conflicts all random events that precluded achieving the 85 allocation threshold for outcome data analysis by PEDro criteria An additional source of bias not addressed in the PEDro analysis is bias from conflict of interest Dr Harch stated conflict of interest in his studies 46 80 Many of the investigators in the low bias studies 69 71 75 81 are employed by the funding source TABLE 5 Levels of evidence for reviewed HBOT mTBI PPCS studies and PEDro Scale methodologic quality bias ratings for randomized trials Studies Levels of evidence Methodologic Quality Bias analysis PEDro Scale 109 Miller et al 75 Harch et al 46 1B 1B 9 5 Boussi Gross et al 70 Wolf et al 69 Cifu et al 71 Weaver et al 81 2B 2B 2B 2B 4 8 9 8 Harch et al 80 3B imaging control Mozayeni 84 4 Harch et al 66 Wright et al 67 Shytle et al 89 5 5 5 Side Effects Side effects and adverse events were minor but some were more frequent than expected and others were both notable and unusual no treatment induced serious adverse events were reported Harch et al 80 reported a higher incidence of minor and unusual adverse events middle ear barotrauma 20 exacerbation of PTSD anxiety 6 7 and transient worsening of symptoms at the midway point 23 Investigators attributed the high incidence of barotrauma to the twice day frequency of HBOT in patients who developed upper respiratory infections during the treatment course Barotrauma occurred in the prodromal phase of their upper respiratory illnesses before patients were symptomatic and could be paused from treatment The twice day frequency prevented the identification of those with developing infections that could have precluded barotrauma Wolf et al 120 documented middle ear barotrauma 5 51 and sinus squeeze confinement anxiety headache nausea numbness heartburn musculoskeletal chest pain latex allergy and hypertension 0 07 0 61 Boussi Gross et al 70 and Cifu et al 71 did not mention report side effects or adverse events Mozayeni et al 84 reported no side effects or complications Churchill et al 85 summarized adverse events for two DoD studies Miller et al 75 and Weaver et al 81 and reported minor adverse events in 40 120 subjects 33 Among these 40 subjects 31 had experienced barotrauma otic 17 sinus 8 3 and tooth 8 The high otic and sinus barotrauma figures were due to 24 otic and 11 sinus barotrauma incidences in one study 81 Scores of

Harch Hyperbaric Oxygen Traumatic Brain Injury TABLE 6 PEDro analysis of methodologic quality and bias for randomized trials Items Wolf et al 69 BoussiGross et al 70 Cifu et al 71 Miller et al 75 Weaver et al 81 Harch et al 46 1 eligibility criteria specified Y Y Y Y Y Y 2 random allocation Y Y Y Y Y Y 3 concealed allocation Y N Y Y Y Y 4 groups similar at baseline Y Y Y Y N Y 5 subject blinding Y N Y Y Y N 6 therapist blinding N N Y N N N 7 assessor blinding Y Y Y Y Y N 8 1 key outcome for 85 subjects Y N Y Y Y N 9 1 key outcome intention to treat analysis N N N Y Y N Y 10 1 key outcome between group statistical comparison Y N Y Y Y 11 1 key outcome point measurements and variability Y Y Y Y Y Y Total 8 4 9 9 8 5 approximately four fold the rates in the other study 6 4 75 Since the treatment pressures were identical in the two studies a difference in chamber operations equipment likely accounts for the high rate and discrepancy between studies Additional adverse events for the two studies included headache 6 7 dizziness vertigo 2 5 vision change 2 5 anxiety and somnolence 1 6 each and dyspnea neck irritation eye pruritis or hyperventilation 0 8 each Shytle et al 89 noted anxiety and GI discomfort in one of 3 patients at 25 HBOTs In Harch et al 46 middle ear barotrauma occurred in 2 along with three notable and unusual adverse events a predicted consented perforation of a multiply previously perforated tympanic membrane in 1 patient 2 and increasing late protocol fatigue with a transient reversal of improved cognitive PPCS symptoms in two patients at 34 and 39 HBOTs These late deteriorations were attributed to oxidative stress independent overlapping and interactive gene expression and suppression effects 123 124 This bioactivity is translated into wound repair and improved symptoms in diverse mostly wound conditions 56 This scientific understanding of hyperbaric oxygen therapy has eluded the hyperbaric medicine field for 359 years particularly in the last 60 years and has been confused and thwarted by the arbitrary definition of HBOT at a minimum pressure of 1 4 ATA of 100 oxygen 55 56 To rectify the confusion help resolve the controversy of the mTBI PPCS studies and enable this systemic review the arbitrary definition was supplanted by the scientific definition 58 Symptom outcomes were chosen as the primary outcome in this review due to their broad reflection of the heterogeneity of TBI their applicability to medical practice and a recommendation by the U S FDA to prioritize a PPCS symptom questionnaire the NSI as a primary outcome for an RCT on HBOT 46 in mTBI PPCS 57 This choice and the FDA s recommendation were reinforced by the results of the Weaver et al 81 study where investigators concluded that the NSI would be a reasonable simple primary outcome measure in future studies after an extensive battery of tests in all of the Department of Defense HBOT mTBI PPCS studies revealed that more resource intensive measures did not prove useful for measuring the change in this population The FDA also recommended that a series of clinical investigations with varying doses of pressure and hyperbaric hyperoxia should be performed 57 Inadvertently the combination of civilian and DoD HBOT mTBI studies above have addressed these criteria with broad based symptom outcomes ImPACT NSI RPQ or both NSI and RPQ and a variety of pressures and levels of hyperbaric hyperoxia This symptom based outcome review shows that regardless of the heterogeneity of TBI and the studies herein as well as negative reporting biases in studies 69 71 75 81 and positive biases in studies 46 80 analysis of the studies by outcomes pressure dose oxygen dose or the composite dose of HBOT reveals that all symptom based outcome studies performed at DISCUSSION Mild TBI is a heterogenous physical injury to the brain 7 27 that causes a wide range of signs symptoms and outcomes that have been defined in the chronic state as the Persistent PostConcussion Syndrome 2 TBI heterogeneity is reflected in the reviewed studies on HBOT in mTBI PPCS in different subject populations civilian and military active duty and veteran a wide range of ages 18 60 years different etiologies blunt and blast varying times to injury 3 months 71 to 46 years 84 PTSD or no PTSD varied dosing protocols of pressure and oxygen and an extensive list of outcome instruments HBOT is classified as a medical gas by the U S Centers for Drug Evaluation and Research of the FDA and consists of two components increased barometric pressure and hyperoxia 57 58 112 which have been argued to have varying effects on mTBI PPCS depending upon the doses of pressure and hyperoxia employed 46 80 99 101 112 Both have demonstrated a wide range of bioactivity 56 99 112 121 125 including Frontiers in Neurology www frontiersin org 17 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury of 6 months post TBI average 5 09 years and had sustained an average of 7 TBIs Patients were treated in a 4 week intensive residential program that included over 20 different therapies at least 20 different therapists and multiple physicians Results from 474 to 1 174 of the most severely symptomatic numbers of subjects varied per outcome instrument reported showed a decrease in NSI of 44 Post Traumatic CheckList of 23 General Anxiety Disorder scale 7 of 50 and Personal Health Questionnaire 8 of 50 Headache a marker of TBI 131 had the least improvement 4 points 6 5 reduction half the amount that is considered a clinically significant change 130 The only study in this review with the same therapy benefit was the Harch et al study 80 also a non controlled study whose subjects were a minimum of 6 months post TBI average 3 35 years with an average of 3 5 TBIs After 4 weeks of treatment with a single biological therapy one technician one physician and a neuropsychologist subjects RPQ decreased by 36 Post traumatic CheckList by 26 General Anxiety Disorder7 by 40 and Personal Health Questionnaire 9 by 48 In contrast to DeGraba et al 130 headache the second most responsive symptom next to dizziness 80 was reduced in 93 of subjects indicating treatment of the underlying TBI 131 This was achieved at a cost for testing and treatment of 14 656 subject figures from personal communication with Harch investigators adjusted from 2012 to 2021 dollars The National Intrepid Center of Excellence is a 65 million center 132 new centers are 14 million 133 and the average cost of treatment in 2021 dollars adjusted from 2015 National Intrepid Center of Excellence costs were 17 153 veteran 134 U S Veterans Administration 2021 adjusted costs from the 2012 U S Congressional Budget Office report were 15 589 veteran for TBI 18 387 for TBI with PTSD 135 The positive findings for the 1 5 ATA oxygen dose in the mTBI PPCS studies are further supported by positive outcomes in multiple studies on HBOT at 1 5 ATA in acute severe TBI 136 143 and one study in moderatesevere subacute TBI 144 suggesting a shared sensitivity of TBI to 1 5 ATA oxygen regardless of acuity or severity An additional RCT performed at 2 5 ATA in acute severe TBI 145 proved to be toxic very similar to the 2 4 ATA oxygen dose in Wolf et al 69 113 The acute treatment course in that study 145 was interrupted in multiple subjects due to pulmonary oxygen toxicity Similar to Wolf et al 69 the neurological results were minimal at this high oxygen dose no difference in mortality or duration of coma was demonstrated between groups 145 The sole positive finding was an interim outcome increased recovery of consciousness at 30 days in a subgroup of HBOT treated subjects with brainstem contusion 1 5 ATA pressure and pressure of oxygen 46 75 80 81 show symptom reductions in mTBI PPCS Two of these 1 5 ATA HBOT studies 75 81 are rated as CEBM Level 1 and 2 with good to excellent methodologic quality low bias PEDro scores of 9 and 8 substantially higher than the average 5 1 scores of 37 417 PEDro scored randomized controlled physiotherapy trials 119 a category of clinical trials that share blinding challenges similar to hyperbaric medicine and surgery The third of these 1 5 ATA oxygen trials the RCT of Harch et al 46 was a Level 1 study with a fair quality greater bias PEDro Scale score of 5 The fourth study 80 is a Level 3 study In addition non significant symptom reductions occurred in a single Level 2 excellent quality low bias study at 2 0 ATA oxygen 71 and significant symptom reductions in single Level 2 and Level 1 moderate and excellent quality low bias studies at 1 3 ATA pressure of air 69 and 1 2 ATA pressure of air 75 respectively Effect sizes for all of the Level 1 4 studies in the present review are at least moderate and in excess of placebo effects for the oxygen groups compared to compressed air or no treatment groups with symptom effect sizes greater than cognitive effect sizes 126 The results achieved with the 1 5 ATA oxygen dose are reinforced when cognitive outcomes are added from these 1 5 ATA oxygen symptom outcome studies and the Boussi Gross et al 70 1 5 ATA Level 2 oxygen study the case series study of Mozayeni et al 84 and the case reports of Wright et al 67 and Shytle et al 89 The symptom and cognitive outcomes are strongly supported by the functional imaging 17 127 changes in two controlled studies 70 80 Both studies revealed significant improvements in brain blood flow In Harch et al 80 the improvements in blood flow were almost exclusively in the white matter the primary site of injury in mTBI 9 10 15 22 24 30 32 35 47 but significant improvements in blood flow were also demonstrated in the hippocampi consistent with the improvements in memory function of the subjects Near identical findings improvements in memory and simultaneous increase in blood vessel density were demonstrated at 1 5 ATA with oxygen in the Harch et al 65 animal model of HBOTtreated mTBI In Boussi Gross et al 70 improvements in cognitive domains matched improvements in HBOT induced brain blood flow to the anatomic areas of injury responsible for cognitive deficits seen in the subjects In addition the increases and decreases in brain blood flow to different regions of the brain measured in Boussi Gross et al 70 were an independent alternative replication of the mathematical texture analysis and visual pattern shift of brain blood flow measured in Harch et al 80 The widespread improvements on SPECT seen in these two studies are contrary to those seen in SPECT studied placebo drug trials 128 129 and are consistent with the symptom and cognitive improvements seen in all of the studies at 1 5 ATA Equally important the beneficial outcomes with the 1 5 ATA dose were achieved with minor side effects or complications The outcomes achieved in the reviewed studies compare favorably with outcomes achieved in a non controlled U S Department of Defense National Intrepid Center of Excellence treatment program for veterans with TBI with or without psychological health problems DeGraba et al 130 reported a 7 5 year experience with 1 456 veterans who were a minimum Frontiers in Neurology www frontiersin org Summary of Main Findings According to the CEBM evidence classification hierarchy 3 the two randomized controlled trials of Miller et al 75 excellent quality low bias and Harch et al 46 fair quality greater bias meet the Level 1 standard for Evidence Based Medicine 18 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury treatment of mTBI PPCS Both of these studies demonstrated the benefit of the treatment groups at 1 5 ATA oxygen and one at 1 2 ATA air compared to control groups One of the studies 75 also demonstrated a positive effect on co morbid PTSD a finding first reported in 2009 66 and reinforced in multiple subsequent reports 68 69 71 80 81 87 89 The comparative dose Level 2 studies of Boussi Gross et al 70 fair quality greater bias and Weaver et al 81 good quality low bias the only study with mismatched treatment groups HBOT group with more serious brain injury demonstrated additional proof of the efficacy of the 1 5 ATA oxygen dose compared to a control group or 1 2 ATA air group along with the casecontrolled series of Harch et al 80 Level 3 and case series of Mozayeni et al 84 Levels 4 and the 3 case reports 66 67 89 Level 5 At the same time one Level 2 good quality low bias study Wolf et al 69 demonstrated efficacy at the 1 3 ATA air dose Compared to a previous Class B recommendation for HBOT in mTBI PPCS 100 the additional randomized controlled trials and randomized comparative dosing studies now meet CEBM Level 1 evidence and American Society of Plastic Surgeons Grade A Practice Recommendation criteria for HBOT treatment of mTBI PPCS 4 Tables 3 5 at 1 5 ATA oxygen a broad range Improvements were greater when patients had comorbid Post Traumatic Stress Disorder Large blinded controlled trials would be ideal to confirm the results of this review but despite small sample sizes the studies using 1 5 ATA oxygen satisfy the Centre for Evidence Based Medicine 3 Level 1 criteria and merit a Class A Recommendation for treatment of mTBI PPCS at 1 5 ATA of oxygen according to the American Society of Plastic Surgeons Grade Practice Recommendations 4 CLASSIFICATION OF EVIDENCE This review provides Centre for Evidence Based Medicine 3 Class 1 evidence that 40 hyperbaric oxygen treatments at 1 5 ATA oxygen are effective and well tolerated in mTBI Persistent PostConcussion Syndrome According to the American Society of Plastic Surgeons Grade Practice Recommendations this evidence meets the threshold for and is a Grade A Practice Recommendation 4 CONTRIBUTION TO THE FIELD STATEMENT Limitations The main limitation is the heterogeneity of the patients inherent in the diagnosis of mTBI PPCS causes of injury circumstances non uniformity of force vector head position anatomy symptoms past injuries co morbidities time from injury to treatment etc Additional limitations are the exclusion of non English language literature the discrepancy in treatment and testing sites with respect to altitude 81 small sample sizes out of proportion to the heterogeneity 46 68 71 75 80 81 84 variability in the design of the studies 46 68 71 75 80 81 84 heterogeneity of outcome instruments 46 68 71 75 80 81 84 failure to appreciate that a pressure experience is not a sham such that randomized controlled trials were actually randomized comparative dosing studies 69 71 75 81 difficulty in structuring a study with a true pressure control 46 69 71 75 81 lack of statistical analysis of outcome changes of one group compared to another or control 69 71 75 and paucity of literature to review Despite all of these limitations which would bias data and conclusions toward rejection of the null hypothesis and Type II Error due to small sample size the studies data support the conclusions drawn herein Mild TBI has been traditionally considered to be innocuous It is now appreciated to cause permanent symptoms the Persistent PostConcussion Syndrome PPCS in a substantial number of patients Hyperbaric oxygen therapy is a treatment for mostly acute and chronic wound conditions that is scientifically defined as a dual drug therapy that uses increased oxygen and increased pressure to treat disease pathophysiology wounds Multiple studies have reported confusing and seemingly conflicting results of HBOT treatment of PPCS due to the use of a non scientific historical definition of HBOT that led to inadvertent maldesign of some of the studies Using the scientific definition of HBOT this review resolves the confusion and conflict by analyzing the studies on HBOT in mTBI PPCS according to individual and composite doses of pressure and oxygen The results show that barometric pressure is the more important component of HBOT compared to the pressure of oxygen and that in multiple studies a composite dose of pressure and oxygen 1 5 ATA oxygen demonstrated significant improvement in symptoms and cognition in patients with PPCS This finding has the potential to change the standard of practice in the treatment of PPCS and has substantial implications for millions of patients worldwide afflicted with PPCS including military war veterans Conclusions In multiple randomized and randomized controlled studies HBOT at 1 5 ATA oxygen demonstrated statistically significant symptomatic and cognitive or cognitive improvements alone in patients with mild traumatic brain injury Persistent Postconcussion Syndrome Positive and negative results occurred at lower and higher doses of oxygen and pressure According to pressure and oxygen dosage analyses increased pressure within a narrow range appears to be the more important effect than increased oxygen which is effective over Frontiers in Neurology www frontiersin org DATA AVAILABILITY STATEMENT The original contributions presented in the study are included in the article Supplementary Material further inquiries can be directed to the corresponding author 19 March 2022 Volume 13 Article 815056

Harch Hyperbaric Oxygen Traumatic Brain Injury AUTHOR CONTRIBUTIONS ACKNOWLEDGMENTS The author confirms being the sole contributor of this work and has approved it for publication Thank you to all of the civilians and veterans who participated in these studies Ms Juliette Lucarini for critical review assistance and support on many levels in the past 17 years and to Liudmyla Osadcha for expert construction of the graphs in Figures 2 3 FUNDING SUPPLEMENTARY MATERIAL LSU School of Medicine paid academic protected time to partially fund the research and writing of this manuscript The remainder of the effort was provided by the sole author without funding The sole author paid all publication fees The Supplementary Material for this article can be found online at https www frontiersin org articles 10 3389 fneur 2022 815056 full supplementary material REFERENCES 17 Raji CA Henderson TA PET and single photon emission computed tomography in brain concussion Neuroimag Clin N Am 2018 28 67 82 doi 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Harch Hyperbaric Oxygen Traumatic Brain Injury 145 Artru F Chacornac R Deleuze R Hyperbaric oxygenation for severe head injuries Eur Neurol 1976 14 310 8 doi 10 1159 000114753 141 Rockswold SB Rockswold GL Vargo JM Erickson CA Sutton RL Bergman TA et al Effects of hyperbaric oxygenation therapy on cerebral metabolism and intracranial pressure in severely brain injured patients J Neurosurg 2001 94 403 11 doi 10 3171 jns 2001 94 3 0403 142 Rockswold SB Rockswold GL Zaun DA Zhang X Cerra CE Bergman TA et al A prospective randomized clinical trial to compare the effect of hyperbaric to normobaric hyperoxia on cerebral metabolism intracranial pressure and oxygen toxicity in severe traumatic brain injury J Neurosurg 2010 112 1080 94 doi 10 3171 2009 7 JNS09363 143 Rockswold SB Rockswold GL Zaun DA Liu J A prospective randomized Phase II clinical trial to evaluate the effect of combined hyperbaric and normobaric hyperoxia on cerebral metabolism intracranial pressure oxygen toxicity and clinical outcome in severe traumatic brain injury J Neurosurg 2013 118 1317 28 doi 10 3171 2013 2 JNS121468 144 Sahni T Jain M Prasad R Sogani SK Singh VP Use of hyperbaric oxygen in traumatic brain injury retrospective analysis of data of 20 patients treated at a tertiary care centre British J Neurosurg 2012 26 202 7 doi 10 3109 02688697 2011 6 26879 Frontiers in Neurology www frontiersin org Conflict of Interest PH declares a co owner of a hyperbaric service and consulting company Publisher s Note All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations or those of the publisher the editors and the reviewers Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher Copyright 2022 Harch This is an open access article distributed under the terms of the Creative Commons Attribution License CC BY The use distribution or reproduction in other forums is permitted provided the original author s and the copyright owner s are credited and that the original publication in this journal is cited in accordance with accepted academic practice No use distribution or reproduction is permitted which does not comply with these terms 24 March 2022 Volume 13 Article 815056

Hyperbaric Oxygen Therapy Can Improve Post Concussion Syndrome Years after Mild Traumatic Brain Injury Randomized Prospective Trial Rahav Boussi Gross1 Haim Golan3 4 Gregori Fishlev1 Yair Bechor1 Olga Volkov3 4 Jacob Bergan1 Mony Friedman1 Dan Hoofien6 7 Nathan Shlamkovitch8 Eshel Ben Jacob2 5 9 10 Shai Efrati1 2 3 10 1 The Institute of Hyperbaric Medicine Assaf Harofeh Medical Center Zerifin Israel 2 Research and Development Unit Assaf Harofeh Medical Center Zerifin Israel 3 Sackler School of Medicine Tel Aviv University Tel Aviv Israel 4 Nuclear Medicine institute Assaf Harofeh Medical Center Zerifin Israel 5 The Raymond and Beverly Sackler Faculty of Exact Sciences School of Physics and Astronomy Tel Aviv University Tel Aviv Israel 6 Department of Psychology The Hebrew University of Jerusalem Jerusalem Israel 7 The National Institute for the Rehabilitation of the Brain Injured Tel Aviv Israel 8 Otolaryngology Head Neck Surgery Assaf Harofeh Medical Center Zerifin Israel 9 Center for Theoretical Biological Physics Rice University Houston Texas United States of America 10 Sagol School of Neuroscience Tel Aviv University Tel Aviv Israel Abstract Background Traumatic brain injury TBI is the leading cause of death and disability in the US Approximately 70 90 of the TBI cases are classified as mild and up to 25 of them will not recover and suffer chronic neurocognitive impairments The main pathology in these cases involves diffuse brain injuries which are hard to detect by anatomical imaging yet noticeable in metabolic imaging The current study tested the effectiveness of Hyperbaric Oxygen Therapy HBOT in improving brain function and quality of life in mTBI patients suffering chronic neurocognitive impairments Methods and Findings The trial population included 56 mTBI patients 1 5 years after injury with prolonged postconcussion syndrome PCS The HBOT effect was evaluated by means of prospective randomized crossover controlled trial the patients were randomly assigned to treated or crossover groups Patients in the treated group were evaluated at baseline and following 40 HBOT sessions patients in the crossover group were evaluated three times at baseline following a 2 month control period of no treatment and following subsequent 2 months of 40 HBOT sessions The HBOT protocol included 40 treatment sessions 5 days week 60 minutes each with 100 oxygen at 1 5 ATA Mindstreams was used for cognitive evaluations quality of life QOL was evaluated by the EQ 5D and changes in brain activity were assessed by SPECT imaging Significant improvements were demonstrated in cognitive function and QOL in both groups following HBOT but no significant improvement was observed following the control period SPECT imaging revealed elevated brain activity in good agreement with the cognitive improvements Conclusions HBOT can induce neuroplasticity leading to repair of chronically impaired brain functions and improved quality of life in mTBI patients with prolonged PCS at late chronic stage Trial Registration ClinicalTrials gov NCT00715052 Citation Boussi Gross R Golan H Fishlev G Bechor Y Volkov O et al 2013 Hyperbaric Oxygen Therapy Can Improve Post Concussion Syndrome Years after Mild Traumatic Brain Injury Randomized Prospective Trial PLoS ONE 8 11 e79995 doi 10 1371 journal pone 0079995 Editor Jinglu Ai St Michael s Hospital University of Toronto Canada Received August 22 2013 Accepted October 4 2013 Published November 15 2013 Copyright 2013 Boussi Gross et al This is an open access article distributed under the terms of the Creative Commons Attribution License which permits unrestricted use distribution and reproduction in any medium provided the original author and source are credited Funding The study was supported by the research fund of Assaf Harofeh medical center by the Tauber Family Foundation and by the Maguy Glass Chair in Physics of Complex Systems at Tel Aviv University None of the supporting bodies had any role in study design data collection and analysis decision to publish or preparation of the manuscript Competing Interests The authors have declared that no competing interests exist E mail efratishai 013 net SE eshel rice edu EB J These authors contributed equally to this work dysfunction Intensive therapy and rehabilitation programs are considered essential for maximizing quality of life but are often just partially successful Clearly new methods for brain repair should be examined in order to provide sustained relief to brain damage patients Recent studies reported that hyperbaric oxygen treatment HBOT can induce neuroplasticity leading to significant neurological improvement in post stroke patients at the convalescent stage and at late chronic stages months to years after the acute event 3 4 Introduction Traumatic brain injury TBI and stroke are the major causes of brain damage Every year close to two million people in the US suffer TBI which is the leading cause of death and disability among the general population Stroke affects almost a million people and is the leading cause of inability to maintain independent life among adults 1 2 There is no effective treatment metabolic intervention in the daily clinical practice for post TBI and stroke patients with chronic neurological PLOS ONE www plosone org 1 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Definitions and classifications function in animal models 26 27 28 29 30 The elevated oxygen levels can have a significant effect on the brain metabolism largely regulated by the glial cells see discussion Improved energy management leads to multifaceted repair including activation of angiogenesis and triggering of neuroplasticity reactivation of quiescent neurons creation of new synapses and new axonal connections and might even induce differentiation of neuronal stem cells 22 The idea that HBOT can promote brain repair is reasonable and has gained experimental support yet is still largely dismissed by the medical community as is discussed next Traumatic brain injury is defined as damage to the brain resulting from external mechanical force such as rapid acceleration or deceleration impact blast waves or penetration by a projectile Consequently to the injury brain function is temporarily or permanently impaired and structural damage may or may not be detectable with current imaging technology TBI is usually classified based on severity anatomical features of the injury and the cause of the injury The severity is assessed according to the loss of consciousness LOC duration the post traumatic amnesia PTA and the Glasgow coma scale GCS grading of the level of consciousness Approximately 70 90 of the TBI in the US are classified as mild TBI mTBI or concussion LOC duration of 0 30 minutes PTA duration of less than a day and GCS grade of 13 15 Post concussion syndrome PCS is a set of symptoms succeeding mTBI in most patients The PCS symptoms include headache dizziness neuropsychiatric symptoms and cognitive impairments 5 6 In most patients PCS may continue for weeks or months and up to 25 of the patients may experience prolonged PCS PPCS in which the symptoms last for over six months 7 8 9 10 11 12 Such individuals are at high risk for emotional and cognitive dysfunction culminating in inability to carry out ordinary daily activities work responsibilities and standard social relationships 9 10 11 12 The medical community reservations A study of the effect of hyperbaric oxygen treatment of severe brain injured patients has been published already two decades ago Several prospective clinical trials on treatment of mTBI have been published in the last decade 31 32 33 and three studies published in the last two years addressed the effect of HBOT on chronic mild TBI patients 34 35 36 However the reported beneficial effects of the hyperbaric treatment were severely questioned by the medical community and triggered high skepticisms to the extent that TBI and stroke patients in the US are rarely treated by hyperbaric oxygen The HBOT option has been dismissed by the medical community on the grounds of 1 Lack of knowledge about the connection between metabolism and neuroplasticity 2 Lack of randomized clinical trial with standard placebo control 3 Sham control with room air at 1 3Atm yielded significant improvements These issues are clarified and elaborated on in the discussion section Associated brain pathology and function impairments Diffuse axonal injury diffuse shearing of axonal pathways and small blood vessels is one of the most common pathological feature associated with mTBI 13 Another primary pathological feature usually caused by a direct hit to the skull is brain contusions which commonly involve the frontal and anterior temporal lobes 12 Secondary pathologies of mTBI include ischemia mild edema and other bio chemical and inflammatory processes culminating in impaired regenerative healing processes resulted from increasing tissue hypoxia 14 Due to the diffuse nature of injury cognitive impairments are usually the predominant symptoms involving deficiencies in several cognitive functions primarily memory attention processing speed and executive functions all localized in multiple brain areas Their potent functions rely on potent network structure and connectivity between different brain areas 12 15 16 We note that the diffuse nature of the mTBI injury renders the pathological damage hard to be detected by common neuroimaging methods such as CT and MRI so that diagnosis largely relies on subjective reports of the patients as well as cognitive and quality of life tests While diffusion tensor imaging DTI has the potential to detect diffuse axonal injuries this method is still not commonly used for diagnosis of mTBI pathology The placebo dilemma People can sense a pressure increase beyond 1 3Atm hence standard placebo with normal air pressure for HBOT could perhaps be attained by exposing the patients to normal pressure combined with falsifying stimulation e g by increasing and decreasing the pressure which generates a fictitious pressure sensation Since breathing normal air under hyperbaric conditions leads to elevated tissue oxygen e g about 50 for 1 3Atm standard placebo could also be attained by giving the patients compressed air with sub normal oxygen concentration In the discussion section we explain that the first approach can be effective only for some patients and poses logistic difficulties and the second approach involves ethical issues In an attempt to evade the placebo dilemma a recent study of HBOT for mTBI compared the effect of 100 oxygen at 2 4Atm with the effect of room air at 1 3Atm as sham control 36 The study found significant improvements in both groups and with slightly higher efficacy at 1 3Atm Based on these results the authors resented a sweeping conclusion that their study shows that HBOT has no effect on post mTBI brain damage and the observed improvements resulted from placebo associated with spending time in the hyperbaric chamber As is discussed in great details in the discussion section we reason that the authors reached wrong conclusions for two main reasons First room air at 1 3Atm cannot serve as a proper sham control since it is not an ineffectual treatment as is required from placebo since it leads to a significant increase in the level of tissue oxygenation which has been shown to be effective 37 38 Second 100 oxygen at 2 4Atm leads to too high oxygen levels which can cause inhibitory effect or even focal toxicity Rationale for hyperbaric oxygen treatment HBOT The brain receives 15 of the cardiac output consumes 20 of the total body oxygen and utilizes 25 of the total body glucose Still this energy supply is only sufficient to keep about five to ten percent of the neurons active at any given time Thus at standard healthy condition at any given time the brain is utilizing almost all oxygen energy delivered to it The regeneration process after brain injury requires much additional energy This is where hyperbaric oxygen treatment can help the increased oxygen level in the blood and body tissues during treatment 17 18 19 can supply the energy needed for brain repair Indeed several previous studies have demonstrated that elevated levels of dissolved oxygen by HBOT can have several reparative effects on damaged brain tissues 3 19 20 21 22 23 24 25 Other studies revealed the beneficial effect of HBOT on the injured brain and cognitive PLOS ONE www plosone org The crossover approach To overcome the placebo issue a randomized crossover approach was successfully used to test the effect of HBOT in post stroke patients at late chronic stage 3 The advantage of the 2 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Evaluation of cognitive state crossover approach is the triple comparison between treatments of two groups between treatment and no treatment of the same group and between treatment and no treatment in different groups Up till now a similar prospective randomized crossover trial to evaluate the brain repair effect of HBOT in mTBI patients at late chronic stage has not been done The aim of our current study was to provide firm evaluation of the HBOT effects on brain activity and cognitive impairments in mTBI patients with prolonged PCS at late chronic stage Cognitive Indices The state of the patients cognitive functions was assessed in terms of the following four cognitive indices ordered from the index associated with most fundamental basic functions to that associated with the higher functions 1 Information Processing Speed IPS index This index is associated with the basic ability to process and respond to stimuli at different levels of speed and complexity 2 Attention related index This index is associated primarily with the ability to remain concentrated and respond effectively throughout relatively extended periods of time 3 Memory related index This index is associated with the learning of verbal and visual new stimuli and the immediate and delayed recognition of these learned stimuli 4 Executive Functions EF index This index is associated with cognitive abilities involved in the initiation planning organization and regulation of behavior Each of above cognitive indices was computed as a normalized combined score of 2 3 cognitive tests from the Mindstreams Computerized Cognitive Test Battery Mindstreams NeuroTrax Corp NY Cognitive tests The Mindstreams battery includes several cognitive tests devised to check various aspects of brain capabilities In the current study we evaluated the cognitive indices based on the scores of the 6 cognitive tests listed below which are expected to be relevant for mild TBI For detailed description of all cognitive tests in Mindstreams battery see 39 The tests are 1 Verbal memory Ten pairs of words are presented followed by a recognition test in which the first word of a previously presented pair appears together with a list of four words from which the patients choose the other member of the pair There are four immediate repetitions and one delayed repetition after 10 min 2 Non verbal memory Eight pictures of simple geometric objects are presented followed by a recognition test in which four versions of each object are presented each oriented in a different direction There are four immediate repetitions and one delayed repetition after 10 min 3 Go No Go test In this continuous performance test a colored square red green white or blue appears randomly on the center of the screen The patient in then asked to respond quickly only for red squares by pressing the mouse button and inhibit his reaction to any other colored square 4 Stroop test Timed test of response inhibition modified from the Stroop paper based test In the first phase patients choose a colored square matching the color of a general word for example the word Cat appears in red letters the patient must choose the red square out of two colored squares in the following screen In the next phase termed the Choice Reaction Time test the task is to choose the colored square matching the name of the color presented in white letter color In the final Stroop interference phase patients are asked to choose the colored square matching the color and not the meaning of a former colornaming word presented in an incongruent color for example the word RED appears in green letters the patient is asked to choose the color green and not red a task requiring the ability to inhibit an automatic response to the meaning of the word 5 Staged information processing test Timed test requiring a reaction based on solving simple arithmetic problems pressing right left mouse button if the answer higher lower than 4 respectively with three levels of information processing load single digit two digits addition subtraction and three digits addition subtraction problems each containing three speed levels 3 2 and 1 second for the presentation of the stimuli Methods The study was performed as a prospective randomized controlled two group trial The study was conducted in the hyperbaric institute and the research unit of Assaf Harofeh Medical Center Israel Enrolment of patients started at 2008 and ended at 2012 All patients signed written informed consent The protocol was approved by Assaf Harofeh institutional review board Participants Inclusion The participants were patients of age 18 years or older who suffered mild TBI less than 30 minutes loss of consciousness 1 6 years prior to their inclusion All patients experienced post concussion syndrome PCS and complained of impaired cognitive functions for over a year yet brain damage was below the detection level of MRI or CT brain imaging Only patients who reported no change in cognitive function during one month prior to the beginning of the study were included Exclusions Exclusions were due to chest pathology incompatible with HBOT inner ear disease claustrophobia and inability to sign informed consent Smoking was not allowed during the study Protocol and End Points After signing an informed consent form the patients were invited for baseline evaluation Included patients were randomized into two groups 1 1 randomization a treated group and a crossover group The neuropsychological functions evaluated by Mindstreams testing battery and brain activity as visualized by SPECT Single photon emission computed tomography were the primary endpoints of the study Secondary end point included quality of life evaluation by the EQ 5D questionnaire Evaluations were made by medical and neuropsychological practitioners who were blinded to patients inclusion in the control crossed or the treated groups Patients in the treated group were evaluated twice at baseline and after 2 months of HBOT Patients in the crossover group were evaluated three times baseline after 2 months control period of no treatment and after subsequent 2 months of HBOT Figure 1 The post HBOT neurological evaluations as well as the SPECT scans were performed more than 1 week 1 3 weeks after the end of the HBOT protocol The following HBOT protocol was practiced 40 daily sessions 5 days week 60 minutes each 100 oxygen at 1 5ATA Patients were not involved in any other cognitive or rehabilitation intervention as part of the study protocol The detailed clinical study protocol copy of the informed consent as well as CONSORT 2010 checklist of information are attached as supporting information Protocol S1 Form S1 Checklist S1 We note that information regarding sample size detectable change and power calculation parameters is included and addressed in the statistical considerations section in the SI1 PLOS ONE www plosone org 3 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 1 Flow chart of the patients in the study doi 10 1371 journal pone 0079995 g001 6 Catch game A test of motor planning that requires participants to catch a falling object on a computer screen by moving a paddle horizontally so that it can catch the falling object To assign scores Mindstreams data was uploaded to the NeuroTrax central server Outcome parameters were calculated using custom software blind to diagnosis or testing site To minimize differences related to age and education each outcome parameter was normalized and fit to an IQ like scale mean 100 STD 15 according to patient s age and education We note that the score evaluation was based on normative data from cognitively healthy individuals collected in controlled research studies that were part of more than 10 clinical sites 40 The cognitive indices scores The computation of the cognitive indices based on the scores of the cognitive tests was done as follows 1 Information Processing Speed index was computed as the combined score for the low and medium load stages of the staged information processing test 2 Attention index was calculated as the mean score of reaction time for Go No Go test and choice reaction time of the Stroop test at second phase mean STD of reaction time for Go No Go test mean reaction time for a low load stage of staged information processing test and mean accuracy for a medium load stage of information processing test 3 Memory index was computed as the mean score for total learning score after four repetitions and delayed recognition phase of verbal and non verbal memory tests 4 Executive PLOS ONE www plosone org Functions index was computed based on the scores of the Stroop test and the Go No Go test and the mean weighted accuracy for catch game For more information regarding the validity of the tests and the construction of the cognitive indices see 41 42 5 In addition we defined the individual s General Cognitive Score as the average of the scores of the four cognitive indices for each individual It is important to note that the above cognitive index scores were specifically designed to represent known impaired cognitive domains in mild TBI In addition the fact that each index is referred to more than one test score ensures the index to be associated more with a cognitive domain score and less with a testdependent score We also utilized a computerized testing battery which supports the inclusion of more accurate measures such as reaction time and elimination of the bias effect of tests administration and hand scoring An important aspect of the tests is the inclusion of the cognitive domain of information processing speed known to be impaired in mild TBI patients Quality of life evaluation Quality of life QOL was evaluated by the EQ 5D questionnaire 43 EQ 5D essentially consists of 2 pages the EQ 5D descriptive system and the EQ visual analogue scale EQ VAS The EQ 5D descriptive system covers mobility self care usual activities pain discomfort and anxiety depression The EQ VAS 4 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI records the respondent s self rated health on a vertical visual analogue scale range 0 worst 100 best Results Brain Functional Imaging SPECT The study included 90 screened patients aged 18 years or older who signed an informed consent Pre study exclusions Nineteen patients had their consent withdrawn before the beginning of the control treatment period 13 in the crossover group 6 in the treated group In study exclusions Four patients decided to drop out during the treatment protocol 3 due to personal reasons and 1 due to ear problem 1 in crossover group 3 in treatment group Seven patients 5 in crossover group 2 in treatment group were excluded due to technical performance problems in their cognitive tests and 4 patients due to inconsistent use of medications such us methylphenidate during the tests period 2 in crossover group 2 in treated group Accordingly 56 patients 32 in the treated group and 24 in crossover group were included in the final analysis Figure 1 Thirty two 57 patients were females the mean age was 44 years range of 21 66 years and the time elapsed since the acute traumatic event ranged from 1 6 years with 33 months average The most frequent etiology of the TBI was a vehicle accident n 38 with some other less common etiologies falls 7 object hit 6 pedestrian accident 3 assault 2 Baseline patients characteristics are summarized in Table 1 As seen from this table there was no significant difference in the included measures between the groups except for years of education where there was a minor advantage for the treated group Participants Profiles Brain single photon emission computed tomography SPECT was conducted with 925 1 110 MBq 25 30 mCi of technetium99m methyl cysteinate dimmer Tc 99m ECD at 40 60 min post injection using a dual detector gamma camera ECAM or Symbia T Siemens Medical Systems equipped with high resolution collimators Data was acquired in 3 degree steps and reconstructed iteratively with Chang method m 0 12 cm attenuation correction 44 Visual analysis was conducted by fusing pre and post treatment studies that were normalized to cerebellum brain activity SPECT images were reoriented into Talairach space using NeuroGam Segami Corporation for identification based on visual inspection of Brodmann cortical areas and in order to compute the mean perfusion in each Brodmann area BA In addition volume rendered brain perfusion images normalized to cerebellum maximal activity were reconstructed All SPECT analysis was done while blinded to the laboratory and clinical data Change in perfusion in all Brodmann areas for each subject was determined by calculating the percentage difference between post period and pre baseline period divided by the pre baseline period perfusion An average of these perfusion changes for each Brodmann area was calculated Statistical analysis The statistical analysis was done using SPSS software version 16 0 Continuous data is expressed as means 6 standard deviations and compared by one tailed paired t test for intragroup comparisons and two tailed unpaired t test for inter group comparisons Effect sizes for main comparisons were calculated using Cohen s d Categorical data is expressed in numbers and percentages and compared by x2 test P values 0 05 were considered statistically significant All randomly allocated patients were included in the safety analysis and those with complete postbaseline assessment were included in efficacy analysis The Effect on Cognitive Functions Changes in cognitive indices The effect of the hyperbaric oxygen treatment on the patients cognitive functions as assessed by the four cognitive indices is summarized in Figure 2 and Table 2 The baseline mean cognitive scores of all four indices were close in the two groups within the standard error but with somewhat higher values in the treated group The HBOT treatments of both groups led to statistically significant improvements in the mean scores of all four indices As is apparent in Figure 2 and detailed in Table 2 a significant improvement was observed in the treated group after HBOT in all cognitive measures Information Processing Speed t 31 4 20 p 0 0001 Attention t 31 3 26 p 0 005 Memory t 31 4 13 p 0 0005 and Executive Functions t 31 3 72 p 0 0005 Effect sizes were medium to large the Cohen s d measures 45 were 0 74 0 57 0 73 and 0 66 respectively No significant improvement was noticed in the crossover group during the control period Information Processing Speed t 23 0 53 p 0 298 Attention t 23 0 33 p 0 368 Memory t 23 0 74 p 0 233 and Executive Functions t 23 0 54 p 0 295 However a significant improvement following HBOT was noticed in the crossover group as well Information Processing Speed t 23 1 98 p 0 05 Attention t 23 2 29 p 0 05 Memory t 23 3 21 p 0 005 and Executive Functions t 23 2 26 p 0 05 Effect sizes were medium to large with Cohen s d measures of 0 40 0 47 0 65 and 0 46 respectively Note that t 31 and t 23 correspond to N 1 where N 32 and N 24 are the number of patients in the treated and crossover group respectively Assessment of a general cognitive score The changes in the four cognitive indices presented in Figure 2 and in Table 2 show noticeable variability For example the mean values of the Information Processing Speed and Executive Functions indices decreased during the control period of the crossover group while the corresponding mean values of the Attention and Memory Scatter plot analysis of the clinical scores The analysis aims to better quantify and compare changes in the clinical scores while taking into consideration the high patient topatient variability following Efrati et al 3 The idea was to inspect for each patient at each time stage the scaled relative differences in each of the clinical scores More specifically we calculated for a specific patient j the scaled relative difference SRDj defined as STD SFj SRDj SFj vSFj w SIj vSIj w STD SIj z vSFj SIj w 1 Where SFj is the value of a clinical score at the end of the time stage either treatment or control and SIj is the score at the beginning of the time stage We note that the symbol indicates average over the values of the patients in the group For example SFj means the average of SFj over all patients j that belong to the group The abbreviation STD means the standard deviation between the values of the patients in the group This analysis enables quantitative inspection of the changes in the clinical scores as is further explained in 3 PLOS ONE www plosone org 5 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Table 1 Baseline patients characteristics Treated group n 32 Crossover group n 24 Comparison Age years 42 5612 6 45 7610 9 p 0 32 Gender male 11 34 13 54 p 0 07 Years of education 16 263 9 14 063 1 p 0 05 Time since injury months 34 6616 7 31 7616 3 p 0 51 None 24 75 14 58 30 minutes 8 25 10 42 Loss of consciousness p 0 18 Etiology Vehicle accident 20 63 18 75 Fall 5 16 2 8 Object hit 4 12 2 8 Pedestrian accident 2 6 1 4 Assault 1 3 1 4 Hypertension HTN 5 15 4 16 Diabetes Mellitus DM 2 6 2 8 Hyperlipidemia 4 12 3 12 Ischemic Heart Disease 0 1 4 Epileptic seizure 0 0 Smoking 1 3 0 Aspirin 2 6 3 12 Glucose lowering drugs 2 6 1 4 Anti HTN 4 12 3 12 Background disease Medications Statins 3 9 3 12 Anti depressant 7 22 4 16 doi 10 1371 journal pone 0079995 t001 indices increased Figure 3 shows the mean values of the individual general cognitive scores with standard error for the treated and crossover groups at each evaluation stage baseline and postHBOT for both groups and after the control for the crossover group It can be seen that the cross group had the same general score at baseline and after the control period This value seems higher than the score of the treated group at baseline 88 vs 85 and the post HBOT general cognitive score of the treated group seems higher than that of the crossover group 96 vs 94 While these differences are within the standard error they still give rise to what appears to be significant differences in the level of changes post vs pre HBOT between the crossover and the treated groups 6 points for the crossover group vs 11 for the treated group Examining the relative changes There is a high patient topatient variability in the cognitive indices with scores ranging from 20 to 120 The magnitude of the change in a cognitive score has different implications for patients at low or high base levels Hence we inspect the effect of the HBOT on the relative changes i e the change relative to the base value We calculated for each person the relative change in each of the cognitive indices for each period control and HBOT for the crossover group and HBOT for the treated group In Figure 4A we show the mean relative changes in all four cognitive indices for the crossover group following the control period and following HBOT and for the treated group following HBOT In Figure 4B we show the mean PLOS ONE www plosone org relative changes in the general cognitive score for the same three periods We note that calculating the mean of the relative changes is more informative than calculating the changes in the mean values especially for small groups with high patient to patient variability Looking at the relative changes elucidates the improvements after the HBOT period vs the control period of the crossover group However it also amplifies the differences mentioned earlier between the crossover and treated groups the bigger relative changes in the treated group vs the crossover group reflect the fact that the baseline values of the treated group were lower and the post HBOT values were higher in comparison to the corresponding values of the crossover group Scatter plot analysis of the cognitive indices As mentioned in the methods section the analysis aims to present the mean relative changes in cognitive indices while superimposing information regarding the patient to patient variability For that we calculated for each patient i the normalized relative change NRC i Next we calculated for each group control and HBOT in the crossover group and HBOT in the treated group the locations of the mean departures from baseline Finally we marked the location of each patient i at a distance NRC i from the location of his her group s mean difference see methods section for details Typical results are shown in Figure 5 More specifically we show the scatter plots for Information Processing Speed vs Executive Functions Figure 5A for Attention vs Memory 6 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 2 Assessment of the cognitive indices Each patient in each group was assigned a score at baseline after the control period for patients in the crossover group and after HBOT The figures show the mean scores and standard errors for the two groups at each stage for the four cognitive indices Information Processing Speed A Attention B Memory C and Executive Functions D as defined in the method section doi 10 1371 journal pone 0079995 g002 Attention EF Interestingly the scattering of the individual patients also follows the linear line for the scatter plots of the specific cognitive indices as function of the general cognitive score Figures 5C and 5D These linear dependences demonstrate high consistency between the changes of the different cognitive indices for each patient As such the analysis provides valuable test for the validity of the test performances and the validity of the general cognitive score Figure 5B for Attention vs General cognitive score Figure 5C and for IPS vs General cognitive score Figure 5D The results illustrate the differences between the three cases control and HBOT for the crossover group and HBOT for the treated group which form three distinct clusters Also clear in all the figures is the linear dependence between the changes in the different cognitive indices similar results are also obtained for the other combinations e g Memory IPS Attention IPS Memory EF and Table 2 Summary of results for Mindstreams cognitive indices scores Treated group n 32 Baseline HBOT Crossover group n 24 P1 P2 Baseline Control Pre HBOT Post HBOT P2 P3 P4 Memory 82 43625 15 96 54617 18 0 567 0005 85 90617 80 88 36617 34 95 61615 54 0 233 0 005 0 835 Executive function 88 26614 74 96 96611 69 0 367 0 0005 91 73613 26 90 20615 77 95 13613 84 0 295 0 05 0 595 Attention 85 13620 28 95 30612 90 0 854 0 005 86 10618 42 87 05620 98 92 02618 95 0 368 0 05 0 443 Information processing speed 85 12615 88 95 04613 75 0 324 0 0001 89 74618 81 88 30619 68 92 47618 25 0 298 0 05 0 55 Abbreviations Values are presented as mean 6 STD P1 stands for the p values for baseline comparison of treated and crossover group P2 stands for the p values for comparison of the second measurement to baseline in the same group P3 stands for the p values for comparison of pre and post HBOT in the crossover group P4 stands for the p values for endpoint scores comparison following treatment in both groups The baseline mean cognitive scores of all four indices were close in the two groups with no significant difference The HBOT treatments of both groups led to statistically significant improvements in the mean scores of all four indices as oppose to no significant improvement after control period alone The tables are discussed in details in the results section doi 10 1371 journal pone 0079995 t002 PLOS ONE www plosone org 7 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 3 Assessments of the general cognitive score The figure shows the level of the general cognitive score defined in the text for the crossover group at baseline after the control period and after HBOT and for the treated group at baseline and after HBOT doi 10 1371 journal pone 0079995 g003 The Effect on quality of life evaluation and evaluator the cognitive function tests were done by a computerized validated method and the SPECT analysis was blind to patients participation in treated crossover group Brain SPECT evaluations were completed for 31 patients in the treated group one patient from the treated group did not complete two SPECT scans and for 24 in the crossover group In supporting information Table S1 we present detailed statistics of the SPECT results for all Brodmann areas BA of all the tested patients The results revealed a significant increase in brain activity perfusion following the hyperbaric oxygen treatments in both groups compared to the change during the control period of the crossover group A summary of the results is presented in Figure 7 To construct the figure we calculated for each patient the relative change in the SPECT measured brain activity during each phase of the trial The relative change was defined as the difference in normalized activity relative to the cerebellum activity see SI4 between the end point and the start point divided by the activity at the start We calculated for each BA the average changes over all patients that underwent HBOT both treated and cross groups These results correspond to the red curve in Figure 7 The blue curve represents the results of similar calculations for the control period averaging for each BA over all the results of all patients in the crossover group during the control period The results revealed significant improvement in brain perfusion following HBOT in the frontal and temporal areas including inferior frontal gyrus BA 45 the middle frontal area BA 46 parts of the orbito frontal cortex BA 47 and 11 most of the temporal cortex BA 36 37 38 20 21 22 28 and parts of the Cingulate gyrus BA 24 25 These fronto temporal regions responsible for the cognitive functions are the most affected in The effect on the QOL is summarized in Table 3 The EQ 5D score significantly improved following HBOT in the treated group t 31 7 41 p 0 0001 and in the crossover group after HBOT t 23 6 17 p 0 0001 As expected there was no improvement in the EQ 5D score in the crossover group following the control period During the control period we have noticed some reduction in this group with respect to the patients subjective perception of their quality of life t 23 2 60 p 0 01 Similar results were obtained for the EQ VAS evaluations as summarized in Table 3 More specifically the EQ VAS score significantly improved following HBOT both in the treated group t 31 4 86 p 0 0001 and in the crossover group following treatment t 23 4 79 p 0 0001 while there was no significant improvement following the control period t 23 0 32 p 0 373 Details are presented in Table 3 Examining the relative changes Figure 6 presents the mean relative changes and standard errors in both measurements of quality of life for the treated group following HBOT and for the crossover group after the control period and after HBOT SPECT assessments of brain activity Motivation Since mTBI involves a diffuse structural and or physiologic metabolic derangement 46 47 48 patients with mTBI have more frequent and more extensive areas of brain damage than can be seen by anatomical imaging conventional CT and MRI scans The preferred brain imaging methods are thus functional metabolic SPECT PET CT perfusion and functional MRI In order to achieve greater validity of the results cognitive function and SPECT analysis were done by a blinded PLOS ONE www plosone org 8 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 4 Assessment of the relative changes A The relative changes as defined in the text for the four cognitive indices The changes are shown for the crossover group following the control period green bars and HBOT blue bars and for the treated group following HBOT red bars B Relative changes in the general cognitive score for the same three cases as in A doi 10 1371 journal pone 0079995 g004 Example 2 SPECT analysis of a 46 year old woman from the treated group following mTBI as a result of a car accident 1 year prior to inclusion in the study The patient s main complaints were related to her memory and concentration capabilities as well as dizziness and tinnitus that interfered with her daily functioning Following HBOT the dizziness and tinnitus disappeared in addition to significant improvement in all cognitive domains The patient s cognitive indices demonstrated significant improvements following treatment increase of 1 5 STD in Memory 57 preHBOT to 78 post HBOT 1 STD increase in Attention 88 to 104 1 5 STD increase in Executive Functions 82 to 102 and 0 6 STD increase in Information Processing Speed 85 to 95 SPECT images of the patient at baseline and following HBOT are shown in Figure 9 The percentage of CBF change demonstrated significant improvements after HBOT in parts of the frontal and temporal lobes in agreement with the improvements in neurological functions Example 3 SPECT analysis of a 44 year old man from the cross group suffering from cognitive impairments due to mild TBI as a result of a fall 2 years prior to treatments The patient complained mainly on short and long term memory difficulties attention and concentration problems decline in naming abilities as well as headaches dizziness anxiety and sleep problems Following HBOT the patient experienced significant improvements in most aspects including concentration and memory headaches dizziness mood and sleep The patient s cognitive indices demonstrated significant improvements in Executive Functions after treatment baseline 60 after control 63 and postHBOT 74 SPECT images of the patient after the control period head trauma 12 The temporal lobe plays a significant role in memory and learning skills 49 50 51 the frontal lobe is associated with attention and executive functions 52 53 and the cingulate gyrus plays an important role in emotions and cognitive self regulation 54 55 56 The SPECT results of elevated activity in these Brodmann areas are consistent with the improvement in cognitive indices following HBOT Further below we show specific examples of SPECT results for four patients two from the treated group and two from the crossover group Representative examples Example 1 SPECT analysis of a 51 year old woman from the treated group Figure 8 The patient suffered from cognitive impairments due to mTBI as a result of a fall in a moving bus 2 years prior to inclusion in the study The patient experienced no loss of consciousness and CT imaging did not reveal anatomic damage The patient s main complaints included headaches dizziness memory and concentration problems and random mood swings The patient was a manager in a private business and since the injury was having difficulties following and completing daily activities and routine Following HBOT the patient reported significant improvement in every aspect of daily functioning The patient s cognitive indices demonstrated significant improvements following treatment increase of 3 5 STD in Memory 56 pre HBOT to 108 post HBOT 2 STD increase in Attention 47 to 81 1 5 STD increase in Executive Functions 65 to 85 and a 0 7 STD increase in Information Processing Speed 85 to 95 PLOS ONE www plosone org 9 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 5 Scatter plot analysis of the changes in cognitive indices The scatter plots show the normalized relative changes NRC as defined in the methods section and explained in the text above A Scatter plot for the changes in IPS as function of changes in EF B Scatter plot for changes in Attention as function of Memory The changes in the Attention and in the IPS as function of the General cognitive score are shown in C and D respectively Circles are for the treated group and diamonds are for the crossover group The color code is Red for changes during HBOT and blue for changes during control doi 10 1371 journal pone 0079995 g005 following treatment in comparison to the changes during the control period Executive Functions at baseline after control and post HBOT were 77 71 and 88 respectively and Attention scores were 62 64 and 78 respectively SPECT images of the patient after the control period and following HBOT are shown in Figure 11 At first global look the CBF changes after HBOT seem to be similar to the changes after the control period demonstrating no significant change However a closer inspection of the SPECT images reveal local improved perfusion in the right anterior temporal and right dorso lateral frontal areas This example is shown to demonstrate that even for patients with relatively mild cognitive improvements there is good correspondence between the change in the cognitive tests and the SPECT results and following HBOT are shown in Figure 10 The CBF change was not significant after the control period but substantial improvement after HBOT in most of bilateral frontal and temporal lobes and right parietal lobe Example 4 SPECT analysis of a 39 year old woman from the cross group suffering from cognitive impairments due to mild TBI as a result of a car hit as a pedestrian 2 years prior to treatments The patient s complaints included dizziness nausea fatigue and decrease in memory and concentration abilities The patient also had troubles in performing every day activities such as attaining the grocery store or reading the newspaper The patient was working as a nurse and could not continue to perform her work after the accident due to the impaired abilities The patient s cognitive indices demonstrated significant improvements Table 3 Summary of results of quality of life questionnaire EQ 5D and EQ VAS Treated group n 32 Baseline HBOT Crossover group n 24 P1 P2 Baseline Control Pre HBOT Post HBOT P2 P3 P4 EQ 5D 7 8761 36 6 4861 07 0 615 0 0001 7 7061 11 8 0661 05 6 7561 06 0 01 0 0001 0 362 EQ VAS 5 0362 31 6 6262 45 0 696 0 0001 5 2661 70 5 2161 66 6 3961 80 0 373 0 0001 0 696 Abbreviations Values are presented as mean 6 STD P1 stands for the p values for baseline comparison of treated and crossover group P2 stands for the p values for comparison of the second measurement to baseline in the same group P3 stands for the p values for comparison of pre and post HBOT in the crossover group P4 stands for the p values for endpoint scores comparison following treatment in both groups EQ 5D as well as the EQ VAS scores significantly improved following HBOT both in the treated group and in the crossover group following treatment while there was no significant improvement following the control period doi 10 1371 journal pone 0079995 t003 PLOS ONE www plosone org 10 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 6 Assessments of the mean relative changes and standard errors in quality of life measurements The changes are shown for the crossover group following control period green bars and following HBOT blue bars and for the treated group following HBOT red bars Note that according to the questionnaire structure in the EQ 5D measurement improvement is reflected as score decrease hence the negative values of change doi 10 1371 journal pone 0079995 g006 Discussion 3 imply that increasing the plasma oxygen concentration with hyperbaric oxygenation is a potent means of delivering to the brain sufficient oxygen for tissue repair HBOT might initiate a cellular and vascular repair mechanism and improve cerebral vascular flow 34 58 59 60 More specifically HBOT induces regeneration of axonal white matter 61 62 63 64 has positive effect upon the myelinization and maturation of injured neural fibers 65 and can stimulate axonal growth and increase the ability of neurons to function and communicate with each other 66 In addition HBOT was found to have a role in initiation and or facilitation of angiogenesis and cell proliferation processes needed for axonal regeneration 67 At the cellular level HBOT can improve cellular metabolism reduce apoptosis alleviate oxidative stress and increase levels of neurotrophins and nitric oxide through enhancement of mitochondrial function in both neurons and glial cells Moreover the effects of HBOT on neurons can be mediated indirectly by glial cells including astrocytes 23 HBOT may promote the neurogenesis of endogenous neural stem cells 24 With regard to secondary injury mechanisms in mTBI HBOT can initiate vascular repair mechanism and improve cerebral vascular flow 58 59 68 69 promote blood brain barrier integrity and reduce inflammatory reactions 28 as well as brain edema 20 21 22 26 34 70 A drawback to the above mentioned findings is that the different effects have been tested at different experimental setups and while utilizing different protocols of HBOT However it is well noticed that there is at least one common denominator to all repair regeneration mechanisms We presented a prospective randomized and controlled cross over study of the effect of HBOT with 100 oxygen at 1 5Atm on mTBI patients at late chronic stage The results clearly demonstrate that HBOT can induce neuroplasticity and significant brain function improvement in mild TBI patients with prolonged PostConcussion Syndrome at late chronic stage years after brain injury Additional statistical validation using simulated randomizations is available as supporting information Text S1 Linking elevated oxygen metabolism and brain activity to neuroplasticity The changes in SPECT images after treatment indicate that HBOT led to reactivation of neuronal activity in stunned areas that seemed normal under CT and MRI imaging While SPECT imaging has a limited spatial resolution compared for example to fMRI the changes in activity were sufficiently robust to be clearly detected by the SPECT images Recently Kan et al 57 discussed the need for potent interventions such as elevated tissue oxygen capable of repairing microenvironment alterations after mTBI e g impairments in vascular changes in cerebral blood flow and in perfusion leading to reduced oxygen availability followed by reduced metabolism which in turn leads to reduced neuronal activity loss of synapses and tampered neuronal connectivity The observed reactivation of neuronal activity in the stunned areas found here along with similar results in post stroke patients PLOS ONE www plosone org 11 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 7 Distribution of the Brodmann areas relative SPECT CBF changes The change for each BA represents and averaging of the relative changes of all the patients as explained in the text The results show a clear difference between the control and the HBOT periods We note that the higher variations for the control period are associated with the fact that the averaging in this case is over 24 patients the crossover group while for the HBOT period the averaging is over all 55 patients doi 10 1371 journal pone 0079995 g007 mainly to the cognitive dysfunction following the injury are commonly based on behavioral compensation methods such as attention training drills teaching memory planning strategies and usage of external aids 16 76 and have limited patient specific success in repair of mTBI impaired brain function 77 Therefore studies at the late chronic stage allow assessment of the power of the HBOT approach to achieve brain function improvements in addition to rather than instead of the standard rehabilitation programs they are all energy oxygen dependent It might be possible that HBOT enables the metabolic change simply by supplying the missing energy oxygen needed for those regeneration processes Rationale for testing the HBOT effect on patients at late chronic stage As stated in the introduction the crossover approach is adopted in order to avoid the inherent difficulties associated with randomized HBOT trial while practicing standard placebo see Text S1 The placebo dilemma and the rationale for a crossover approach are further discussed below following the rationale to selecting patients at late chronic stage First as explained in 3 applying hyperbaric oxygen in the acute or early phase after brain injury makes it almost impossible to signify and assess the HBOT effects vs the effects of the spontaneous natural repair mechanism that are effective at this stage Moreover in some patients the elevated oxygen can inhibit natural regeneration or even cause toxicity In 3 it was proposed that this might explain the contradictive results in studies using HBOT at early stage after stroke 71 72 73 74 75 One can assume that any added energy during the degenerative stage immediately after brain injury could further increase the unwanted post injury damage On the other hand elevated oxygen supply during the regenerative stage would supply the energy needs for the innate brain repair processes Second as also explained in 3 patients at the chronic late stage demonstrate neurological stability with low probability of spontaneous changes unrelated to treatment Third typically patients at this stage years after injury have already gone through rehabilitation programs These programs which attempt to attend PLOS ONE www plosone org The placebo dilemma and debate There are inherent ethical and logistic difficulties in handling the sham control in HBOT trial according to the standard placebo definition Medically ineffectual treatment for medical conditions intended to deceive the recipient from knowing which treatment is given First the minimal pressure for the patients to sense pressure increase is 1 3Atm Second breathing regular air under hyperbaric conditions of 1 3Atm leads to more than 50 elevation in tissue oxygenation There are many case reports illustrating significant effects due to small increases in air pressure including effects on the brain 38 78 79 80 Moreover even a slight increase in partial pressure such as to 1 05 ATM at altitude 402 m below sea level the Dead Sea can lead to noticeable physiological effects 81 82 83 84 85 Since 50 elevation in tissue oxygen can have significant physiological effects treatment with room air at 1 3Atm is not an ineffectual treatment as is required from a proper sham control Yet a recent randomized controlled trial on mTBI patients by Wolf et al 36 used room air at 1 3Atm as sham control for treatment with 100 oxygen at 2 4Atm Both groups 12 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 8 Volume rendered Brain SPECT perfusion maps of Example 1 a 51 year old woman from the treated group suffering mTBI that had occurred 2 years prior to inclusion in the study Comparison of the baseline activity upper row with the post HBOT activity middle row and the CBF changes bottom row demonstrated significant improvements after HBOT in bilateral orbito frontal and lateral parietal regions and left ventro lateral frontal region correlating to BAs 45 47 and 11 doi 10 1371 journal pone 0079995 g008 test this issue by evaluating the specific dose response in post mTBI patients A potential way to comply with standard placebo could be to expose the patients to normal pressure combined with falsifying stimulations e g by increasing and decreasing the pressure which generates a fictitious pressure sensation This approach poses non trivial logistic difficulties Some patients especially in long term repeated treatments can detect pressure fluctuations Another potential way to avoid the increase in tissue oxygen at 1 3Atm in order to attain a standard placebo is to let the patients breath air with lower than normal oxygen level Obviously this is an unsuitable approach as it involves ethical issues and leaves an open question with regards to the pressure effect Nevertheless Cifu et al 35 conducted a randomized blinded clinical study in which 2 0 ATA with 10 5 oxygen was used as the sham control More specifically the patients were at 2 0 ATA but were randomly assigned to one of three groups breathing either 10 5 75 or 100 oxygen to mimic normal air at 1 0 ATA 100 air at 1 5 ATA and 100 air at 2 0 ATA respectively The authors concluded that This study demonstrated that HBO2 at either 1 5 or 2 0 ATA equivalent had no effect on post concussion symptoms after mild traumatic brain injury when compared with sham revealed significant improvements in cognitive symptoms and in the measure of post traumatic stress disorder PTSD We find these results very important they actually demonstrate that the significantly less expensive and logistically simpler treatment of mTBI patients with mild HBNO2 mild hyperbaric pressure of 1 3Atm and regular air can lead to meaningful improvements Our interpretation is based on previous studies demonstrating that mild HBNO2 conditions can be effectual treatment The authors of that study presented a very different interpretation Overlooking the fact that mild HBNO2 can be an effectual treatment they regarded it as sham control and concluded that the observed improvements must be due to placebo and that HBOT has no therapeutic effect on mTBI patients In other words they implicitly assumed that bringing the patients many times to spend long duration in the hyperbaric chamber can trigger such a powerful placebo effect that it can lead to a significant repair of chronic brain damage due to mTBI Remembering that for mTBI patients with intact macro vascular bed breathing 100 oxygen at 2 4ATA generate very high oxygen levels in tissues which can cause an inhibitory effect or even focal toxicity it is conceivable that HBOT using 2 4 ATA can be less effective than 1 3 ATA or other lower levels of pressure 86 Future studies are needed to PLOS ONE www plosone org 13 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 9 Volume rendered Brain SPECT perfusion maps of Example 2 The results are of a patient in the treated group suffering mTBI that had occurred 1 year prior to inclusion in the study Comparison of the baseline activity upper row with the post HBOT activity middle row and the CBF changes bottom row demonstrated significant improvements after HBOT in bilateral orbito frontal regions the medial aspect of the temporal lobes and the temporal poles that correspond to BAs 11 25 27 28 and 38 doi 10 1371 journal pone 0079995 g009 crossover approach involves two groups a treated group in which the patients went through two months of 40 HBOT sessions and a crossover group in which the patients first went through two month of no treatment followed by two months of HBOT sessions The advantage of the crossover approach is the triple comparison between treatments of two groups between treatment and no treatment of the same group and between treatment and no treatment in different groups see Text S1 For both groups the HBOT sessions induced statistically significant improvement in cognitive functions according to four cognitive indices Information Processing Speed Attention Memory and Executive functions in brain activity according to SPECT imaging and in quality of life according to the EQ 5D and the EQ VAS scores compared to the control period of the crossover group To gain better validity of the results we used the scatter plot analysis of the changes of the cognitive indices in terms of the corresponding scaled relative changes The scatter plots figure 5 show correlations in the improvements of the different indices both for the group means and the individual patients The good correspondence between the improvements in the cognitive indices the quality of life scores and the elevated brain activity as revealed by the SPECT imaging which was done in a compression Unfortunately the HBOT effect in this study was assessed merely based on the self administered Rivermead PostConcussion Symptoms Questionnaire RPQ which is known to display several flaws in implementation and in its ability to accurately reflect test taker experience Moreover interpretation and accuracy of the RPQ can vary widely due to selfadministration and the various confounding variables involved 87 Put aside this weakness the study suffers from a logical flaw The authors mention that their study was motivated by the results of Wolf et al 36 and they accepted the interpretation that any observed improvements should be a reflection of placebo effect and have nothing to do with the HBOT If indeed placebo can be so powerful in mTBI patients one would expect that stress related to the idea of breathing half the normal level of air may trigger powerful negative placebo effect Rationale for the crossover approach In the current study we tested the effect of 1 5 ATA using the crossover approach As stated in the introduction the approach is adopted in order to avoid the inherent difficulties associated with conducting HBOT trial while practicing standard placebo The PLOS ONE www plosone org 14 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 10 Volume rendered Brain SPECT images representing the percentage of CBF change post control period and post HBOT of the cross group patient described in example 3 As can be clearly seen the improvement in perfusion following HBOT was significantly high in most areas of the brain as opposed to insignificant change following the control period The most significant improvements were in both frontal and temporal lobes and right parietal lobe doi 10 1371 journal pone 0079995 g010 how to optimize patient specific protocol is important subject for future research In conclusion this study provides for the first time convincing results based on a crossover study demonstrating that HBOT can induce neuroplasticity and significant brain function improvements in mild TBI patients with prolonged Post Concussion Syndrome at late chronic stage years after injury The results call for better understanding of how to set the optimal HBOT protocol for the specific patients and how to determine which patients benefit the most from this treatment The findings reported here bear the promises that HBOT can be effective in treating other brain impairments like easing PTSD symptoms or repairing radiation damage It is also reasonable to expect that HBOT can help slow down or even reverse metabolic disorders associated neurodegenerative diseases completely blinded fashion further substantiates the clinical findings Implications Combined with previous studies of the HBOT effects on TBI and CVA patients the results presented here show that treatment with hyperbaric oxygen can significantly repair the chronically impaired brain functions and dramatically improve the quality of life of these patients Yet HBOT did not become a common acceptable treatment for TBI and CVA largely because of the debate regarding the placebo issue and the optimal time for administration Additional larger scale clinical studies are required to asses if and to what extent placebo effects might be operative However since the improvements are significant with no significant side effects it seems reasonable to let patients benefit from HBOT now rather than wait until future studies are completed We foresee that the future oxygen pressure dose response studies described in the discussion section will have significant therapeutic implications In particular we expect that HBOT treatment with room air at 1 3ATA will have significant brain repair effects and its effect should be compared with the 1 5ATA protocol used in this study In the current study the HBOT effects were assessed shortly after treatment ended Future follow up studies are needed in order to investigate the durability of the effect It might be that some patients will need more than 40 HBOT sessions The issue of PLOS ONE www plosone org Supporting Information Table S1 SPECT based measurements of changes in brain activity This SI includes data regarding the SPECT imaging for all the patients Table S1 1 S1 3 The data was normalized according to Cerebellum activity and the relative change percentage from baseline was calculated for each subject for each Brodmann area Average and STD of all subjects were then calculated for each BA The data is available for all three groups of subjects control group after waiting period control group after HBOT crossover and treated group after HBOT 15 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI Figure 11 Volume rendered Brain SPECT images representing the CBF change in percentage post control period and post HBOT of the cross group patient described in example 4 The overall changes after the control period and the HBOT show normal variations of brain perfusion in the 10 to 10 range from green to orange colors However close inspection reveals localized significant changes white circles in the in the right temporal pole and in the right dorso lateral area These changes in perfusion are in good agreement with the improvements in the cognitive indices as the SPECT detected changes correspond to Brodmann areas 45 46 11 38 and 39 doi 10 1371 journal pone 0079995 g011 use of their software Dr Glen Doniger in particular for his great help in administering the software s data and results and Prof Avraham Schweiger for his contribution to implementing the software in our study We are grateful to Dr Hanna Levi for valuable help with the statistics and data analysis and Dr Yaron Gross for his help with the randomizations analysis We are thankful Dr Alexander Vol and Dr Orna Gribova for enlightening discussions regarding the design of the proper hyperbaric oxygen therapy for TBI patients We thank Dr Nachum Gal and Dr Laura Herzog for their help in initiating this study We thank the following individuals for their important contribution in patients management during this study Alona Esterin Mazi Aski Sela Angela Chanimov Malca Katovski Lea Shkolnic Eyal Malca Vitali Triban Talia levy Doing so ease associating the changes in SPECT measurements of brain activity with the assessed changes in the cognitive indices PDF Protocol S1 Clinical study protocol DOCX Form S1 Informed consent form English translation DOCX Checklist S1 CONSORT 2010 checklist DOCX Text S1 Crossover approach and simulated randomi zation DOC Author Contributions Conceived and designed the experiments RB G HG SE Performed the experiments HG GF YB OV NS Analyzed the data RB G HG JB MF DH EB J SE Contributed reagents materials analysis tools HG OV SE Wrote the paper RB G HG EB J SE Acknowledgments The authors express special thanks to Michal Ben Jacob for her significant help in editing the manuscript We thank NeuroTrax Corporation for the References 5 Bazarian JJ Wong T Harris M Leahey N Mookerjee S et al 1999 Epidemiology and predictors of post concussive syndrome after minor head injury in an emergency population Brain injury BI 13 173 189 6 McCauley SR Boake C Pedroza C Brown SA Levin HS et al 2005 Postconcussional disorder Are the DSM IV criteria an improvement over the ICD 10 The Journal of nervous and mental disease 193 540 550 7 Kashluba S Paniak C Blake T Reynolds S Toller Lobe G et al 2004 A longitudinal controlled study of patient complaints following treated mild traumatic brain injury Archives of clinical neuropsychology the official journal of the National Academy of Neuropsychologists 19 805 816 8 Iverson GL 2005 Outcome from mild traumatic brain injury Current opinion in psychiatry 18 301 317 1 Coronado VG Xu L Basavaraju SV McGuire LC Wald MM et al 2011 Surveillance for traumatic brain injury related deaths United States 1997 2007 Morbidity and mortality weekly report Surveillance summaries 60 1 32 2 Faul M XL Wald MM Coronado VG 2010 Traumatic Brain Injury in the United States Emergency Department Visits Hospitalizations and Deaths 2002 2006 Atlanta GA Centers for Disease Control and Prevention National Center for Injury Prevention and Control 3 Efrati S Fishlev G Bechor Y Volkov O Bergan J et al 2013 Hyperbaric oxygen induces late neuroplasticity in post stroke patients randomized prospective trial PloS one 8 e53716 4 Cao H Ju K Zhong L Meng T 2013 Efficacy of hyperbaric oxygen treatment for depression in the convalescent stage following cerebral hemorrhage Experimental and therapeutic medicine 5 1609 1612 PLOS ONE www plosone org 16 November 2013 Volume 8 Issue 11 e79995

Hyperbaric Oxygen Therapy for TBI 9 Bohnen N Jolles J Twijnstra A 1992 Neuropsychological deficits in patients with persistent symptoms six months after mild head injury Neurosurgery 30 692 695 discussion 695 696 10 Binder LM 1997 A review of mild head trauma Part II Clinical implications Journal of clinical and experimental neuropsychology 19 432 457 11 Bazarian JJ McClung J Shah MN Cheng YT Flesher W et al 2005 Mild traumatic brain injury in the United States 1998 2000 Brain injury BI 19 85 91 12 Kushner D 1998 Mild traumatic brain injury toward understanding manifestations and treatment Archives of internal medicine 158 1617 1624 13 Medana IM Esiri MM 2003 Axonal damage a key predictor of outcome in human CNS diseases Brain a journal of neurology 126 515 530 14 Kochanek PM Clark R S B Jenkins L W 2007 TBI Pathobiology In Zasler ND Katz D I Zafonte R D editor Brain injury medicine NY Demos medical publishing pp 81 92 15 Levin HS Mattis S Ruff RM Eisenberg HM Marshall LF et al 1987 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Hyperbaric Oxygen Therapy for TBI 66 Neubauer RA Walker M 2000 Hyperbaric Oxygen Therapy Garden City Park NY Avery Publishing Group 67 Kuffler DP 2011 The role of hyperbaric oxygen therapy in enhancing the rate of wound healing with a focus on axon regeneration Puerto Rico health sciences journal 30 35 42 68 Rockswold SB Rockswold GL Defillo A 2007 Hyperbaric oxygen in traumatic brain injury Neurological research 29 162 172 69 Zhou Z Daugherty WP Sun D Levasseur JE Altememi N et al 2007 Protection of mitochondrial function and improvement in cognitive recovery in rats treated with hyperbaric oxygen following lateral fluid percussion injury Journal of neurosurgery 106 687 694 70 Calvert JW Cahill J Zhang JH 2007 Hyperbaric oxygen and cerebral physiology Neurological research 29 132 141 71 Anderson DC Bottini AG Jagiella WM Westphal B Ford S et al 1991 A pilot study of hyperbaric oxygen in the treatment of human stroke Stroke a journal of cerebral circulation 22 1137 1142 72 Nighoghossian N Trouillas P Adeleine P Salord F 1995 Hyperbaric oxygen in the treatment of acute ischemic stroke A double blind pilot study Stroke a journal of cerebral circulation 26 1369 1372 73 Rusyniak DE Kirk MA May JD Kao LW Brizendine EJ et al 2003 Hyperbaric oxygen therapy in acute ischemic stroke results of the Hyperbaric Oxygen in Acute Ischemic Stroke Trial Pilot Study Stroke a journal of cerebral circulation 34 571 574 74 Vila JF Balcarce PE Abiusi GR Dominguez RO Pisarello JB 2005 Improvement in motor and cognitive impairment after hyperbaric oxygen therapy in a selected group of patients with cerebrovascular disease a prospective single blind controlled trial Undersea hyperbaric medicine journal of the Undersea and Hyperbaric Medical Society Inc 32 341 349 75 Imai K Mori T Izumoto H Takabatake N Kunieda T et al 2006 Hyperbaric oxygen combined with intravenous edaravone for treatment of acute embolic stroke a pilot clinical trial Neurologia medico chirurgica 46 373 378 discussion 378 76 Glisky EL 2004 Disorders of memory In Ponsford J editor Cognitive and Behavioral Reabilitation From Neurobiology to Clinical Practice New York The Guilford Press pp 100 129 PLOS ONE www plosone org 77 de Frias CM Dixon RA Backman L 2003 Use of Memory Compensation Strategies Is Related to Psychosocial and Health Indicators The Journals of Gerontology Series B Psychological Sciences and Social Sciences 58 P12 P22 78 Golding FC Griffiths P Hempleman HV Paton WD Walder DN 1960 Decompression sickness during construction of the Dartford Tunnel British journal of industrial medicine 17 167 180 79 Austin D 1998 Gammow bag for acute mountain sickness Lancet 351 1815 80 Mychaskiw G 2nd Stephens PL 2013 Hyperbaric Oxygen Mild Traumatic Brain Injury and Study Design An Elusive Target Journal of neurotrauma 81 Goldbart AD Cohen AD Weitzman D Tal A 2007 Effects of rehabilitation winter camps at the Dead Sea on European cystic fibrosis patients The Israel Medical Association journal IMAJ 9 806 809 82 Kramer MR Springer C Berkman N Glazer M Bublil M et al 1998 Rehabilitation of hypoxemic patients with COPD at low altitude at the Dead Sea the lowest place on earth Chest 113 571 575 83 Falk B Nini A Zigel L Yahav Y Aviram M et al 2006 Effect of low altitude at the Dead Sea on exercise capacity and cardiopulmonary response to exercise in cystic fibrosis patients with moderate to severe lung disease Pediatric pulmonology 41 234 241 84 Abinader EG Sharif D Rauchfleich S Pinzur S Tanchilevitz A 1999 Effect of low altitude Dead Sea location on exercise performance and wall motion in patients with coronary artery disease The American journal of cardiology 83 250 251 A255 85 Gabizon I Shiyovich A Novack V Khalameizer V Yosefy C et al 2011 Impact of descent and stay at a Dead Sea resort low altitude on patients with systolic congestive heart failure and an implantable cardioverter defibrillator The Israel Medical Association journal IMAJ 13 402 407 86 Mychaskiw Ii G Stephens P 2012 Hyperbaric oxygen mild traumatic brain injury and study design an elusive target Journal of neurotrauma 87 Potter S Leigh E Wade D Fleminger S 2006 The Rivermead Post Concussion Symptoms Questionnaire a confirmatory factor analysis Journal of neurology 253 1603 1614 18 November 2013 Volume 8 Issue 11 e79995

Research Effect of hyperbaric oxygen therapy on Amir Hadanny 1 2 3 4 Stefanie Abbott 2 Gil Suzin 2 Yair Bechor 2 Shai Efrati2 4 5 6 To cite Hadanny A Abbott S Suzin G et al Effect of hyperbaric oxygen therapy on chronic neurocognitive deficits of post traumatic brain injury patients retrospective analysis BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 Prepublication history for this paper is available online To view these files please visit the journal online http dx doi org 10 1136 bmjopen 2018023387 EUBS 2017 EANS 2017 Received 5 April 2018 Revised 27 June 2018 Accepted 24 July 2018 Author s or their employer s 2018 Re use permitted under CC BY NC No commercial re use See rights and permissions Published by BMJ 1 Neurosurgery Department Galilee Medical Center Nahariya Israel 2 Sagol Center for Hyperbaric Medicine and Research Assaf Harofeh Medical Center Zerifin Israel 3 Galilee Faculty of Medicine Bar Ilan University Ramat Gan Israel 4 Sackler School of Medicine Tel Aviv University Tel Aviv Israel 5 Research and Development Unit Assaf Harfoeh Medical Center Zerifin Israel 6 Sagol School of Neuroscience Tel Aviv University Tel Aviv Israel Correspondence to Dr Amir Hadanny amir had gmail com ABSTRACT Objectives The aim of the study is to evaluate the effect of hyperbaric oxygen therapy HBOT in participants suffering from chronic neurological deficits due to traumatic brain injury TBI of all severities in the largest cohort evaluated so far with objective cognitive function tests and metabolic brain imaging Methods A retrospective analysis was conducted of 154 patients suffering from chronic neurocognitive damage due to TBI who had undergone computerised cognitive evaluations pre HBOT and post HBOT treatment Results The average age was 42 7 14 6 years and 58 4 were men All patients had documented TBI 0 3 33 years mean 4 6 5 8 median 2 75 years prior to HBOT HBOT was associated with significant improvement in all of the cognitive domains with a mean change in global cognitive scores of 4 6 8 5 p

multiple brain areas 9 10 cognitive impairments are usually the predominant symptoms Global brain hypoperfusion and its related tissue ischaemia detected in patients suffering from TBI serves as a rate limiting factor for any regenerative process 11 13 By increasing the oxygen level in blood and body tissues HBOT can augment the repair mechanisms 5 Various models have strongly suggested that HBOT can induce angiogenesis improve brain plasticity enhance neurogenesis and synaptogenesis and foster functional recovery 14 15 Conflicting clinical HBOT data and objective measurements in PCS Some of the previous studies which evaluated the effect of HBOT on chronic neurological and cognitive impairments due to TBI mainly used self assessment questionnaires as their primary endpoints 16 18 Such endpoints have several inherent disadvantages First they lack an objective evaluation that is not biased by the patients perspectives Second self administrated questionnaires are exposed to various confounding variables such as litigation and compensation 19 Unlike the questionnaires standardised cognitive tests with high test retest reliability can and should be used as objective evaluations of neurocognitive impairments 20 In addition novel brain imaging techniques such as single photon emission computed tomography SPECT and perfusion sequences in MRI which evaluate cerebral blood flow and brain metabolism can shed new light in PCS diagnosis and in evaluating therapeutic interventions 20 In clinical studies which used objective cognitive assessments HBOT was found to induce significant improvements in patients suffering from PCS due to mild TBI 5 6 15 21 However to the best of our knowledge the objective effect of HBOT on chronic neurocognitive impairments stemming from moderate to severe TBI in addition to mild has not been investigated In addition to objective evaluations there are inherent ethical and logistic difficulties in handling the sham control in HBOT trials 4 5 20 22 HBOT includes two active ingredients pressure and oxygen Pressure is needed to increase plasma oxygen but the pressure change alone may also have significant cellular effects 5 Additionally the greatest effect of pressure is in human tissues that are under tight autoregulation pressure control such as the brain where the intracranial pressure is normally 0 0092 0 0197 atm 23 24 To generate a pressure sensation the chamber pressure must be 1 2 ATA or higher However such a change in environmental pressure from 1 ATA to 1 2 ATA and subsequent tissue oxygenation with an increase of tissue oxygenation by at least 50 has a significant biological effect 25 26 Thus sham therapy in previous studies using 1 2 ATA on 21 inhaled oxygen ie air cannot be regarded as an inert or sham control but rather as a lower dose of the active ingredient 4 20 In regards to a possible effect of vasoconstriction of the large blood vessels induced by hyperbaric oxygen it has been 2 well established that the tissues are saturated by hyperoxia and do not suffer from hypoxia as the vasoconstriction effect is compensated by increased plasma oxygen content and microvascular blood flow 27 Any increase in pressure even with reduced oxygen percentage cannot serve as a true placebo but rather as a low dosage of the active ingredient further supporting the need for objective data gathered from large cohorts of patients suffering from PCS and treated by HBOT The aim of the current study was to evaluate the objective effects of HBOT on patients with TBI suffering from chronic neurological deficits stemming from mild moderate and severe TBI in the largest cohort evaluated until now Since all the patients had metabolic brain imaging and a computerised neurocognitive test battery before and after HBOT correlations between specific cognitive indexes and their related brain regions activity were also evaluated MATERIALS AND METHODS Participants A retrospective analysis was conducted on patients suffering from TBI related chronic neurocognitive damage more than 3 months from injury treated by HBOT between January 2008 and January 2017 at the Sagol Center for Hyperbaric Medicine and Research Assaf Harofeh Medical Center Israel Patients were included if they had pre HBOT and post HBOT computerised cognitive evaluations Patients with a history of potential additional brain insults such as spontaneous subarachnoid haemorrhage anoxic brain injury or history of prior cognitive impairment were excluded figure 1 Patients and public involvement Patients and public weren t involved in the study due to its retrospective nature Figure 1 Patients flowchart TBI traumatic brain injury HBOT hyperbaric oxygen therapy Hadanny A et al BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 BMJ Open first published as 10 1136 bmjopen 2018 023387 on 28 September 2018 Downloaded from http bmjopen bmj com on October 5 2022 by guest Protected by copyright Open access

TBI severity TBI severities were rated according to the TBI admission documents Mild TBI was defined as loss of consciousness LOC with duration of 0 30 min post traumatic amnesia PTA with duration of less than a day and a Glasgow Coma Scale GCS grade of 13 15 28 Moderate TBI was defined as LOC with duration of more than 30 min and up to 24 hours PTA with duration of 1 7 days and GCS grade of 9 12 Severe TBI was defined as LOC with duration of more than 24 hours PTA with duration of more than 7 days and GCS less than 9 In addition if there was imaging evidence of an injury such as a haematoma contusion or haemorrhage then the TBI was classified as moderate to severe 28 Hyperbaric oxygen treatment Patients were treated with 40 70 daily hyperbaric sessions 5 days a week Each session consisted of 60 90 minutes of exposure to 100 oxygen at 1 5 2 ATA Cognitive assessment The patients cognitive functions were assessed by NeuroTrax computerised cognitive tests NeuroTrax 29 The NeuroTrax tests evaluate various aspects of brain functions and include verbal memory immediate and delayed recognition non verbal memory immediate and delayed recognition go no go response inhibition problem solving Stroop interference finger tapping catch game staged information processing speed single digit two digit and three digit arithmetic verbal function and visual spatial processing Cognitive index scores were computed from the normalised outcome parameters for memory executive function attention information processing speed visual spatial verbal function and motor skills domains 30 A global cognitive score was computed as the average of all index scores for each individual After administration the NeuroTrax data were uploaded to the NeuroTrax central server and outcome parameters were automatically calculated using software blind to diagnosis or testing site To account for the wellknown effects of age and education on cognitive performance each outcome parameter was normalised and fit to an IQ like scale mean 100 SD 15 according to the patient s age and education The normative data used by NeuroTrax consist of test data from cognitively healthy individuals in controlled research studies at more than 10 sites 31 Specifically the patients were given two different versions of the NeuroTrax test battery before and after HBOT to allow repeated administrations with minimal learning effects Test retest reliability for these versions was evaluated and found to be high with no significant learning effect 32 33 Regarding the current study cohort in a previous randomised controlled trial in patients suffering from TBI the NeuroTrax scores were found to be stable in the retest of the control group 21 Hadanny A et al BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 Brain SPECT imaging Brain activity was assessed using SPECT 1 2 weeks prior to and after the HBOT period The SPECT method was selected for evaluation due to its known normal range and test retest established validity The imaging was conducted using 925 1110 MBq 25 30 mCi of a technetium 99 methyl cysteinate dimmer Tc 99m ECD at 40 60 min postinjection using a dual detector gamma camera ECAM or Symbia T Siemens Medical Systems equipped with high resolution collimators Data were acquired in three degree steps and reconstructed iteratively using the Chang method of attenuation correction 0 12 cm 34 Both pretreatment and post treatment SPECT images were normalised to the median maximal brain activity in the entire brain and were then reoriented into Talairach space using NeuroGam software Segami to identify Brodmann cortical areas and to compute the mean perfusion in each Brodmann area BA In addition volume rendered brain perfusion images were reconstructed and normalised to the entire brain median maximal activity All SPECT analyses were done by study team members who were blinded to the laboratory and clinical data SPECT scans were performed late morning to midday On the day of the SPECT scan patients were treated with only their chronic medications and were instructed not to smoke Changes in perfusion in all Brodmann areas for each subject were determined by calculating the percentage of the difference of the normalised activity values between post treatment and pretreatment divided by the pretreatment value Statistical analysis Continuous data were expressed as means SDs The normal distribution for all variables was tested using the Kolmogorov Smirnov test The mean differences between cognitive index scores before and after HBOT were analysed using one way analysis of variance ANOVA with post hoc Bonferroni tests Multiple linear regression models and multivariate logistic regression models were performed to control for potential confounders and to determine independent predictors for clinical outcome The alpha level was set to 0 05 Data were statistically analysed using SPSS software V 22 0 RESULTS Patient profiles Of the 242 patients suffering from neurocognitive impairment due to TBI treated by HBOT between January 2008 and January 2017 25 patients had potential additional brain insults and 63 did not have repeat computerised neurocognitive evaluations Therefore 154 patients were included in the final analysis of whom 100 patients completed pre HBOT and post HBOT SPECT imaging figure 1 The patients baseline characteristics are summarised in table 1 The average age was 42 7 14 6 years and 58 4 3 BMJ Open first published as 10 1136 bmjopen 2018 023387 on 28 September 2018 Downloaded from http bmjopen bmj com on October 5 2022 by guest Protected by copyright Open access

Table 1 Baseline patient characteristics Characteristics Total Mild TBI Moderate TBI Severe TBI Significance Patients n Age years 154 100 42 7 14 6 69 44 8 48 8 12 0 24 15 6 41 7 12 7 61 39 6 36 2 15 3

Table 2 Cognitive indices pre HBOT and post HBOT of the entire study cohort Baseline PostHBOT Mean change P values General Memory 88 3 15 2 81 7 23 2 92 9 14 2 89 9 21 9 4 6 8 5 8 1 16 9

Figure 4 Mean changes of post HBOT compared with pre HBOT across the different TBI severities Both patients who suffered mild and severe TBI groups had improvements in general memory attention information processing speed and motor skills scores whereas patients who suffered moderate TBI had significant improvement in memory P10 change in cognitive indices proportions across traumatic brain injury TBI groups General Memory Total Mild TBI 36 23 4 64 41 6 15 21 7 28 40 6 Moderate TBI Severe TBI P values 7 29 2 9 37 5 14 23 0 27 44 3 0 756 0 830 Executive functions 51 33 1 23 33 3 7 29 2 21 34 9 0 897 Attention Information processing speed 62 40 3 48 31 2 27 39 1 23 33 3 8 33 3 12 50 27 44 3 13 21 3 0 631 0 032 6 Hadanny A et al BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 BMJ Open first published as 10 1136 bmjopen 2018 023387 on 28 September 2018 Downloaded from http bmjopen bmj com on October 5 2022 by guest Protected by copyright Open access

Figure 5 The mean relative change in Broadmann areas posthyperbaric oxygen therapy for the entire study cohort This study has several limitations The major one relates to its retrospective methodology This limitation is diminished when considering that this large cohort of patients was treated at late chronic stages The findings presented here are in agreement and reinforce the findings from previous prospective controlled trials in which the neuroplasticity effects of HBOT were demonstrated in chronic stages of different types of brain injuries 15 21 42 43 Moreover the correlation between the changes in cognitive function and the metabolic brain imaging gives further strength to the results Another important limitation relates to the HBOT protocol which was inconsistent across the cohort Although significant neurotherapeutic effects were seen with 60 min of 1 5 ATA the optimal protocol needed to induce maximal neuroplasticity for the specific individual with minimal side effects has not been investigated The strengths of the study are worth mentioning First objective cognitive assessments using computerised tests were performed on each patient both pretreatment and post treatment Objective measures are significantly superior to PCS questionnaires which are inaccurate variable and contain various confounders rather than reflect the true PCS state 44 Second most of the patients in the study underwent an objective ancillary brain SPECT to confirm PCS diagnosis prior to HBOT This practice is crucial when considering the differential diagnosis following TBI PTSD depression etc Moreover post treatment brain SPECTs revealed an anatomical functional correlation in regards to HBOT s Figure 6 Cognitive functions correlated with Brodmann areas Each of the traumatic brain injury TBI groups mild moderate and severe had perfusion metabolism increase in specific Brodmann areas correlated with improved cognitive function Hadanny A et al BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 7 BMJ Open first published as 10 1136 bmjopen 2018 023387 on 28 September 2018 Downloaded from http bmjopen bmj com on October 5 2022 by guest Protected by copyright Open access

effect in brain neuroplasticity Third the study cohort consisted of a civilian population that does not have any potential secondary gain such as financial compensation by reporting sick Previous studies included patients with PCS who suffered mild TBI injury Considering its strengths and limitations the current study implies that the cognitive function of patients with post TBI can be improved significantly irrespectively of whether the primary brain injury was classified as mild moderate or severe Although long term data are still lacking considering the high safety profile of the treatment these results are promising and should encourage rehabilitation centres to consider HBOT for patients with chronic neurocognitive deficits following TBI Future studies should monitor these patients in the long term 6 months 12 months as well as their return to activities of daily living CONCLUSIONS HBOT was associated with significant cognitive improvements in patients who suffer from chronic neurocognitive deficits due to mild moderate and severe TBI Improvement in memory correlated with activation of the perirhinal cortex improvement of executive functions correlated with activation of the inferior frontal gyrus and improvement in attention correlated with activation of the anterior cingulate gyrus Contributors AH concept data collection data analysis manuscript draft and manuscript review SA data collection and data analysis GS YB data collection and manuscript review SE concept data analysis manuscript draft and manuscript review Funding This research received no specific grant from any funding agency in the public commercial or not for profit sectors Competing interests None declared Patient consent Data collected retrospectively were anonymised Ethics approval The study was approved by the institutional review board of Assaf Harfoeh Medical Center Israel Provenance and peer review Not commissioned externally peer reviewed Data sharing statement Extra data are 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32 Melton JL Psychometric evaluation of the Mindstreams neuropsychological screening tool Panama City Navy Experimental Diving Unit US 2005 33 Schweiger GMD A Dwolatzky T D Jaffe E S Simon reliability of a novel computerized neuropsychological battery for mild cognitive impairment Acta Neuropsychologica 2003 1 407 13 34 Jaszczak RJ Chang LT Stein NA et al Whole body single photon emission computed tomography using dual large field of view scintillation cameras Phys Med Biol 1979 24 1123 43 35 Hadanny A Golan H Fishlev G et al Hyperbaric oxygen can induce neuroplasticity and improve cognitive functions of patients suffering from anoxic brain damage Restor Neurol Neurosci 2015 33 471 86 36 Efrati S Fishlev G Bechor Y et al Hyperbaric oxygen induces late neuroplasticity in post stroke patients randomized prospective trial PLoS One 2013 8 e53716 37 Chen J Zhang ZG Li Y et al Intravenous administration of human bone marrow stromal cells induces angiogenesis in the ischemic boundary zone after stroke in rats Circ Res 2003 92 692 9 Hadanny A et al BMJ Open 2018 8 e023387 doi 10 1136 bmjopen 2018 023387 38 Brown MW Aggleton JP Recognition memory what are the roles of the perirhinal cortex and hippocampus Nat Rev Neurosci 2001 2 51 61 39 Aron AR Robbins TW Poldrack RA Inhibition and the right inferior frontal cortex Trends Cogn Sci 2004 8 170 7 40 Miller EK Cohen JD An integrative theory of prefrontal cortex function Annu Rev Neurosci 2001 24 167 202 41 Bush G Luu P Posner MI Cognitive and emotional influences in anterior cingulate cortex Trends Cogn Sci 2000 4 215 22 42 Harch PG Hyperbaric oxygen in chronic traumatic brain injury oxygen pressure and gene therapy Med Gas Res 2015 5 9 43 Harch PG Andrews SR Fogarty EF et al A phase I study of low pressure hyperbaric oxygen therapy for blast induced postconcussion syndrome and post traumatic stress disorder J Neurotrauma 2012 29 168 85 44 Potter S Leigh E Wade D et al The rivermead post concussion symptoms questionnaire a confirmatory factor analysis J Neurol 2006 253 1603 14 9 BMJ Open first published as 10 1136 bmjopen 2018 023387 on 28 September 2018 Downloaded from http bmjopen bmj com on October 5 2022 by guest Protected by copyright Open access

The American Institute of Stress COMBAT STRESS BRINGING YOU ALL THE WAY HOME Volume 7 Number 2 HBOT May 2018 Brain Healing Hidden in Plain Sight Volume 2 in a 2 Part HBOT Series 3 May 2018 AIS Combat Stress www stress org

CONTENTS 4 Overview of this Edition Part 2 Expanding Options 6 MAJ RET Ben Richards and his HBOT Treatment Experience 12 To Treat a Concussion Using Hyperbaric Oxygen to Mitigate the Duration of Symptoms of Acute Concussions 18 All the Right Moves The Need for the Timely use of Hyperbaric Oxygen Therapy for Treating TBI CTE PTSD 29 A Discussion of the Use of Hyperbaric Oxygen Therapy as a Primary and Adjunctive Treatment Modality for TBI Patients 34 Patriot Clinics Press Release 38 TBI PTSD and the Suicide Epidemic Breaking the Cycle 42 IN THE NEWS TBI Treatment The Invisible Scars of War 44 Epilogue May 2018 AIS Combat Stress www stress org 3

Overview of this Edition Part 2 Expanding Options his conquest of years of neglect reflect grandly on himself and the caregivers he discovered outside conventional ineffective DOD VA Army medicine Rob Beckman PhD building on the work of many particularly Dr Daphne Denham updates how much smarter we are in 2018 about the Mechanisms of Action underlying the wound healing actions of hyperbaric oxygen Current worldwide debates about the By Robert L Beckman PhD Concussion Protocol fall miserably Executive Director TreatNOW org short of addressing a simple fact you It is not entirely impossible that don t heal wounds to the brain without perhaps sometime in the next decade oxygen and lots of it professors of medicine will have difficulty in explaining why the treatment Dr Ken Stoller s All the right moves with hyperbaric oxygen was not the need for the timely use of hyperwidely adopted much earlier Edward baric oxygen therapy for treating TBI Teller PhD 1999 In Textbook of CTE PTSD crystallizes a fundamental Hyperbaric Medicine 2017 dilemma for citizens parent ballplayers and warriors Should people who are Unless I am convinced by proof or by making money dispensing drugs make plain and clear reasons and arguments decisions about what therapies can be I can and will not retract for it is neither used to treat those brain wounds In safe nor wise to do anything against his words HBOT has met interference conscience Here I stand I can do no because other agendas are present other Martin Luther 1521 be they the protection of the status quo myopic budgetary constraints Once again famous lines from history or perceived liability issues after all tell a true story without being quite when you treat TBI directly the way true Teller and Luther upset historical that HBOT does the problems creating inevitability So too will the authors in those TBI s in the first place are harder this edition and its companion in last to ignore and the unconscious way quarter s Part 1 of the Combat Stress those problems have been dealt with E Magazine s HBOT issue attest to how are harder to deny evidence and science will change the history of medicine for brain wounds John Hughes D O and James Lyons Weiler PhD are at the forefront MAJ Ben Richards USA ret and his of neuro regenerative medicine Their family are testimony to how TBI wounds methods point to the role real science the warrior and the family His story of can play in cutting through the dilatory resilience in the face of years of the tactics of nay saying researchers best military medicine could do and pressurized oxygen is analogous 4 May 2018 AIS Combat Stress www stress org

for eight years Veterans Active Duty Military National Guard Reserves First Responders citizens and athletes returned to healthier lives after being told there was little to nothing conventional medicine could do to heal the wounds to their brains The latest count First Coast News in Jacksonville FL from 70 plus Coalition clinics across filmed a special on a Fort Hood mass the US have revealed more than 3 000 shooting survivor who is seeing drastic success stories changes after Hyperbaric Oxygenation Eight years after the attack Army In a final word evidence continues to Veteran Patrick Zeigler is another accumulate that corroborates previous example of the power of oxygen under independent studies finding that pressure to help awaken and heal the Alpha Stim and other microcurrent brain after injury See http www devices provide a viable treatment firstcoastnews com article news local option for pain anxiety insomnia and fort hood mass shooting survivor depression When used in concert sees drastic changes after hyper with hyperbaric oxygenation these baric chamber 77 506629334 non invasive safe and effective alternatives are powerful antidotes to the Partly in response to the slaughter conventional ineffective expensive at Fort Hood with which the Editor and frequently fatal standard of care is intimately connected the Patriot dispensed by military medicine Clinics movement in Oklahoma evolved to answer the call at the state level 70 000 plus suicides since 9 11 Lacking federal energy funding focus a continuing stream of 20 or more action or ethical incentive Dr William suicides a day an opioid epidemic Duncan PhD organized the first down that gained momentum due to VA payment on a state centered model prescribing protocols 1 800 000 brain to provide the care to brain wounded Veterans and public servants that is wounded Service Members since 9 11 denied them by traditional medicine and an equal number from previous The model has been replicated and wars mental health costs destroying enhanced in four more states legis military medicine budgets dramatically lation encouraging hyperbaric oxygen understaffed VA hospitals in mental treatment for the brain wounded with health positions non stop mis and some providing funding to offer the malfeasance in VA corporate structure care to those essentially relegated to lack of White House Congressional and VA leadership taking action to help permanent welfare status heal veterans brain wounds when is In TBI PTSD and the Suicide ENOUGH enough Epidemic Breaking the Cycle the TreatNOW Coalition presents six Reference stories that are illustrative of the work 1 Newsweek How VA Fueled the National Opioid Crisis and is the Coalition has been performing Killing Thousands of Veterans Art Levine October 12 2017 to plugging the brain the computer hardware into a wall socket with enough voltage to begin its self healing Then the software of multiple other therapies have a chance of running correctly May 2018 AIS Combat Stress www stress org 5

MAJ RET Ben Richards and his HBOT Treatment Experience By Ben Richards B S M A Major RET US Army During the spring and summer of 2007 I MAJ Ben Richards had the privilege of leading Bronco Troop 1 14 CAV a Stryker equipped cavalry troop during intense combat operations in and around Baqubah Iraq Bronco Troop was blessed with the deep bench of top quality Noncommissioned Officers that distinguishes great units from good ones Five of the six officers in the troop were West Pointers At one point all six of us were captains and the experience paid dividends in a challenging operating environment At the peak of operations a new second lieutenant arrived straight from the basic course to take over a scout platoon I greeted him shortly after he arrived at our dilapidated combat outpost and told him we would have a Combat Action Badge CAB for him the next day His face showed that he clearly thought I was joking By the following evening he had survived an IED hit to his Stryker been in two firefights and earned his CAB The rest of us had earned our CABs on our first day in town two months earlier as well A few weeks later he was wounded by a grenade fragment while leading his platoon in a dismounted close combat assault on an al Qaeda fighting 6 May 2018 AIS Combat Stress www stress org position The courage competence and character of these young officers was in every way a credit to our alma mater and a testimony to West Point s continuing role as the corner stone of our Nation s defense During those several months of combat operations ninety percent of my men hit at least one IED often more than one In May 2007 a suicidebomber driving a sedan laden with explosives rammed into my Stryker and destroyed it A few weeks later we hit a second plain vanilla IED buried in the road that damaged our second Stryker sufficiently that it was later coded out as not being worth fully repairing After each hit we got back up and returned to the fight because we knew that there was going to be a fight and we fought as a team even when it hurt On returning home I like so many others began a personal battle against an enemy that I could not see could not anticipate and was neither trained nor equipped to combat Six months after arriving back at Fort Lewis I was diagnosed with Post Traumatic Stress Disorder PTSD To be honest I only sought help after being command directed by my wife At that time I was

Ben Richards Stryker armored vehicle after being hit by a suicide vehicle borne IED in Baqubah Iraq in May 2013 Richards and his crew survived the attack but all with TBIs Photo Credit Ben Richards not intimidated by PTSD I had every confidence that it was something I could beat I was surprised and not a little embarrassed that I had it all I gave it a year tops By then I would be fully back in the saddle The extent of damage to my brain caused by the pair of mild traumatic brain injuries was not recognized until more than three years after the injuries and was not fully diagnosed until yet another year had passed While I was serving in Iraq I was extremely fortunate to be selected by the History Department at the US Military Academy West Point to return for a tour as an instructor I arrived in the summer of 2010 in pretty rough shape Less than a year into the assignment I collapsed under the weight of disabling chronic pain memory problems cognitive deficits sleep deprivation drugs the legal kind emotional problems and all the noise that often accompanies invisible injuries At one point heavily under the influence of prescription medications I even seriously considered taking my own life West Point was up to the challenge The History Department leadership kept me in the department so that they could personally oversee my care My fellow instructors both civilian and military took on the burden of my workload without complaint as they would have carried me my rifle and May 2018 AIS Combat Stress www stress org 7

my ruck to the CASEVAC point I am sure theirs was a long hard walk out It was real leadership at real personal cost and sacrifice The Department s Colonels breached every administrative and bureaucratic obstacle to ensure I literally received the best care available in the Department of Defense for my injury profile When it turned out that the best care was not enough and after they had done everything within their power to assure my future well being they fare welled me with honors and fanfare well beyond those merited by a junior major The day I took off my uniform for the last time was one of the saddest in my life I saw only an empty husk of the new cadet who had marched in the rain on R Day eighteen years earlier and so full of the potential that enables a first year cadet to sit with generals and presidents while a second lieutenant hides from majors in the motor pool I was permanently broken The natural processes of neural plasticity had run their course and come up wanting at the end Medications could only partially mitigate the pain while causing new problems of their own The results of evidence based psychotherapies became part of the new canon of evidence that those therapies so promising for victims of rape and traffic accidents are disappointingly much less effective against combat related PTSD Acceptance and accommodation were all that was left to aspire to 8 May 2018 AIS Combat Stress www stress org It was at that moment of hopelessness that the Long Gray Line extended its hand to drag me back from the edge John Batiste class of 74 a retired general officer and president of the Veteran serving non profit Stand for the Troops founded by the legendary COL David Hackworth SFTT org hunted me down to deliver a lifechanging message We will help you he told me and by that I mean really help you and not in the sense of providing a palliative weekend retreat or the cathartic commiseration of other wounded warriors Had John not been a graduate and a Soldier of such well known reputation I would have hung up the phone I did not have the hope left to waste on vain promises with unlikely outcomes but because John was who he was I gave him the time He gave me my life back The problem of invisible wounds and injuries was one that merited a Manhattan Project Instead it had the Army Medical Corps bureaucracy that ran Walter Reed into scandal regularly abused invisibly wounded warriors exiled to Warrior Transition Units and never seemed to get past the word excuse so clearly bookmarked in their dictionary to the word execution It was a corps of capable and dedicated medical operators who did not deserve their uninspired and ineffective leaders Their obvious failures were difficult for me to understand after having spent a career in the company of men and women I would follow anywhere And then there was the VA

Unwilling to accept defeat at the hands of inefficacious bureaucracies John and SFTT recruited a team of medical experts and began scouring the country for new and more effective approaches to treating traumatic brain injury TBI and PTSD Their rescue mission had led them to Doctor Paul Harch a practitioner of Hyperbaric Medicine at the Louisiana State University Medical School in New Orleans Harch John said would treat me Dr Harch had become the point man for the league of medical practitioners and researchers using Hyperbaric Oxygen Therapy to treat brain damage caused by TBIs By the time I arrived in New Orleans these practitioners had already treated over a hundred invisibly wounded warriors as well as several well known NFL football players to include the legendary quarterback Joe Namath Dr Harch had personally completed a research study with 20 Soldiers and Marines whose brains had been damaged by combat related TBIs The results were unprecedented When I was being evaluated by the military s top neurologists in 2011 the prevailing medical wisdom was that modern medicine could do very little if anything to help a brain heal after being damaged by a mild TBI There was a period of natural healing of up to several years but at four years postinjury they had no expectation that my brain would improve and many reasons to suspect that it would instead begin to degrade I arrived in New Orleans with repressed expectations I found Dr Paul Harch to be a dedicated and innovative professional He exhibited a reserved persona that I soon found to be a fa ade masking a burning passion for healing and especially for healing those that hope had passed by Harch is a man of great moral courage conviction and compassion a classical gentleman endowed with the noblesse oblige of an heir of a great inheritance of character and natural capacity Dr Harch and his colleagues had pioneered a protocol for using hyperbaric oxygen therapy HBOT to treat brain injuries The medicinal effects of oxygen at higher the atmospheric pressure have been recognized empirically for over a century It is perhaps best known as a treatment for diving injuries It is also widely used for healing hard to treat wounds and is approved by the FDA for over a dozen different medical conditions Using HBOT to treat brain injuries like most of the prescription medications I had been prescribed by the DOD and VA is considered off label but its safety has been recognized by the Institute of Medicine Treatment consisted of 40 one hour dives in a Plexiglas tank that I would describe as similar to a torpedo tube These were conducted at a rate of one dive sometimes two a day The tube is filled with 100 percent oxygen which is then pressurized to 1 5 atmospheres Protocols for wound healing and dive injuries use higher pressures The pressure loads oxygen into the blood stream like carbonation in an unopened can of soda May 2018 AIS Combat Stress www stress org 9

Ben Richards with his family in 2017 The ability to build and maintain meaningful social relationships has been one of the most important treatment outcomes Photo Credit Ben Richards The introduction of the extra oxygen into the brain initiates a cascade of chemical interactions that are good for you The end result is the creation of new blood vessels angiogenesis and the repair or regrowth of brain cells scans of other types had verified multiple points of structural damage The image meant that my brain was not using the amount of oxygen that a normal brain would have been That difference was apparent not only in the scans but in the neuropsychoBefore I began treatment we did a logical testing and other measures of SPECT neuro imaging scan of my cognitive and emotional impairment brain A SPECT scan uses an injective with which I had been evaluated radioactive agent to image blood flow in the brain It is one of the more sensitive imaging tools for detecting By the time I had completed 20 dives brain damage caused by mild traumatic the changes I was experiencing brain injuries mTBIs and in many were becoming undeniable Nearly cases is superior to CT or MRI scans every facet of my injury profile began especially if more than a few months to improve Pain levels dropped have elapsed since the time of injury Sleep improved Memory improved The images showed the poor blood Attention span lengthened Irritability perfusion typical of a brain damaged decreased I started feeling things I by TBIs not unexpected as previous had not felt in years Good things 10 May 2018 AIS Combat Stress www stress org

Happy things I was able to sustain a light workout program for the first time since 2008 We scanned my brain again The amount and extent of blood perfusion had increased significantly matching the subjective results that even my guarded skepticism was compelled to recognize The SPECT image is one of the most reliable predictors of the long term prognosis of brain injury and mine had just changed radically saved my marriage It has enabled me to participate in and experience life in ways that I and my DOD and VA doctors had assumed were gone for good I have even been able to contribute a little bit back I am no longer a husk Looking back on those dark days I do not believe it would be unfair to say that Dr Paul Harch and SFTT probably saved my life The Harch s covered the cost of my treatment from their own pockets as they have for dozens of other Veterans before me at no small sacrifice John and SFTT rallied donors mostly West Pointers to help cover living expenses for four months of care Gulf Coast Alumni quickly assumed an overwatch position and contributed several thousand dollars I could not have covered the costs alone Even a 100 percent VA disability rating only matches the pay of a private first class not enough to maintain dual household with four kids at home Ben Richards is a combat Veteran who suffered disabling brain damage from a Traumatic Brain Injury TBI caused by a suicide bomber in Iraq and suffered from Post Traumatic Stress Disorder PTSD He experienced significant healing from HBOT He served in the Army for 16 years in the Armor and Cavalry Branch and is a graduate of the United States Military Academy at West Point and Georgetown University He currently serves on the boards of the National Hyperbaric Association NHA and the International Hyperbaric Medical Associations IHMA where he advocates for improved access for invisibly wounded combat Veterans to hyperbaric oxygen therapy He also serves as an executive director for the Veteran serving nonprofit organization Stand for the Troops HBOT has not completely healed my wounds but it has given me more back than I thought possible More than five years after leaving Iraq a husband and a father finally came home to his family The treatment that Dr Harch provided unquestionably About the Author Army Veteran Patrick Zeigler was shot in the head at the Fort Hood massacre on November 5 2009 Thanks to Hyperbaric Oxygen treatment years after he was not expected to live much less return to a sense of normalcy Zeigler is testimony to the power of the community and the wound healing powers of oxygen under pressure Watch the video here https fcnews tv 2rg2vC2 May 2018 AIS Combat Stress 11 www stress org

TO TREAT A CONCUSSION Using Hyperbaric Oxygen to Mitigate the Duration of Symptoms of Acute Concussions By Robert L Beckman PhD A concussion is a form of traumatic brain injury TBI It is a wound to the brain The current medical standard of care for concussions relies on a protocol that essentially recommends rest It is thought that this resting period will allow the brain to recover Indeed it is common knowledge that 80 to 90 percent of concussions get better on their own 1 While it is true that some or all symptoms will diminish over time it is equally true that merely waiting for those symptoms to go away is not our best medicine nor is this good for the wounded brain risk of concussions and head injuries This includes advice about continuing play after a concussion or head injury Concussion Management in Illinois public and private schools is governed by the Youth Sports Concussion Act Public Act 099 0245 Illinois directs school boards to the Centers for Disease Control and Prevention s CDC Injury Center www cdc gov Concussion developing a formal public health response to concussion Through their HEADS UP campaign concussion educational materials are available at no cost for youth sports coaches school coaches parents and In the State of Illinois each school school and health care professionals district must use educational materials provided by the Illinois High School What the CDC does not discuss let Association to educate coaches alone the Mayo Clinic your hospital student athletes and parents guardians emergency room the NCAA nor your of student athletes about the nature and average concussion or TBI clinic is 12 May 2018 AIS Combat Stress www stress org

that a concussion is a wound to the brain that should be treated according to wound healing principles to allow the brain to heal There is no disputing the fact that in most of the millions of concussion cases experienced every year the average person will recover within a few weeks and symptoms will abate In a large fraction of cases however those symptoms do not go away and in no case will the brain become healed unless necessary steps are taken to address the wound to the cervical spine and peripheral vestibular system The literature has not evaluated early interventions as most individuals recover in 10 to 14 days A variety of treatments may be required for ongoing or persistent symptoms and impairments following injury The data support interventions including psychological cervical and vestibular rehabilitation In addition closely monitored active rehabilitation programs involving controlled subsymptom threshold submaximal exercise has been shown to be safe and may be of benefit in facilitating recovery A collaborative approach to treatment including controlled cognitive stress pharmacological treatment and school accommodations may be beneficial Further research evaluating rest and active treatments should be performed using high quality designs that account for potential confounding factors and have matched controls and effect modifiers to best inform clinical practice and facilitate recovery after SRC 2 We are all becoming familiar with the symptoms of concussions headaches confusion memory loss nausea vomiting dizziness fatigue sleepiness and emotional instability These symptoms are linked to complex pathophysiological processes affecting the brain induced by biomechanical forces These forces can lead to brain inflammation and swelling damage to blood vessels and brain cells ringing in the ears tinnitus visual and balance problems and a myriad of other overt They note that SRC is considered to and or subtle physical and emotional be among the most complex injuries in and functional difficulties sports medicine to diagnose assess and manage 3 Let us look at how Some in the worldwide sports we are now changing that paradigm community recognize a concussion and the long standing nonchalance as an injury to the brain yet nowhere about aggressively intervening in the in the gold standard Consensus concussion cascade to reverse the Statement of the 2017 Concussion damage and wounding made to a major in Sport Group Berlin 2017 is there organ of the body the brain mention of a wound to the brain In the eleven major sections of the Statement Critical to understanding the work of only one is devoted to rehabilitation It integrative medicine doctors trained in is worth quoting in full Sports related wound healing are these facts medicine concussions SRCs can result in knows a lot about how to heal wounds diverse symptoms and problems and A physical wound to the brain is like a can be associated with concurrent injury physical wound to any other organ in May 2018 AIS Combat Stress 13 www stress org

the league s command center for all games On ESPN it was reported that an estimated 400 people were involved in the concussion process including every team physician every athletic trainer every UNC and every booth spotter 6 Imagine these 400 people worried about whether or not an athlete Starting in the fall of 2015 Dr Daphne had suffered a concussion but not one Denham MD in her clinic began of them considering how to undertake treating concussions 4 We know that healing these wounds to the brain we can do better than recognition rest and recovery Simply placing a A concussion leads to inflammation patient in the Concussion Protocol5 is and cerebral ischemia deficient supply insufficient to promote recovery Team of blood to the brain that is due to trainers doctors coaches teachers obstruction of the inflow of arterial and families are then responsible for blood A host of negative chemical managing symptoms and after a while processes begin along with mitochonand with the passage of time these drial cell damage oxidative damage patients are somehow expected to be and apoptosis cell death There can better We now know that we can be be a breakdown of the blood brainfar more effective in treating wounds to the brain than waiting for these wounds barrier and brain swelling Numerous to heal on their own Consider the animal studies on concussion and blast injuries confirm that blows to the head following are just not good for your brain All There were tens of thousands of the discoveries by Dr Ann McKee and concussions sustained by athletes at her Chronic Traumatic Encephalopathy all educational levels in 2017 There CTE team in Boston confirm a strong were 540 concussion evaluations correlation between numerous hits to and 281 concussions from the NFL the head and onset of CTE At last count pre season that occurred until the week admittedly a contentious statistic before Super Bowl LII To improve their given the selection criteria she had treatment protocols the NCAA brought found CTE in 99 percent of the brains the chief medical officer of a college into studied or 110 out of 111 of former the mix demanding that an additional 7 sign off was required before returning NFL players Common sense is slowly to play The NFL went as far as causing parents coaches trainers placing more NFL independent certified and even some medical personnel to athletic trainers ATC spotters at every pay attention to these correlations game As recently as December of This is one of the primary reasons that 2017 the NFL placed a central unaffil word of mouth referrals to HBOT foriated neurotrauma consultant UNC in Concussions are increasing nationwide the body Hyperbaric Oxygen Therapy has already been approved by the FDA for certain types of wound healing air embolism arterial insufficiencies compromised skin grafts and flaps acute thermal burns crush injuries and other acute traumatic injuries that lead to oxygen and blood constriction 14 May 2018 AIS Combat Stress www stress org

A Mom s Tale Told by M J client of Dr Daphne Denham On Monday April 23rd 12 year old W J was on the monkey bars at his school Six feet off the ground he was skipping bars swinging his body a fair amount skipped a few bars grabbed the next bar and lost his grip He fell to the ground on his back shoulder and head He was helped to the nurse s office by a friend The school nurse determined he had no recollection of the fall or what he ate for lunch immediately preceding his fall W J was also having balance issues had a big headache and was lethargic Additionally he was sensitive to light and noise I picked him up from school and brought him home at 1 pm We stopped to see our pediatrician who diagnosed W J with a concussion It was shortly after we arrived home that I reached out to Dr Denham W J went into the hyperbaric chamber at 3 30 pm on the day of his accident He still had a headache when he came out of the chamber but it was not as strong He was also dizzy sleepy and generally lethargic for the remainder of the day He continued to be sensitive to light and noise On Tuesday April 24th as we drove to see Dr Denham W J commented that the oncoming cars headlights were bothering him He went into the hyperbaric chamber at 7 am and 11 30 am Both times the headache lessened but was not resolved At home he only wanted to lay on the sofa and listen to audible books He was sleeping more than normal and was still experiencing dizziness when he would get off the sofa On Wednesday April 25th two days after his fall W J went back into the hyperbaric chamber His headache improved much more after this visit It was still hovering in the 3 4 range but it would improve after each treatment His dizziness was gone and he no longer was sensitive to light and sounds W J stopped asking for Tylenol to treat his headache He was also starting to act more like himself at this point He was still hurting though he was not interested in playing video games and watching TV W J was relatively content to listen to audible books on the sofa and rest his brain We did manage to walk the dog together and he felt good following that activity On Thursday April 26th W J went into the hyperbaric chamber around 11 30 am He went in with a headache at a 2 and came out with no headache W J was feeling much better he was more energetic and had no lingering dizziness no headache and no light or noise sensitivity W J has been headache free since 1 pm on Thursday April 26th His energy levels are good He has watched TV read on his phone and a book played video games and has been a normal kid and has no headache or other concussion symptoms His pediatrician saw him on Friday April 27th and released him to go back to school on Monday April 30th May 2018 AIS Combat Stress 15 www stress org

We treat wounds to the brain Hyperbaric oxygen has been proved to enhance several natural processes related to wound healing it reduces inflammation inhibits apoptosis cell death reduces Intracranial pressure and promotes neurogenesis and angiogenesis 8 This is significant HBOT promotes neurogenesis growth of new neuronic tissue and angiogenesis growth of new blood vessels The combination of oxygen typically 100 percent O2 and pressure varying depending on the diagnosis leads to the production of more stem cells available for wound healing as well All this has been proven by rigorous scientific studies that have explored the role of oxygen and pressure in the brain healing process 9 What is not controversial is that HBOT aids tremendously in wound healing typically 20 to 40 percent faster healing than is considered the norm In the case of acute concussions and within ten days of the injury Denham demonstrated http bit ly 2jwdUwI that patients 51 out of 52 diagnosed with acute concussions completely resolved her his symptoms in five or less treatments average of 2 4 treatments per concussion Her most recent numbers of success as of February 2018 are 108 successes out of 110 patients diagnosed with acute concussions 10 Readers can learn more about this by viewing a short film entitled Concussion Help in a Hyperbaric Chamber https tinyurl com ybldktqn If brain injury patients primarily high school athletes playing contact sports like football hockey lacrosse and field hockey can be evaluated and treated 16 May 2018 AIS Combat Stress www stress org over the weekend we normally find that their symptoms can be resolved because of the wound healing 11 they can return to school symptom free on Monday Much more importantly however are the comments from the concussed patients Time and again we hear within one or two one hour treatments that the patient had no idea how messed up they were It is as if the fog has cleared as the symptoms resolve The patient is wounded and brain function including judgment are typically impaired Naturally athletes want to get back into the game and are not always the best judges of their level of recovery and ability to return to playing football Objective tests coupled with subjective measures such as IMPACT 12 may offer much more insight into the actual functional recovery Practitioners are familiar with athletes and warriors who game the computer and paper tests purposely obtaining low scores on baseline tests so that post concussion testing fails to demonstrate a significant departure from normal 13 To summarize We are much smarter in 2018 than we were in the 20th century about the Mechanisms of Action both stemming from the negative consequences of concussions and about the remarkable wound healing actions of hyperbaric oxygen The Concussion Protocol must be upgraded to reflect what Dr Denham and hundreds of other HBOT clinics are demonstrating daily that wounds to the brain can be successfully treated and helped to heal using HBOT 14

References 1 2 3 4 5 6 7 8 9 10 11 12 13 14 The Center for Disease Control and Prevention keeps track of Traumatic Brain Injury and Concussion at https www cdc gov traumaticbraininjury get_the_facts html See also https www brainline org article facts about concussionand brain injury For a more thorough look at those who do not recover so well see Carmen Hiploylee et al Longitudinal Study of Postconcussion Syndrome Not Everyone Recovers Journal of Neurotrauma 34 1511 1523 April 15 2017 http bit ly 2Cv7VwA McCrory P et al Br J Sports Med April 28 2017 0 1 10 doi 10 1136 bjsports 2017 097699 http bjsm bmj com content 51 11 838 Ibid p 2 Daphne Denham MD is a general surgeon who began practicing wound care exclusively in 2009 She is now a Diplomate of the American College of Wound Medicine and Surgery demonstrating her extensive knowledge and passion for the art of wound care She declares her interest in a private clinic in Northbrook Illinois Comprehensive Wound Care LLC This article is an outgrowth of her presentation at HBOT 2017 in New Orleans LA Her clinic has now treated 108 acute concussions successfully http comprehensivewoundcare com There is no single Concussion Protocol The Protocols differ in various ways but they all generally speak to some variation on graduated return to play and a series of steps For example the NFL Concussion Protocol calls for these steps in their Return to Practice Protocol 1 Rest and Recovery 2 Light Aerobic Exercise 3 Continued aerobic exercise and strength training 4 Football specific activities and 5 Full football activity The Protocol was upgraded in December 2017 by NFL Head Neck and Spine Committee s Protocols Regarding Diagnosis and Management of Concussion available at https www nytimes com 2017 12 24 sports football nfl concussion protocol html http www espn com nfl story _ id 21864620 nfl making significant changes concussion protocol https www nytimes com interactive 2017 07 25 sports football nfl cte html Amir Hadanny Shai Efrati 2016 Treatment of persistent postconcussion syndrome due to mild traumatic brain injury current status and future directions Expert Review of Neurotherapeutics DOI 10 1080 14737175 2016 1205487 To link to this article http dx doi org 10 1080 14737175 2016 1205487 See Philip B James MD Oxygen and the Brain The Journey of Our Lifetime Best Publishing North Palm Beach FL 2014 See also Harch PG Andrews SR Fogarty EF Lucarini J Van Meter KW Case control study hyperbaric oxygen treatment of mild traumatic brain injury persistent post concussion syndrome and post traumatic stress disorder Med Gas Res 2017 7 3 156 174 http bit ly 2zyFrAr and HBOT2017 11th International Symposium TBI No Need to Die A review of HBOT in Acute Severe Traumatic Brain Injury with an Extension to Acute Concussion and an Update on Chronic Mild TBI Paul G Harch M D http bit ly 2x4tWUf Personal correspondence A Case Report is being prepared Each concussion is unique Variables such as number of previous head injuries and severity are taken into consideration The protocol for each patient s treatment is similarly designed for each patient based on the presentation ImPACT Immediate Post Concussion Assessment and Cognitive Test the most widely used and most cientifically validated computerized concussion management tool available and the newly launched ImPACT Pediatric both the first of their kind to earn FDA clearance See https www impacttest com Jacob Resch PhD ATC et al ImPact Test Retest Reliability Reliably Unreliable J Athl Train 2013 Jul Aug 48 4 506 511 doi 10 4085 1062 6050 48 3 09 In cases of active duty Service Member testing it is common to hear that the stigma of brain injury in a warrior pre conditions study participants to game the system A warrior fears loss of security clearance in certain cases to say nothing of being considered weak or a malingerer Reichert WM ed Indwelling Neural Implants Strategies for Contending with the In Vivo Environment John D Stroncek and W Monty Reichert Chapter 1 Overview of Wound Healing in Different Tissue Types Boca Raton FL CRC Press Taylor Francis 2008 About the Author Robert L Beckman PhD is the Executive Director for TreatNow org and Chief Knowledge Officer Foundation for the Study of Inflammatory Disease He is part of a nationwide Coalition focused on ending the Service Member suicide epidemic through clinical research and healing TBI and PTSD in brain injured Wounded Warriors He is responsible for sustaining a national network of hyperbaric clinics as well as architecting the technology platform for data collection and analysis Dr Beckman has been building knowledge management systems most of his professional career primarily in the Intelligence Community and DOD He has participated in and led revolutionary efforts in strategic planning organizational and management reengineering knowledge based targeting and decision support competitive intelligence war room development targeting and thwarting foreign targeting of US assets fraud detection counter drug counter terror money laundering and organized crime investigations He was a senior member of an elite data integration effort for the counter intelligence CI and counterterrorism CT communities and has built an insider threat prototype for law enforcement entities engaged in CI CT and homeland security He had an extensive academic and publishing career In a former life he taught at the US Naval Academy and was a co owner of consulting and software companies His PhD is in International Relations and he is an expert on nuclear non proliferation and knowledge management He is a former USAF KC 135 pilot and a Vietnam Veteran He lives in Arlington VA is married and has three sons May 2018 AIS Combat Stress 17 www stress org

All the Right Moves The Need for the Timely Use of Hyperbaric Oxygen Therapy for Treating TBI CTE PTSD By Kenneth P Stoller Abstract Background The modern age of hyperbaric medicine began in 1937 however today few know about hyperbaric oxygen s effects on the body and medical conditions outside of diving medicine and wound care centers a serious ethical issue as there are 20 US military veterans committing suicide every day directly related to Traumatic Brain Injury Post Traumatic Stress Disorder The problem is not whether hyperbaric oxygen is effective for treating brain injuries but why the interference in offering this therapy to those who need it Discussion Up against black boxed anti depressants that are not efficacious it should be a no brainer to use a safe off label drug but in the case of military veterans every suicide might be seen as a tremendous cost saving to certain technocrats The unspoken rationale is that if the military were to embrace hyperbaric oxygen as the efficacious therapy that it is then current active troops that have suffered injuries will come forward and seek treatment and benefits for their Traumatic Brain Injuries now that they know there is a viable therapy and in so doing troop strength will be decimated 18 May 2018 AIS Combat Stress www stress org So to attempt to delay the acceptance of hyperbaric oxygen the Department of Defense has funded faux studies claiming low pressure room air to be a placebo or sham and then proclaiming there is no statistical difference between treatment arms and sham or placebo treatment arms With few who understand hyperbaric medicine there is almost no one to call them on this subterfuge and prevarication Many peer reviewed articles have been published in the last decade that demonstrate hyperbaric oxygen is effective in repairing an injured brain even long after that injury took place One of the most notable showed that blast induced brain injured war veterans experienced a 15 point IQ increase p 0 001 Summary Hyperbaric oxygen is an efficacious benign and humanitarian way to affect brain repair but it has not been adopted because it lacks patent protection and has no large corporate sponsors It has also met interference because other agendas are present be they the protection of the status quo myopic budgetary constraints or perceived liability issues Keywords Football TBI Brain Injury HBOT Hyperbaric Oxygen CTE Encephalopathy

Background Hyperbaric oxygen therapy HBOT saturates the body s tissues with oxygen using a pressure vessel HBOT is most often recognized as the treatment for decompression sickness DCS or the bends DCS causes significant neurological injury and post initial injury similar pathophysiology is virtually identical to that caused by trauma Thus oxygen under pressure has been used to treat neurological injuries since 1937 for 75 years No one has found a replacement for or substitute treatment for the bends that works as well as oxygen HBOT results in a 95 acute treatment cure rate for DCS in all of the navies of the world Three currently accepted HBOT indications are for neurological conditions and three are for various kinds of non healing wounds Thus there is more evidence for using HBOT for neurological acute and chronic wound treatment with better clinical outcomes than any other single or combined treatment The good news that combining HBOT with other therapies that help brain injured patients enhances the effect of those treatments and makes these other therapies less costly with creating additional recovery in a given patient Because a patient s future income is directly tied to recovery after injury it is important for policy makers to set maximizing patient recovery as a goal to maximize productivity and tax revenue from each given individual An increase of one half to a 1 atmosphere increase will raise the oxygen levels in plasma 7 12 normal 700 to 1200 Under this increased pressure oxygen acts like a drug and DNA signaling agent This treatment s mechanisms of action simply follow the general gas laws for saturating liquids with a gas similar to the way Coca Cola makes their product No one yet has found a substitute for oxygen in human physiological processes and any injury caused by a lack of oxygen can be expected to benefit with the right oxygen dosage Saturating with oxygen is a safe procedure when all of the correct protocols are followed and significant side effects are extremely rare The history of hyperbaric medicine reaches back to the year 1620 when Drebbel developed a one atmosphere diving bell and 40 years later Boyle joined forces with Gay Lussac to develop the General Gas Law Moving the sands of time to the near present day the modern age of hyperbaric medicine began in 1937 when Behnke and Shaw used a hyperbaric chamber to treat DCS However it was not until 1955 that there was major interest in using hyperbaric oxygenation HBO outside of treating DCS That year Churchill Davidson began to use oxygen therapy in a hyperbaric chamber to treat the damage induced by radiotherapy in cancer patients In 1956 Boerema Holland performed the first reported heart surgery on blue babies in a hyperbaric chamber He became the Father of Hyperbaric Medicine when he treated a woman who had been badly beaten was unconscious and was about to lose her leg This became the first recorded prevention of an amputation with HBOT and the woman did well The next year his famous Life without Blood study was published He referred to the treatment as oxygen drenching 1 May 2018 AIS Combat Stress 19 www stress org

In 1962 Sharp Smith Scotland were first to treat carbon monoxide poisoning by HBOT 1963 Hitchcock testifies before the House Labor Health Education and Welfare committee on the need for hyperbaric chambers in surgery and congress appropriates money for building a score of them 2 1965 Perrins United Kingdom showed the effectiveness of HBOT in osteomyelitis 1965 Japanese researchers treat the first burn patients 1966 Saltzman et al USA showed the effectiveness of HBOT in stroke patients 1970 Boschetty and Cernoch Czechoslovakia used HBOT for multiple sclerosis MS 1971 Lamm France used HBOT for treatment of sudden deafness 1973 Thurston showed that HBOT reduces mortality in myocardial infarction 1976 Hollbach Wasserman determine 1 5 ATA atmospheres absolute maximizes oxygen content and glucose metabolism in the brain 1983 first double blind RCT using HBOT to treat MS 1987 Jain Swiss treats paralysis of stroke with HBOT 1989 the U S Navy discovers that bubbles are gone within 5 min so while DCS is caused by bubbles the secondary injury cascade is the same as in all brain insults 1992 Harch treats the first delayed decompression sickness which lead to the treating dementia pugilistica in boxers cerebral palsy children and autistic children and nearly 50 neurological conditions in 700 patients In 1992 Rockswold USA conducts first double blind RCT showing HBOT reduces mortality in acute traumatic brain injury TBI by 59 the largest single reduction in mortality since the invention of the ambulance In 2002 US Army study confirms Harch s HBOT 20 May 2018 AIS Combat Stress www stress org repairs white matter damage in children with cerebral palsy CP and a Canadian group shows hyperbaric air the original treatment for DCS and Mountain Sickness and HBOT 1 75 effective in treating CP in double blind RT 2005 Stoller USA first child with fetal alcohol syndrome treated 3 and Thom USA finds HBO causes stem cell mobilization 2007 Harch et al USA chronic TBI treated in animal model and in 2009 in military veteran 4 2010 Godman discovers HBOT activates 8 101 genes reducing inflammation and increasing growth and repair hormones In 2011 Stoller treats first retired National Football League NFL player treated for CTE 5 2012 Harch et al demonstrates blast induced post concussion syndrome and posttraumatic stress disorder treatable with HBOT in phase 1 clinical trial 6 The above is not meant to be a comprehensive timeline nor is this a meta analysis of HBOT for various conditions rather an opportunity to understand why a benign yet beneficial therapy has been ignored and even treated with great disdain Bureaucratic concerns have repeatedly trumped medical and scientific evidence For someone with training in decision theory and bureaucratic behavior the problems are very clear Each time in history when a decision was made about the deployment of hyperbaric oxygen therapy starting with Behnke s discovery of oxygen improving the outcomes for DCS bureaucratic concerns over budget constraints trumped science to the determent of the persons those bureaucracies were intended to serve Sometimes it was

you don t know the decision logic one often comes to the conclusion it must be fraud The reports about HBOT s impact on patients have never changed What has changed is we now There is no evidence of pharmaceutical understand the mechanisms of action company interference although such and how vital oxygen is to healthy interference could be inferred after a functioning human metabolism 1983 study was published in the New 8 England Journal of Medicine showing For example a 2002 Canadian study HBOT to benefit patients with multiple found that even room air under sclerosis B H Fischer M D a tenured relatively low pressure 1 3 atmospheres professor at New York University improved the clinical outcomes of became the principal investigator the children in this thirteen year old of a study funded by the National double blind randomized study Ten Multiple Sclerosis MS Society which times more progress was made in is directly funded by pharmaceutical Gross Motor Function GMF during the interests Apparently this society had 2 months of hyperbaric therapy while great difficulty accepting the results of all other therapies were ceased than the work Dr Fischer had completed during the 3 months of follow up with OT PT restarted The editorial in the and multiple revisions were made to Lancet where the article was published weaken the conclusions sufficiently to pointed out that both groups of children satisfy the editors of the New England improved substantially with respect Journal of Medicine In this doubleto GMF speech attention memory blind controlled study of patients and functional skills The Canadian with advanced chronic disabilities government which financed the study Fischer found significant improvement after being pressured by parents of in objective measurements and the children with cerebral palsy falsely treatment effect persisted for at least 1 claimed the pressurized room air was year 7 a placebo and therefore there was no difference between the placebo group For reasons hard to explain to a logical and the group of children receiving mind this study was never followed 100 oxygen While this is sadly up despite the positive results and amusing it kept HBOT from becoming the treatment languished for lack of standard of care for children with CP in financial support and sponsorship Canada and in the United States This Indeed Fischer lost his position and is a gross tragedy and disservice and his chamber was destroyed not using hyperbaric oxygen therapy to treat these children when their brains are It is almost always a bureaucratic plastic and recovery can be dramatic decision that makes no sense to those leaves them as adults with continued who have to enforce it because if high care costs and lost productivity concerns about costs Other times it was incorrect assumptions about the impact of the new science on the system May 2018 AIS Combat Stress 21 www stress org

HBOT has truly been the Cinderella of conventional medicine which means it has been an attractive therapy that has been shown to be efficacious in treating many conditions and yet is treated with derision or ignored at best Because no patent is possible on oxygen or any other element there is no profit to spark a large pharmaceutical interest to prove or promote it Few know about HBOT s effects on the body and medical conditions outside of diving medicine and wound care centers But this has become much more than an issue of lack of marketing and poor public relations This is now a serious ethical issue as there are now 20 US military veterans committing suicide every day directly related to TBI PTSD Suicide losses now exceed combat casualties Even National Football League NFL veterans are starting to commit suicide The clinical trial called the National Brain Injury Rescue and Rehabilitation project15 showed HBOT can virtually eliminate suicidality in this population once they are treated with HBOT while reducing depression by 51 That is a larger and broader effect on depression than anything advertised on television Yet even on the basis of compassionate use it is not possible to get HBOT paid for to treat TBI even though HBOT has more on label indications for brain injury than any other drug or therapy in medicine The issue at hand is not whether HBOT is effective for treating TBI PTSD HBOT is an effective treatment the real issue is why the interference for the time is upon us to expose the obfuscation of this humanitarian therapy literally people are dying because they are not getting into hyperbaric chambers to breathe oxygen Discussion In 2009 and again in 2010 Paul Harch MD Director of the LSU Hyperbaric Medicine Department delivered testimony to both the House and Senate Armed Services Committee reminding them that the epidemic of suicides amongst military veterans was most likely due to cocktail of off label antidepressants they were being prescribed Black Boxed antidepressants None of which are approved for treating TBI Others have delivered this warning as well The exact FDA warning states Antidepressants increased the risk compared to placebo of suicidal thinking and behavior suicidality in children adolescents and young adults in short term studies of major depressive disorder MDD and other psychiatric disorders Anyone considering the use of insert name of antidepressant or any other antidepressant in a child adolescent or young adult must balance this risk with the clinical need While antidepressants are modestly effective in reducing the symptoms of severe depression they increase the brain s susceptibility to future episodes after they have been discontinued This fact contradicts pharmaceutical company sponsored research as antidepressants cause neuronal damage and mature neurons to revert to an immature state both of which may explain why antidepressants also cause neurons to undergo apoptosis programmed death 9 If antidepressants cause death on a micro level then it is much easier to understand why certain patients commit suicide given the human body macro level is made up of these cells So not May 2018 AIS Combat Stress 23 www stress org

only are black boxed off label Selective Serotonin Reuptake Inhibitors SSRI s prescribed to a vulnerable population but they are done so without any regard to an ability tometabolize this class of drugs10 which further exacerbates suicidal behaviors And then it seems SSRIs actually deplete both catecholamine and serotonin 11 which is exactly what isn t in a depressed individual s interest Up against black boxed anti depressants that are not efficacious it should be a no brainer to use a safe off label drug i e oxygen at hyperbaric doses to treat those who have received a TBI now with two decades of use treating various neurological conditions the double blind RCT by Rockswold 12 showing HBOT effective in treating acute severe TBI was published in 1992 decreasing mortality in the acute treatment on severe TBI by 59 the largest reduction in mortality since the invention of the ambulance the use of helicopters in Vietnam for battle casualties and penicillin for infection So what is the problem What does it take to become standard of care As already pointed out HBOT is non patentable Research on non patentable or off patent drugs or with insufficient marketing prospects orphan drugs is funded by nonprofit or charitable organizations only Drugs for which a patent cannot be granted are not being developed and or marketed even when they respond to a public health need Patients pharmacists physicians and other caregivers consequently cannot take advantage of potentially effective treatments they can t even find out about them 24 May 2018 AIS Combat Stress www stress org But while HBOT won t make any entity large profits doesn t it have other monetary incentives For each active duty brain injured solider returned to duty the lifetime savings to the government is 2 6 million dollars and 2 million for each injured service member returned to work or school Between 60 and 80 percent of the veterans participating in the National Brain Injury Rescue and Rehabilitation NBIRR project are returning to work duty or school after receiving HBOT One would think that would move the powers that be to action but it has not and the reason for that may be because all they can see is what an injured veteran will cost should they stay alive That is a serious accusation so to better understand this controversial area on how the Department of Defense DoD handles TBI the NFL provides an example of what takes place on a smaller scale For many decades evidence has linked repetitive traumatic brain injury to long term neurological problems in many sports The NFL as the organizer marketer and face of the most popular sport in the United States in which head trauma is a regular occurrence was aware of the evidence and the risks associated with repetitive traumatic brain injuries and concussions for decades but apparently ignored and worse actively concealed the information from those who participated in organized football at all levels That is now the basis of hundreds of lawsuits So what seems to have taken place is the NFL inserted itself into the scientific research and discussion concerning the relationship between concussions and short term and long term impairment of

the brain After doing so the NFL then intentionally and fraudulently mislead present and former players and all people who reasonably rely upon the NFL s expertise about its own sport regarding the short term and long term risks posed by concussions and head trauma Rather than warn players that they risked permanent brain injury if they returned to play too soon after sustaining a concussion the NFL actively deceived players by misrepresenting to them that concussions did not present serious life altering risks The NFL created the Mild Traumatic Brain Injury Committee the MTBI Committee in 1994 to research and ameliorates the impact of concussions on NFL players Notwithstanding the purported purpose of the MTBI Committee and despite clear medical evidence that on field concussions led directly to brain injuries with tragic results for players at every level of the sport the NFL failed to inform its current and former players of the true risks associated with such head trauma and purposefully misrepresented and or concealed medical evidence on that issue The NFL also stonewalled on an intervention and therapy that could be helping injured players regardless of whether those injuries were acute or chronic The author has firsthand experience dealing with the NFL s 88 Plan in an attempt to get veteran NFL players with dementia HBOT But if a plan member tries to get HBOT using the plan because they have been diagnosed with CTE they will be told CTE does not cause dementia and therefore HBOT which treats CTE will not be a covered benefit Obviously that is irrational but there is often madness behind the reason for not allowing an effective treatment to be utilized by those that need it In the case of military veterans for 20 suicides every day might be seen as a tremendous cost saving to certain decision makers Yet it goes beyond money for if the military embraced HBOT as a viable therapy of TBI PTSD then many many troops who are on active duty with TBI PTSD will come forward once they know there is an efficacious intervention that is recognized by the military In so coming forward troop strength could be decimated This is the fear and this is what has driven faux research funded by the Department of Defense All civilian studies published have been reported as positive even a randomized Israeli study while all the Department of Defense studies have been consistently negative One study had to go so far as to say that it was biologically implausible for compressed room air to have a healing effect on the brain That would be called conformational bias if only this were about bias but this was a deliberate and orchestrated attempt to scuttle HBOT as a recognized therapy for the above reason Yes it is illegal to waste tax payer dollars knowing you are creating pseudo science but that is beyond the scope to this editorial The 88 Plan is designed to assist players who are vested under the Bert Bell Pete Now this is truly misanthropic but no Rozelle NFL Player Retirement Plan and more so than what tobacco corpowho are determined to have dementia rations do and we still tolerate their May 2018 AIS Combat Stress 25 www stress org

malfeasance In the case of the NFL the reason for prevaricating about TBI and potential treatments is just a business decision The exposure of the hit squad of the New Orleans s Saints where there was a bounty put on players from opposing teams is a clear example of what kind of business this is about A great deal of time has been lost by those who believe hyperbaric oxygen is either a placebo sham or should be subjected to placebo controlled studies or want others to believe this But oxygen can never be a placebo HBOT is an FDA approved drug that affects non specific biological repair in fact it is the only non hormonal FDA approved treatment known to repair and regenerate human tissue It does so at a DNA level by activating growth factors and reviving mitochondrial function 13 The beneficial effects of HBOT apply no matter where a wound or injury is located in the body Many peer reviewed articles have been published in the last decade that demonstrates HBOT is effective at repairing an injured brain even long after that injury took place One of the most notable was the article published by Harch et al 6 using only one half of the NBIRR protocol Forty 60 min treatments at 1 5 atmospheres The blast induced TBI war veterans experienced a 15 point IQ increase p 0 001 39 reduction in post concussion symptoms 30 reduction in PTSD symptoms and a 51 decrease in depression This is all consistent with past published reports of HBOT in chronic brain injury including research by the US Army on brain injured children 14 26 May 2018 AIS Combat Stress www stress org The first battle casualty to be treated with HBOT 1 5 and one of the few to be treated was General Patt Maney retired for his blast induced brain injury in Afghanistan His treatment was ordered after 9 months of therapy at Walter Reed had shown minimal improvement As a result of his injuries he was non functional and unable to return to his job let alone redeploy back to Afghanistan After HBOT treatment he was discharged from Walter Reed and returned to his civilian job as a Florida state judge He received treatment from George Washington University Medical Center at the Tricare Reimbursement rate of 250 per treatment Counting lost time and hospital costs his months at Walter Reed making no progress the DoD spent 400 950 with a permanent disability loss to the service of 1 3 million Had he received HBOT at 1 5 atmospheres earlier he would have been able to remain on active duty a savings of 1 3 million but more importantly the 5 months of recovery once he began receiving HBOT 1 5 cost 133 650 a savings to the government of 287 300 No other patients were treated at the Walter Reed s brain injury center despite the General s remarkable recovery that everyone on the staff witnessed The 20 000 for his hyperbaric medical treatment was 12 000 less than what a RAND report states the annual ongoing costs per year of the current treatments for mild TBI is 16 and that is a lot of SSRI s

Since every working person represents 1 million in tax revenue over their working life to the government that government should be interested in and foster payment for biological repair of brain injury Thus every brain injured veteran all 700 000 of them at a cost of 60 000 per year to the economy every year in increased costs and lost productivity is a 42 billion drain on the economy Treated they immediately set about doing what young people do they begin to form families and create the next American generation Injured they are unable to do so and it is highly likely that these untreated brain injuries are a major cause of our nation s economic challenges But bureaucracies do not think logically their ability to think laterally is limited When Medicare approved HBOT to treat diabetic foot ulcers at the end of 2002 they only made it available for Wagner III and IV lesions osteomyelitis and gangrene so afraid they were of budgetary constraints HBOT prevents 75 of major limb amputations in Wagner III and IV ulcers but if they had included Wagner II lesions HBOT would be preventing 88 of amputations So the result is there are a lot of unnecessary amputations not because of bad science but because technocrats were afraid of having a short term budget problem Summary HBOT is an efficacious benign and humanitarian way to affect brain repair but it has not been adopted because it lacks patent protection and has no large corporate sponsors It has also met interference because other agendas are present be they the protection of the status quo myopic budgetary constraints or perceived liability issues after all when you treat TBI directly the way that HBOT does the problems creating those TBI s in the first place are harder to ignore and the unconscious way those problems have been dealt with are harder to deny It brings the true cost and repercussions of war to the fore and football itself after all is just a form of organized war This perspective should it be adopted by the general public will be a catalyst for change So whether that means changing the way football is played to not allowing our leaders to guide us into unending military forays for the sack of warprofiting run amuck all of these sub terrain issues come into play when a very straight forward and effective therapy tries to assert itself hence the resistance both on a conscious and subconscious level Veterans are considered a threat by the security apparatus in the USA Is someone thinking that it is better to let 20 potential threats kill themselves every day This is a dark rabbit hole to go down but the human cost of war is very much a deep rabbit hole and one that many want to keep hidden Someone is making money when someone else bleeds to death from a cluster bomb It is that black white someone is making money giving dangerous SSRIs for treating TBI Technocrats rotating between corporation and state are the last people you want making medical decisions but that is exactly who has been making medical decisions These are all things to be looked at when asking the question why a therapy like HBOT is being suppressed If the man May 2018 AIS Combat Stress 27 www stress org

on the street understood why certain medical therapies are not available while other dangerous and nonefficacious therapies are favored then change would be demanded Right now the man on the street is being kept in the dark about money driving medical decisions to the extent it is today including the suppression of medical knowledge if it will interfere with the flow of that money or some other irrational fear such as troop strength decimation if soldiers were officially offered HBOT by the military to treat their TBI PTSD Appeared in Medical Gas Research 2015 5 7 Abbreviations TBI Traumatic brain injury ATA Atmospheres absolute CTE Chronic traumatic encephalopathy PTSD Post traumatic stress disorder HBOT Hyperbaric oxygen therapy DCS Decompression sickness HBO Hyperbaric oxygen CP Cerebral palsy NFL National Football League GMF Gross motor function MTBIC Mild Traumatic Brain Injury Committee NBIRR National Brain Injury Rescue and Rehabilitation DoD Department of Defense RCT Randomized Controlled Trial FDA Federal Drug Administration SSRI Selective Serotonin Reuptake Inhibitor MS Multiple sclerosis Competing interests Financial competing interests Dr Stoller practices hyperbaric medicine in New Mexico and California Non financial competing interests Dr Stoller was president of the International Hyperbaric Medical Association for over a decade References 1 2 3 4 5 28 Trimble Vance In hyperbaric oxygen the uncertain miracle the little known maverick medical treatment which has saved the lives of thousands of people First edition Unknown Binding 1974 83 84 U S House of Representatives Labor HEW Public Witness Testimony 1963 p 274 9 Stoller KP Quantification of neurocognitive changes before during and after hyperbaric oxygen therapy in a case of fetal alcohol syndrome Pediatrics 2005 116 e586 91 Harch PG Fogarty EF Staab PF van Meter K Low pressure hyperbaric oxygen therapy and SPECT brain imaging in the treatment of blast induced chronic traumatic brain injury post concussion syndrome and post traumatic stress disorder a case report Cases J 2009 2 6538 Stoller KP Hyperbaric oxygen therapy 1 5 ATA in treating sports related TBI CTE two case reports Med Gas Res 2011 1 17 May 2018 AIS Combat Stress www stress org 6 7 8 9 10 11 12 13 14 15 16 Harch PG Andrews SR Fogarty EF Amen D Pezzullo JC Lucarini J et al A phase I study of low pressure hyperbaric oxygen therapy for blast induced post concussion syndrome and post traumatic stress disorder J Neurotrauma 2012 29 1 168 85 Fischer BH Marks M Reich T Hyperbaric oxygen treatment of multiple sclerosis a randomized placebo controlled double blind study New Eng J Med 1983 308 181 6 Carole S Larivee S Richard E Marois P Hyperbaric oxygenation therapy in the treatment of cerebral palsy a review and comparison to currently accepted therapies J Am Phys Surg 2007 12 4 Andrews PW Thomson Jr JA Amstadter A Neale MC Primum non nocere an evolutionary analysis of whether antidepressants do more harm than good Front Psychol 2012 3 117 Epub 2012 Apr 24 Lucire Y Crotty C Antidepressant induced akathisia related homicides associated with diminishing mutations in metabolizing genes of the CYP450 family Pharmacogenomics Personalized Med 2011 2011 4 65 81 Delgado PL Moreno FA Onate L Sequential catecholamine and serotonin depletion in mirtazapine treated depressed patients Int J Neuropsychopharmacol 2002 5 63 6 Rockswold GL Ford SE Anderson DC Bergman A Sherman RE The results of a prospective randomized trial for treatment of severely brain injured patients with hyperbaric oxygen J Neurosurg 1992 76 929 34 Gutsaeva DR Suliman HB Carraway MS Oxygeninduced mitochondrial biogenesis in the rat hippocampus Neuroscience 2006 137 2 493 504 Waalkes P Fitzpatrick DR Stankus S Topolski R Adjunctive HBO treatment of children with cerebral anoxic injury Army Med Dept J 2002 April June 13 21 http clinicaltrials gov ct2 show NCT01105962 RAND Report Invisible Wounds of War Psychological and Cognitive Injuries Consequences and Services to Assist Recovery Tanielian Terri Jaycox Lisa April 2008 page xxii xxiii Two year costs within the first two years the service member returns home PTSD 5 904 to 10 298 depending on whether we count the lives lost to suicide Two year costs for major depression 15 461 25 757 co morbid PTSD and major depression 12 427 to 16 884 One year costs for traumatic brain injury diagnosis 25 572 to 30 730 in 2005 for mild cases 27 259 to 32 759 in 2007 dollars and 252 251 to 383 221 for moderate or severe cases 268 902 to 408 519 in 2007 dollars These costs largely treating symptoms continue to have out year costs and out year consequences in terms of disability payments inability to work etc Given that the HBOT ONE TIME cost for service members who need all 80 treatments averages 16 000 at Medicare Reimbursement rates for a 1 h treatment HBOT alone and even HBOT in conjunction with other treatments is very cost effective If provided acutely within hours of injury the treatment is even more effective and massively more cost effective About the Author Ken Stoller is an Adjunct Assistant Professor of the AT Still University School of Medicine a Fellow of the American College of Hyperbaric Medicine and a principal investigator of the NBIRR clinical trial

A Discussion of the Use of Hyperbaric Oxygen Therapy as a Primary and Adjunctive Treatment Modality for TBI Patients By John Hughes D O and James Lyons Weiler PhD Traditional Approach Most treatments for chronically debilitated traumatic brain injury TBI patients have involved pharmaceutical drugs occupational and physical rehabilitation speech therapy and cognitive maintenance Many patients resign to merely manage their condition and gain very little improvement or begin a slow decline of cognitive or motor function It is true that some mild traumatic brain injury mTBI patients recover to a degree using mainline therapies but this recovery time can take 7 or more years with significant loss of cognition learning and psychological affect during that time of recovery TBI Therapy s Approach Using TBI Therapy s standard of care protocols patients at Dr John Hughes TBI treatment facilities optimally receive HBOT therapies before and after intranasal and IV infusions as well as with other healing modalities such as cranial osteopathic treatment IV nutrition and hydration and other adjunctive therapies to aid in neuro regeneration The procedures and protocols used by Dr Hughes Colorado based TBI treatment clinic are patent pending uniquely sophisticated presently scarce and at the forefront of neuroregenerative medicine The brain is in a metabolic crisis in a concussion Robert Cantu MD 2013 It is this understanding of what happens in a traumatic brain injury even a mild TBI that forms the basis of TBI Therapy s unique treatment protocols Most mainstream therapies just work to treat the symptoms of a TBI using pharmaceutical drugs speech therapies vision therapy and physical therapy While these therapies have importance most TBI patients hit a plateau in terms of treatment benefits These TBI patients continue to suffer from post concussive symptoms sound and light hypersensitivity sleep disturbances memory loss loss of concentration headache pain loss of libido loss of decision making mood changes loss of motivation and inability to handle daily stressors Many TBI patients suffer so significantly from these conditions that they may not recover They eventually lose hope sometimes even at the expense of families and their own lives Most all of the above post concussive symptoms of TBI are related to one major disturbance to the brain in Dr Robert Cantu s 2013 words a metabolic crisis Standard of care for concussion includes rest to avoid perpetuation of May 2018 AIS Combat Stress 29 www stress org

a crisis of gliosis Gunther Queen 2013 A TBI is like a bomb that goes off in the biochemistry lab of the brain all the experiments are in disarray and it will require tremendous energy to get the lab back in working order once again Pressurized oxygen is analogous to plugging a computer the hardware of the brain into a wall socket with enough voltage to begin its self healing This energy must come from somewhere The challenge is however that the brain even at a resting state is always using energy It can never fully rest because it is always working or healing itself The brain cannot be placed on the couch for a week with an icepack on it to heal like a sprained ankle We do not need our ankle to work every day but we do need the brain to work at all times in order to sustain life Traumatic brain injuries TBIs are frequent among Soldiers football players and victims of assault Studies of the impact of HBOT on TBIs have been inconsistent One small study has had a dampening impact on the medical community s enthusiasm for clinical adoption of HBOT for the treatment of TBI with only 50 participants per group the study was likely underpowered for small but important positive or negative effects of HBOT Wolf Cifu Baugh Carne Profenna 2012 A full review of studies and their limitations have been provided by Eve Steele Sanberg Borlongan 2016 including extensive consideration of the difficulty in obtaining a control group that is free of confounding influences of the procedure itself The difficulty with this explanation however is that other uses of HBOT show clear benefits in spite of this limitation which applies to all randomized clinical trials with obvious intervention A randomized clinical trial of patients with long standing TBIs by Boussi Gross et al 2013 found improved cognition and quality of life measures during the HBOT treatment period Clinical improvements can be seen at exposures of 1 1 atmospheres ATA or less therefore teasing the placebo effect out is challenging Outcomes should be studied for correlation along the axes of the number of treatment sessions completed So how can a working human TBI brain consuming at least 20 percent of the oxygen inhaled by the lungs recover If we cannot rest the brain the most important way to heal the TBI brain in a metabolic crisis is to give it more energy The primary way to get energy to the brain is with more oxygen Will breathing a few extra liters of O2 per minute be enough to help these TBI patients Not really The brain requires a significantly higher dose of oxygen to heal from injury It is hypothesized that the brain needs a continuous 3 to 4 times its regular inhaled dose of oxygen to begin healing from TBI depending on its severity The only way to significantly increase oxygen in the brain is with a medical grade hyperbaric chamber providing 8 to12 times as much oxygen or home hyperbaric chamber 5 to 6 times as much oxygen as normally inhaled when the patient is at rest With enough oxygen the brain can begin repairing its hardware 30 May 2018 AIS Combat Stress www stress org Hyperbaric Oxygen Therapy and TBI

the duration of each session and the dosage in terms of atmospheres and oxygen concentrations Studies of this type should also be restricted to clearly homogeneous clinical populations which is extremely difficult in the heterogeneous population of patients with TBI which can involve any number of regions of the brain differing in severity and degree of injury A priori power calculations should be conducted to ensure that negative results are not due to small sample sizes Studies of hyperbaric oxygen therapy HBOT for treatment of traumatic brain injuries TBIs vary in results in part due to the heterogeneity in the type of scope of TBI cases and challenges in isolating the effect of pressure because subjects are prone to potential placebo effect Well designed dose escalation studies of both pressure and oxygen concentrations compared to low pressure control would provide knowledge on dose efficacy Other variables include treatment intensity frequency and length of dives and duration length of treatment period in months These sources of variation are usually well controlled within but not between HBOT studies Meaningful clinical outcome variables in mild traumatic brain injuries mTBIs include localization and extent of injury In a landmark study Harch et al 2012 applied hyperbaric oxygen therapy to individuals who had developed PTSD after blast induced post concussions All subjects in the study were given half of the clinical protocol for the study forty 60 minute treatments at 1 5 ATA Those receiving the full dose of HBOT showed a 14 8 point increase in intelligence quotient p

reported sustained improvement six months after treatment with functional improvements in cognition 39 percent physical well being 45 percent emotional areas 96 percent and significant reductions in anger issues There is concern that improvements in both treatment and placebo groups demonstrated the Hawthorne effect the study shows that significance of difference between groups outcomes which were not likely due to chance can be obtained in a manner that obliviates the concern of the placebo effect when the effects are unidirectional A small clinical study of concussion patients reported cognitive improvements in both groups treated at 1 2 ATA air or 2 4 ATA 100 percent oxygen There were in fact no differences in cognitive improvements between the two groups Wolf et al 2015 either 1 or 8 hours after TBI in rats led to reduced cerebral infarction microglial activation TNF alpha expression and neuronal apoptosis all factors that can influence inflammatory and neurotrophic processes in the human brain More recently Tal Hadanny Sasson Suzin Efrati 2017 found that HBOT stimulated both angiogenesis and nerve fiber regeneration in TBI patients including improvements in memory executive functions information processing speed and global cognitive scores post treatment compared to pre treatment There is solid evidence that hyperbaric oxygen therapy provides cognitive and neuropsychological improvements for TBI patients However the appropriate dosing of hyperbaric oxygen required to gain significant benefits for these TBI patients has not yet been determined It is hypothesized that patients may gain more benefits from using HBOT at milder pressures from 1 1 to 1 3 ATA than at higher pressures of 1 5 2 4 ATA These milder HBOT treatments at pressures from 1 1 to 1 3 ATA can be performed daily in FDA approved home hyperbaric chambers over extended time periods 3 to 12 months at significantly lower cost per treatment than in an outpatient medical facility References In a study highly relevant for those using off label neurological applications Lin et al 2012 found that HBOT 32 May 2018 AIS Combat Stress www stress org Further research backing the use of hyperbaric medicine for the treatment of TBI patients is available online see https tbitherapy com tbi protocol references 1 2 3 4 5 6 7 Boussi Gross R Golan H Fishlev G Bechor Y Volkov O Bergan J Efrati S 2013 Hyperbaric oxygen therapy can improve post concussion syndrome years after mild traumatic brain injury randomized prospective trial PloS one 8 11 e79995 Eve D J Steele M R Sanberg P R Borlongan C V 2016 Hyperbaric oxygen therapy as a potential treatment for post traumatic stress disorder associated with traumatic brain injury Neuropsychiatric disease and treatment 12 2689 Gunther N Queen E 2013 What Physical and Cognitive Rest Really Mean After a Concussion Brainline Retrieved from http www brainline org content multimedia php id 9022 Harch P G Andrews S R Fogarty E F Amen D Pezzullo J C Lucarini J Van Meter K W 2012 A phase I study of low pressure hyperbaric oxygen therapy for blast induced post concussion syndrome and post traumatic stress disorder Journal of Neurotrauma 29 1 168 185 Lin K C Niu K C Tsai K J Kuo J R Wang L C Chio C C Chang C P 2012 Attenuating inflammation but stimulating both angiogenesis and neurogenesis using hyperbaric oxygen in rats with traumatic brain injury Journal of Trauma and Acute Care Surgery 72 3 650 659 Tal S Hadanny A Sasson E Suzin G Efrati S 2017 Hyperbaric oxygen therapy can induce angiogenesis and regeneration of nerve fibers in traumatic brain injury patients Frontiers in Human Neuroscience 11 Wolf G Cifu D Baugh L Carne W Profenna L 2012 The effect of hyperbaric oxygen on symptoms after mild traumatic brain injury Journal of Neurotrauma 29 17 2606 2612

About the Authors Dr James Lyons Weiler PhD is the CEO and President of The Institute for Pure and Applied Knowledge With over 50 publications in major research journals Dr Lyons Weiler has designed and analyzed the data from over 100 basic translational and clinical studies His current research interests include finding commonalities in the mechanisms of physical and chemical brain injury especially focused on neurotoxic metals and finding treatment options from traumatic brain injury that might translate over to pediatric neurodevelopmental disorders An expert in genetics genomics proteomics systems biology research design and bioinformatics he has authored three books He is alumnus of SUNY Oswego BA Biology Ohio State University MSc Zoology and The University of Nevada Reno PhD Ecology Evolution and Conservation Biology He was awarded an NIH National Service Award Post doctoral Fellowship and a US DOE AP Sloan Foundation Fellowship He has served on the faculty of the Departments of Biology UMASS Lowell Department of Pathology Department of Biomedical Informatics University of Pittsburgh and served as Full Faculty at the University of Pittsburgh Cancer Institute prior to creating and assuming the role of Senior Research Scientist and Scientific Director of the Genomics and Proteomics Core Laboratory Bioinformatics Analysis Core at the University of Pittsburgh Visit http ipaknowledge org for more information and for updates on current research publications Dr John Hughes D O practices osteopathic integrative and regenerative medicine His clinic Aspen Integrative Medicine Inc provides the latest innovations in modern and natural medical care in Basalt and Aspen Colorado Dr Hughes graduated from the Arizona College of Osteopathic Medicine in 2007 and trained in family practice at the University of Arizona He has specialized in integrative and regenerative medicine prolotherapy platelet rich plasma PRP therapy and osteopathic manual medicine since 2009 In 2013 Dr Hughes gained an interest in the treatment of traumatic brain injuries after incidentally treating a patient with a severe TBI using a regenerative medicine protocol of hyperbaric oxygen an intranasal PRP insulin nutrient cocktail IV nutrition cranial therapy and autologous stem cells In 2014 Dr Hughes founded TBI Therapy LLC a TBI clinic in Colorado focused on the development of specific protocols for mild to severe brain injury patients In 2016 Dr Hughes applied for a patent status for his TBI Therapy Protocol and intranasal cocktail formulation His TBI rehabilitation work has been recognized by the patients from the US military the regenerative medical community and brain injury specialists Dr Hughes sees TBI patients from around the US and the world on a regular basis in Basalt Colorado and Lafayette Colorado Read more about the innovative work of Dr Hughes at tbitherapy com May 2018 AIS Combat Stress 33 www stress org

7815 NW 23rd St Bethany OK 73008 405 999 0965 http www patriotclinics com February 22 2018 CONTACT Dr William A Duncan President Patriot Clinics Inc 405 999 0965 williamaduncan msn com FOR IMMEDIATE RELEASE In fact Department of Defense HBOT medicine could easily repair a braininjured Veteran while still in service with an 80 treatment protocol while still in the service at a cost of 480 The VA can do the job for about 4 800 while civilian treatment facilities increase the cost to 25 000 Instead VA and DoD medicine are spending about 32 000 per year on ineffective medications that have significant negative side effects Each untreated war veteran with Traumatic Brain Injury TBI or Post Traumatic Stress Disorder PTSD is costing the economy federal state and local governments about 60 000 per year if they are not homeless That almost doubles if they are homeless Today we are losing more Veterans to suicide than we lost in combat in theater and under fire in World War I 120 000 suicides to 116 000 deaths in WWI Sadly this is because one the biggest myths in medicine is there is no treatment for brain injury There are over one million current war Soldiers whose lives have been significantly When hyperbaric medicine becomes affected by these injuries the standard of care society will save billions as lives are restored The cost The truth is we have two major treat of oxygen treatment is exponenments for brain injury in the United tially less than the millions it costs our state each year to leave our States Hyperbaric Oxygen Therapy wounded Veterans untreated or HBOT has been in use since 1937 treated ineffectively Dr William and Cognitive Rehabilitation has been A Duncan President of Patriot utilized since 1980 Clinics Inc explained Experience 34 May 2018 AIS Combat Stress www stress org

shows that if you take 100 Veterans in transitional living and give them hyperbaric medicine half will be back to work within 40 treatments and 80 percent will be back to work after completing HBOT treatment Patriot Clinics in Oklahoma City was born from the fact that veterans were not being effectively treated The non profit corporation was set up in 2014 and is modeled after 90 charity hyperbaric centers serving patients in Great Britain The treatment regimen follows the National Brain Injury Rescue and Rehabilitation Project s HBOT 1 5 ATA 99 O2 protocol Patriot Clinics focus on effective medical care and interventional hyperbaric medicine Recently a Church Pastor called that one of his parishioners was in trouble Joe is a young Afghanistan War Veteran and in serious danger of suicide He came to Patriot Clinics and within 3 hours had a BrainMaster QEEG brain scan took the Automated Neuropsych Assessment Metric ANAM had a Dive Physical and a hyperbaric treatment He received a second treatment that night an additional treatment in the morning and acupuncture for his PTSD and massage therapy for his injuries aggravated by blasts overseas The improvements in mood and outlook were immediate and welcomed For over 500 Veterans like Joe over the past four years doctors practitioners techs and volunteers at Patriot Clinic delivered over 4 million worth of treatments in 14 000 hyperbaric dives The clinic also delivered other therapies to help those with PTSD such as 3 000 chiropractic adjustments and 1 000 acupuncture treatments for PTSD and pain This care replicates research results from Louisiana State University Oklahoma State University and the country of Israel where people gained 15 IQ points the difference between a high school dropout and a college graduate a 39 reduction in postconcussion syndrome sleep disorder light sensitivity executive function difficulty 30 reduction in PTSD the largest percentage of any published treatment a 51 reduction in depression and a 96 improvement in emotional control Clinically significant improvement can be expected in every patient who follows the protocol Suicidality is reduced as Dr Harch recently published in his study At Patriot Clinics we simply got tired of an attitude of throw away people and managed care that treats only symptoms Instead we have implemented effective care using therapies that actually work to improve a patient s medical condition Duncan added Patriot Clinics works to make hyperbaric treatment accessible to everyone Combat veterans are often treated using donated funds Civilians pay based upon income and family resources Third parties like workers compensation and automobile May 2018 AIS Combat Stress 35 www stress org

carriers will be billed when appropriate Duncan explained the clinic is able to deliver these treatments for veterans and others who cannot pay because of charity donations We have to raise as much as 10 000 each month to accomplish our goals We ask all patients that experience success in treatment to donate at least 10 for the future treatment for vets Patriot Medical Foundation at www PatriotMedicalFdn org raises money for clinics across the nation to fund this kind of care We treated over 500 at our clinic Duncan noted Many injured Veterans have lost hope of ever recovering After years of living with shattered life symptoms they are more than pleased when the headaches disappear normal sleep returns the anger issues subside and the ability to think reason and have healthier family relationships returns volunteer Office Manager Stanley Maness explained Volunteers work every week to keep the clinic Patriot s facebook page flows with operating gratitude from those who have experienced the outcomes first hand The clinic also treats people with Too many Veterans are committing cancer diabetic wound care and Lyme suicide HBOT therapy is healing the disease It offers pre and post surgery hidden wounds that drugs cannot cure treatments resulting in improved and the typical medical community does not understand Patriot Clinics healing reduced complications and has seen substantial measurable pain and greater patient satisfaction successes despite operating on a compared to surgery without the shoestring budget of donations You benefit of pre and post hyperbaric will not find a harder working group treatment The clinic has also begun of volunteers than at Patriot Clinics treating car accident and workers most of whom have been successfully compensation patients The addition treated with HBOT returning to of the BrainMaster QEEG system give them a sense of pride in being able to the clinic the ability to do brain scans give back for patients in real time which greatly improves the ability to help acutely Two thirds of patients were Vets injured patients referred by Vet service organizations and their family members One third With hyperbaric medicine and the were civilians whose donations contributed toward the treatment for other integrated therapies Patriot Vets At over 30 treatments per day the Clinics uses tools to do better for clinic is one of the busiest hyperbaric those who served their country Learn centers in the nation To put this into more or donate to help heal our perspective since 2008 only 2 000 warriors at www patriotclinics com or Vets have been treated nationwide www patriotmedicalfdn org 36 May 2018 AIS Combat Stress www stress org

Post injury Pre Hyperbaric Oxygen Therapy HBOT Senior Force Recon Marine Survived Fallujah and over 7 concussions Constant ER visits with migraines and 6 different pills for 13 years until wife worried about suicide talk found HBOT 21 year Marine SpecOps Multiple missions Blown up by 2 IEDs serious parachute accident over 1 200 breaching exposures Constant ringing in ears eye pain and migraines Ditto for his unit Recently retired SEAL Over 2 dozen tours and dozens of concussive events IEDs friendly fire breaching Reports dozens of his unit in equally bad shape hollowed out and needing help AFSOC Silver Star Winner In police custody for road rage Rescued from jail by HBOT physician who provided 80 dives on pro bono basis Post HBOT Refused HBOT by his unit Cmdr approved 30 day leave 40 dives in one month from pro bono civilian clinic Now instructor reports no longer the anger gunny HBOT made available to this warrior and his Cmdr 40 dives each Both report lives drastically changed for the better Results are consistent across other fellow Marines misdiagnosed with only PTSD Treated quietly off base over 40 dives after being told HBOT could ruin his career Feels he has been given new lease on life Advocating for forward deployed HBOT chambers and HBOT treatment as standard of care for combat Returned to active duty Further tours and decorations Ordered not to talk about his treatment with HBOT 17 year Special Operator Breacher Multiple tours innumerable exposures Multiple failed marriages Suicidal Subjected to 3 months intensive care in PREP program Regressed to original state and psychotropic meds within 2 months Green Beret Foundation provided support for HBOT treatment with Dr Zant After 40 dives reports off all medicines migraines gone wife and children ecstatic Still on active duty and fully functional May 2018 AIS Combat Stress 39 www stress org

Post injury Pre Hyperbaric Oxygen Therapy HBOT Margaux Mange Iraq Veteran Major head injury brain surgery failed marriage depression suicidal Told to get used to her new normal Nothing more could be done MAJ Ben Richards USA ret Iraq Veteran and 3 TBI survivor Misdiagnosed TBI for 5 years Genius IQ reduced to full disability DVBIC and Army had him on multiple drugs Judge Patt Maney BG USA ret IED survivor Afghanistan 15 months without progress at Walter Reed Army Medical Center First HBOT patient in 2006 Over 120 dives CPT Matt Smotherman OK Natl Guard Law student before being blown up 3 times in Iraq Could not fill out paperwork read sleep focus Drugged out of pain and into stupor GnySgt Rotenberry USMC ret IED in 2011 Pain fatigue depression headaches withdrawn Hyperactivity Late TBI diagnosis 15 daily pills Medically separated 40 May 2018 AIS Combat Stress www stress org Post HBOT 90 dives Hiked to South Pole with Prince Harry hiked across US and up Denali Essentially drug and symptom free 4 time Wounded Warrior Olympics champion after being relegated to life on welfare Poster child for HBOT Genius IQ returned after being told his new normal was all he could ever expect In PhD program and spokesman nationwide for HBOT Returned to elected Judgeship Instituted Veteran Courts in FL and lectures nationwide on need for HBOT and special treatment of Veterans with brain injury Finished law school after 80 dives Promoted to CPT after being told he would have to retire on full disability Commended personally by POTUS for efforts in OK tornado relief Wife found Dr Harch in LA Donations paid for 40 dives Employed at DHS as dog handler Life returning to old normal Getting 40 more dives for lingering pain and some headaches

Post injury Pre Hyperbaric Oxygen Therapy HBOT NFL legend Joe Namath 5 big concussions Memory loss balance brain fog Awash in pain and suicides and CTE of friends Steve Bowman Alabama NY Giants Concussion drove him to early retirement Life and wife rapidly going downhill Progressive dementia like symptoms Joe Delamielleure Buffalo Bills HOF 2003 Memory brain fog pain 80 dives after plea to help him in Charlotte NC NFL Pro Bill Romanowsky 20 concussions over 16 years 80 dives to fight memory loss headaches physical pain Marv Fleming Packers Dolphins Patriots Memory loss balance headaches physical pain Post HBOT Poster child for HBOT Lectures on his recovery and deterioration of his friends Strong advocate for HBOT and against delay and denial of concussions in NFL Went from inability to walk think remember sleep to mobile husband Wife cries gratefully I got my man back Spokesman for HBOT Lectures to NFL Players and veterans about his recovery and need for HBOT in every locker room He feels HBOT should be standardof care for concussions Continues a dive a month and proselytizes for HBOT at Super Bowl 50 Spokesman at SB50 for HBOT Works to encourage other braindamaged players to get HBOT and demand treatment May 2018 AIS Combat Stress 41 www stress org

Introduction Head injury is the leading cause of death and disability among young adults 1 In Indonesia head injuries account for almost half of all deaths caused by trauma 2 4 The symptoms of head injury range from mild to severe depending on the extent of brain damage 5 Patients with head injuries often experience considerable cognitive behavioral and communication impairment 6 These patients deserve prompt effective treatments that are not only lifesaving but also preserve their brain function Hyperbaric oxygen therapy HBOT involves the inhalation of 100 oxygen at a higher pressure than the atmospheric standard Patients inhale 100 oxygen and pressure increases gradually to 2 3 absolute atmosphere ATA 7 10 HBOT has become popular in the field of neurology because it inhibits apoptosis suppresses inflammation and protects the integrity of the blood brain barrier BBB in addition to stimulating angiogenesis and neurogenesis The neuroprotective effects of HBOT are most effective during the acute phase which is the first 24 hours post head injury 11 The anti neuroinflammatory properties of HBOT are at least partially exerted by suppressing matrix metallopeptidase 9 MMP 9 expression 12 MMP 9 is a Zn dependent endopeptidase enzyme that maintains and remodels the extracellular matrix ECM MMP 9 is produced by microglia neurons oligodendrocytes astrocytes and the vascular endothelium 13 In chronic brain injury HBOT increases cerebral blood flow improves any related neuropsychological disorders and promotes neurophysiological and electrophysiological recovery 14 As such this therapy improves the quality of life in patients with post concussion syndrome and prevents its progression to more advanced stages 15 Taken together these data suggest that HBOT represents a promising therapeutic modality for various forms of head injury The persistence of symptoms associated with mild traumatic brain injury TBI is referred to as post concussion syndrome The majority of patients with post concussion syndrome recover within three to six months 16 In most clinical trials on the survey based patient progress was evaluated by neuropsychological examination such as the Rivermead Post Concussion Symptoms Questionnaire RPQ The RPQ is one of the most commonly used instruments for determining the severity of the symptoms caused by mild to moderate TBI Individual item scores reflect the presence and severity of post concussion symptoms that overlap with a wide range of conditions e g pain fatigue and mental health conditions including depression The RPQ is divided into two groups RPQ 3 and RPQ 13 RPQ 3 consists of three initial symptoms such as headaches feelings of dizziness nausea and or vomiting The RPQ 13 is a progression of these initial symptoms such as noise sensitivity sleep disturbance fatigue feeling irritable feeling depressed or impatient forgetfulness poor concentration taking longer to think blurred vision light sensitivity double vision and restlessness 17 Questionnaires are repeatedly administered to monitor the patient s progress over time and promptly identify changes in the severity of symptoms The recovery process usually requires three to six months of conservative treatment to resolve the symptoms 18 However these conservative treatments are slow and sometimes ineffective leaving patients with life long symptoms ranging from headaches to impaired cognitive function Therefore the present study aims to determine whether and how HBOT accelerates post concussion syndrome recovery by analyzing RPQ scores and MMP 9 levels respectively We hypothesize that HBOT can improve post concussion symptoms by decreasing MMP 9 levels in patients with TBI undergoing HBOT compared with traditional therapeutic approaches Methods Study design This study used a randomized controlled trial design to assess levels of MMP 9 and Rivermead Post Concussion Symptoms Questionnaire RPQ scores at pre treatment baseline and post treatment at weeks one three and five Ethical approval The study was conducted after obtaining approval from the research ethics committee of Prof Dr R D Kandou Hospital Manado Code number 125 EC KEPK KANDOU XII 2020 This study was registered with the Research Registry with a registration number no 6465 on January 17 2021 Written informed consent was obtained from all the participants The work was carried out in line with the Consolidated Standards of Reporting Trials CONSORT guidelines 19 20 Population and sample This study included 20 patients with mild TBI The patients were randomly divided according to treatment into a control group and an experimental group The experimental group received HBOT in addition to the Advanced Trauma Life Support ATLS protocol while the control group only received the latter In accordance with randomized controlled trial

design the patients in the control group were so selected because they refused to receive HBOT The study was carried out at Dr R D Kandou Hospital Manado North Sulawesi Indonesia Procedure All patients received standard mild TBI treatment according to the ATLS protocol including head CT scans using computed tomography SOMATOM Scope Siemens Healthineers AG Erlangen Germany to identify any brain abnormalities The HBOT group also received 60 minute HBOT sessions breathing in 100 oxygen at 2 3 ATA three times during this study at weeks one three and five 21 RPQ score Items in the questionnaires are divided into two groups The first group RPQ 3 consists of three items and the second group RPQ 13 consists of 13 subsequent items 17 18 22 The total score for the RPQ 3 items range from 0 12 and reflect the baseline symptom pool of post concussion symptoms A higher RPQ 3 score requires reassessment and close monitoring 23 MMP 9 measurements Prior to initiating treatment in both groups the RPQ was administered blood samples were collected for serum MMP 9 assays and head CT scans were performed to identify any brain abnormalities such as subarachnoid hemorrhage SAH epidural hematoma EDH subdural hemorrhage SDH and intracerebral hemorrhage ICH MMP 9 levels were measured in the Biomolecular and Immunology Laboratory of the Faculty of Medicine at Sam Ratulangi University in Manado Indonesia after every HBOT session for both the HBOT and control groups At the end of week 1 3 and 5 all patients were reevaluated for serum MMP 9 levels and RPQ scores An enzyme linked immunosorbent assay ELISA was used to quantify MMP 9 levels in ng mL as described in the MMP9 Human ELISA Kit protocol Invitrogen Corporation Carlsbad CA USA catalog number BMS2016 2 9 21 24 Statistical analysis Microsoft Excel was used for data entry and R Statistical Software version 3 6 325 was used for all statistical analyses For the categorical variables frequency tables were used to assess distributions Both center and dispersion values were calculated according to the type of variable as was the normality of the distribution for numeric variables Normally distributed numeric variables are presented as means and standard deviations SD If the distribution is abnormal median values and interquartile ranges IQR are given Differences for each variable according to the treatment group HBOT vs control were analyzed using the t test for numerical variables and the chi square or Fisher s exact test for categorical variables Changes in serum MMP 9 levels in the treatment group according to the time of examination were visually evaluated using graphs and linear mixed model analysis with random intercept these analysis measurements were repeated for each study subject The effects of HBOT on serum MMP 9 levels and RPQ scores were evaluated using linear regression models The modeling results are reported as changes in the outcome value for each unit increase in the independent variable lower and upper limits of the 95 confidence interval CI and p values which were considered statistically significant below 0 05 Results Patient characteristics including age sex and the presence of intracranial bleeding are presented in Table 1 The average patient age was 39 years and this was not significantly different between the HBOT and control groups The male tofemale ratio was approximately 6 5 3 5 Intracranial bleeding was found in over half of the cases and these were equally distributed between the HBOT and control groups No patient dropped out during the 6 week study period Serum MMP 9 levels and RPQ scores Table 2 displays the MMP 9 levels and RPQ scores at different measurement time points The two patient groups had different MMP 9 serum levels even before treatment was initiated however this difference was not statistically significant While both groups experienced decreases in MMP 9 levels during the course of this study the reduction in the HBOT group was significantly greater than in the control group 40 6 ng mL vs 21 7 ng mL p 0 001 Therefore even though the control group had higher MMP 9 levels at baseline this was compensated for by the highly significant endpoint difference the difference between both scores delta after five weeks RPQ scores did not differ between the two study groups at baseline However by the end of week five patients receiving HBOT had marked improvements in their RPQ scores compared with controls mean 3 1 vs 6 5 for RPQ 3 14 2 vs 29 6 for RPQ 13 both p values 0 001 As a result the delta scores from baseline to week five in the HBOT group were higher compared with the control group 8 0 vs 4 2 for RPQ 3 23 4 vs 7 2 for RPQ 13 both p values 0 001

Table 1 Patient characteristics Characteristics Total N 20 Control n 10 HBOT n 10 Pa Age SD 38 9 15 1 38 0 15 0 39 7 16 0 0 809 Male 13 65 7 30 6 60 1 000 Female 7 35 3 30 4 40 6 30 3 30 3 30 SAH 5 25 2 20 3 30 EDH 3 15 1 10 2 20 Sex Intracranial bleeding Negative 1 000 Positive SDH 1 5 1 10 0 0 ICH 5 25 3 30 2 20 Note Data were successfully collected from twenty mild traumatic brain injury patients who participated in the study The average age of these patients was 40 years but with quite a large variation SD 15 years Table 1 There was no significant difference in the ages of patients in the HBOT group compared to controls The male to female ratio was approximately 6 5 3 5 and the ratio was relatively unchanged when stratified according to the HBOT vs control group Overall patient characteristics in both groups were relatively similar This is important to consider as the allocation of treatments in the study was not completely random The absence of difference between both groups suggests that selection bias related to age sex or location of intracranial hemorrhage might be excluded Another positive thing that supports the fairly even distribution of patients in the two study groups thereby minimizing the possibility of selection bias that might be feared due to the allocation of treatment without randomization is shown in Table 5 2 RPQ scores in the treatment group did not differ from the control group at baseline Abbreviations SD standard deviation HBOT hyperbaric oxygen therapy SAH subarachnoid hemorrhage EDH epidural hematoma SDH subdural hemorrhage ICH intracerebral hemorrhage a t test for numeric variables Fisher s exact test for categorical variables Figure 1 illustrates the changes in serum MMP 9 levels in both groups at four different time points MMP 9 levels in the HBOT group are consistently lower and decline notably after two weeks compared with the control group Figure 2 shows that the patients receiving HBOT had overall lower MMP 9 levels than those in the control group Similarly Figure 3 depicts the declining MMP 9 concentration in both groups over time and highlights that HBOT causes MMP 9 values to plummet within four weeks of initiating post injury treatment compared with controls Regression analysis using a linear mixed model revealed that changes in MMP 9 level depended on the interaction between both groups HBOT vs control and time Figure 4 The p values for the group time and the interaction of both variables were highly statistically significant p 0 001 There was a decrease in MMP 9 concentration in both groups over time however this decline was accelerated in the HBOT group compared with controls Effects of HBOT on serum MMP 9 levels and RPQ scores Table 3 presents the regression analysis of the relationship between the outcome variables MMP 9 concentration and RPQ scores and HBOT administration Scores for the individual variables were marked as delta and represent the difference between the results at week five and baseline Compared with controls the HBOT group exhibited significant declines in MMP 9 concentration 127 91 vs 9 76 p 0 001 RPQ 3 4 78 vs 2 82 p 0 001 and RPQ 13 scores 19 62 vs 12 78 p 0 001 Discussion Based on their timing and different distinctive underlying pathomechanisms head traumas are categorized as primary or secondary brain injury 26 Secondary brain injury occurs in the weeks following and in response to the primary brain injury which occurs at the time of and is directly caused by the trauma itself Patients with secondary brain injury experience biochemical metabolic and cellular changes that are orchestrated by a complex biochemical cascade that causes increased intracranial pressure BBB damage neuroinflammation brain edema brain hypoxia ischemia and neurodegeneration 27 The outcome of TBI is dependent on the secondary brain injury process 28 The starting point of our study is based on this secondary pathomechanism and its reversible dynamic nature The ECM and BBB both play important roles in neuroplasticity 29 An imperative factor in the damage of the BBB is MMP 9 which is produced by microglia the first line of defense against brain injury As the sensors and effectors of the brain s immune system microglial activity is the primary marker of neuroinflammation 30 31

Table 2 Serum MMP 9 levels and RPQ scores in the HBOT and control groups Variableb Total N 20 Control n 10 HBOT n 10 pa 100 6 17 4 113 1 16 2 88 2 5 9 0 001 MMP 9 ng mL Baseline Week 1 Week 3 Week 5 Delta RPQ Score 91 17 9 78 2 21 1 69 5 24 2 105 13 96 6 11 5 91 3 10 2 77 1 8 8 59 8 7 4 47 6 8 3 0 001 0 001 0 001 31 2 13 5 21 7 8 5 40 6 10 7 0 001 10 9 0 9 10 7 0 9 11 1 0 9 0 340 Baseline RPQ 3 RPQ 13 Week 5 RPQ 3 RPQ 13 Delta RPQ 3 RPQ 13 37 2 3 1 4 8 1 9 21 9 8 1 6 1 2 2 15 3 9 36 8 2 8 6 5 1 37 6 3 4 3 1 0 3 0 574 0 001 29 6 2 4 14 2 1 2 0 001 4 2 1 8 0 1 1 0 001 7 2 3 5 23 4 3 7 0 001 Note Table 2 shows even distribution of RPQ scores from patients in the two study groups since baseline thereby minimizing the possibility of selection bias that might be feared due to the allocation of treatment without randomization Similarly the mean reduction in serum levels in the HBOT group was significantly greater than controls 40 6 ng mL vs 21 7 ng mL p

Figure 2 Changes in MMP 9 levels in the HBOT and control groups Figure 3 MMP 9 concentration over time Enzymes such as occludin and claudin cause the basal degradation of endothelial lamina by targeting laminin fibronectin collagen proteoglycan and tight junction proteins degradation of ZO 1 which are also the two main functional elements of the BBB 32 The downstream outcomes of the enzymatic digestion of these structural proteins include cytoskeletal damage the disruption of cellular homeostasis ischemia inflammation tissue necrosis and apoptosis 33 The stability of the microenvironment surrounding neurons including the ECM is necessary for healthy brain function This stability is sustained by the BBB to maintain brain homeostasis and prevent cell death and dysfunction 32 The integrity of the BBB is the key to the restoration of brain homeostasis after physical injury The BBB plays a pertinent role in this process because its integrity is influenced by MMP 9 activity a major effector in secondary brain injury Therefore MMP 9 represents a reliable relevant biological marker for predicting TBI outcome 34

Figure 4 Linear mixed model output for MMP 9 serum levels treatment group time of examination and their interaction as fixed effects Table 3 Regression model of the relationship between treatment type MMP 9 serum level and RPQ scores Outcome HBOT vs Control 95 CI P value MMP 9 18 83 27 91 9 76 0 001 RPQ 3 3 80 4 78 2 82 0 001 RPQ 13 16 20 19 62 12 78 0 001 Note CI confidence interval In the present study HBOT was administered to prevent or inhibit MMP 9 production HBOT acts on the neutrophil adhesion mechanism in endothelial cells More specifically HBOT reduces the expression of endothelial adhesion molecules and inhibits neutrophil adhesion molecule clustering thus reducing the number of neutrophils adhered to endothelial cells Decreasing the number of neutrophils activates pro inflammatory processes including the release of pro inflammatory cytokines e g IL1B IL 6 IL8 and TNFa and decreases the expression of MMP 9 35 The results of this study demonstrate HBOT s ability to inhibit the production of MMP 9 Similarly a study in Canada found improved cognitive function quality of life and elevated brain activity in patients with mild TBI and prolonged post concussion syndrome after 40 HBOT sessions over two months 36 In another study 40 rats were subjected to dynamic cortical deformation DCD and then divided into three treatment groups three hours after the initial trauma 20 rats received HBOT with 100 oxygen at 2 8 ATA for 45 minutes 10 received 100 oxygen for 45 minutes under normobaric conditions 1 ATA and 10 were untreated controls Neuroinflammatory markers TIMP 1 and TIMP 2 and MMP 9 levels were measured 96 hours after treatment MMP 9

levels were found to be significantly lower in the HBOT group relative to controls Inflammatory infiltration around the focus of necrotic brain tissue was prominent in most untreated animals and was composed predominantly of myeloperoxidase positive neutrophils Both normobaric and hyperbaric hyperoxia resulted in a significant decrease of neutrophil infiltration GLM ANOVA main effect of treatment group P 0 0001 This reduction in the neuroinflammatory response was more extended with HBOT in comparison with normobaric hyperoxia Tukey Kramer P 0 05 12 HBOT decreases intracranial pressure ICP reduces cerebrospinal fluid CSF pressure in patients with acute brain injury restores the metabolic activity of the substance grisea in a single photon emission computerized tomography scans of the closed head injury and to restore glucose metabolism after brain injury 37 38 HBOT decreases mortality rates and improves the functional outcome of patients with severe head injuries In such chronic brain injuries HBOT increases cerebral blood flow improves neuropsychological disorders and promotes neurophysiological and electrophysiological recovery 39 Several studies have reported that multiple HBOT sessions can improve neurological deficits and cognitive impairment in both acute and advanced chronic phases of head injury in rats 40 42 The long term therapeutic effects of HBOT are derived from the induction of angiogenesis neuroplasticity and proliferation and differentiation of nerve stem cells When HBOT was administered within three hours post injury in a fluid percussion mouse model of TBI there was a significant increase in the number of endothelial cells neurons and new glial cells four days after the initial injury 40 Ten daily HBOT sessions at 2 5 ATA for 60 minutes enhances neuroplasticity by increasing axonal sprouting and synapse remodeling contributing to the restoration of locomotor function in rats with TBI 42 Harch et al study on the blood vessel density was measured bilaterally in the hippocampus using diaminobenzidine staining and correlated with MWT performance Morris Water Task They found increased vascular density in the bruised hippocampus and improved cognitive function in these rats after consecutive HBOT sessions 7 days week 1 5 ATA 90 minute sessions HBOT over 40 days 43 Several signaling pathways and transcription factors have been implicated in HBOT induced neurogenesis including wingless related integration site hypoxia inducible factors and cAMP response element binding 44 The outcome of this study was evaluated by the RPQ RPQ 13 scores range from zero to 52 with higher scores reflecting more severe post concussion syndrome The RPQ 13 items encompass a group of advanced symptoms while the RPQ 3 symptoms include headaches dizziness and nausea All those symptoms included in RPQ highly impact patient participation in social activities psychosocial functioning and lifestyle 17 During the three to six months typically required to resolve these symptoms patients are advised to gradually resume their routine activities If symptoms do not resolve within this period patients are usually often referred to a specialist for further assessment and treatment services 45 The data collected in the present study showed a notable decrease in the RPQ 3 and RPQ 13 scores There was no significant difference in the baseline RPQs of the groups at the beginning of the study However by the end of the five weeks of treatment the patients receiving HBOT had much better RPQ scores mean 3 1 vs 6 5 for RPQ 3 and 14 2 vs 29 6 for RPQ 13 both p values 0 001 As a result the delta score from baseline to the end of week five in the HBOT group was significantly lower than the control group 8 0 vs 4 2 for RPQ 3 p 0 001 23 4 vs 7 2 for RPQ 13 p 0 001 The decline in RPQ scores over time was significant RPQ 3 3 80 p 0 001 and RPQ 13 16 20 p 0 001 HBOT administration in patients with mild TBI contributed to this improvement RPQ scores Based on the findings described here HBOT confers great benefits by improving the quality of life of patients with mild TBI and likely prevents further brain damage by increasing cerebral blood flow improving neuropsychological disorders and promoting neurophysiological and electrophysiological recovery Given that repeated HBOT sessions restored neurological deficits and cognitive impairment HBOT should be incorporated as a therapeutic modality for the treatment of patients with head injuries Although there are many benefits of HBOT there are also several drawbacks such as seizures oxygen poisoning pneumothorax and middle ear injuries Although this study supports the efficacy of HBOT in treating post concussion syndrome the small sample population and single biomarker are limitations that must be addressed in future studies The kinetics and underlying mechanisms of HBOT also warrant further investigation to maximize the many beneficial effects of HBOT Conclusions HBOT effectively relieves the symptoms associated with post concussion syndrome through a mechanism that involves repressing MMP 9 activity

Consent statement Written informed consent for publication of the patients details was obtained from the patients a guardian of the patient Data availability Underlying data Zenodo Hyperbaric oxygen therapy ameliorates the symptoms of post concussion syndrome by inhibiting MMP 9 activity A Randomized Controlled Trial in Indonesia http doi org 10 5281 zenodo 4785703 46 This project contains the following underlying data Maximilian Data Post Concussion Syndrome 2 docx Patient characteristics Maximilian Data Post Concussion Syndrome xlsx Human Matrix Metalloproteinase 9 MMP 9 VALUE Reporting guidelines This project also contains the following reporting guidelines CONSORT checklist CONSORT flowchart Data are available under the terms of the Creative Commons Attribution 4 0 International license CC BY 4 0 References 1 Jalali R Rezaei M A Comparison of the Glasgow Coma Scale Score with Full Outline of Unresponsiveness Scale to Predict Patients Traumatic Brain Injury Outcomes in Intensive Care Units Crit Care Res Pract 2014 2014 1 4 PubMed Abstract Publisher Full Text Free Full Text 10 Oley MH Oley MC Aling DMR et al Effects of hyperbaric oxygen therapy on the healing of thermal burns and its relationship with ICAM 1 A case control study Ann Med Surg 2021 61 104 109 PubMed Abstract Publisher Full Text Free Full Text 2 Samma L Widodo D A case evaluation of traumatic brain injury in Wahidin Sudirohusodo Hospital Makassar during January 2016 December 2017 Bali Med J 2019 8 542 Publisher Full Text 11 S nchez EC Mechanisms of Action of Hyperbaric Oxygenation in Stroke Crit Care Nurs Q 2013 36 290 298 PubMed Abstract Publisher Full Text 12 3 Wardoyo MS Widodo D Ihwan A et al The relationship between different dosages of mannitol 20 and osmolarity blood sugar serum and coagulation factors in moderate brain injury patients with increased intracranial pressure Med Cl nica Pr ctica 2021 4 100235 Publisher Full Text Vlodavsky E Palzur E Soustiel JF Hyperbaric oxygen therapy reduces neuroinflammation and expression of matrix metalloproteinase 9 in the rat model of traumatic brain injury Neuropathol Appl Neurobiol 2006 32 40 50 PubMed Abstract Publisher Full Text 13 4 Prasetyo E The primary secondary and tertiary brain injury Crit Care Shock 2020 23 4 13 Rempe RG Hartz AM Bauer B Matrix metalloproteinases in the brain and blood brain barrier Versatile breakers and makers J Cereb Blood Flow Metab 2016 36 1481 1507 PubMed Abstract Publisher Full Text Free Full Text 5 NASEM Evaluation of the Disability Determination Process for Traumatic Brain Injury in Veterans In Veterans Diagnosis Assess Trauma Brain Inj Washington D C National Academies Press 2019 PubMed Abstract Publisher Full Text 14 Wolf G Cifu D Baugh L et al The Effect of Hyperbaric Oxygen on Symptoms after Mild Traumatic Brain Injury J Neurotrauma 2012 29 2606 2612 PubMed Abstract Publisher Full Text 15 6 Sepahvand E Jalali R Mirzaei M et al Glasgow coma scale versus full outline of unresponsiveness scale for prediction of outcomes in patients with traumatic brain injury in intensive care unit Turk Neurosurg 2015 PubMed Abstract Publisher Full Text Cifu DX Hart BB West SL et al The Effect of Hyperbaric Oxygen on Persistent Postconcussion Symptoms J Head Trauma Rehabil 2014 29 11 20 PubMed Abstract Publisher Full Text 16 D Souza M Trivedi R Singh K et al Traumatic brain injury and the post concussion syndrome A diffusion tensor tractography study Indian J Radiol Imaging 2015 25 404 PubMed Abstract Publisher Full Text Free Full Text 17 Potter S Leigh E Wade D et al The Rivermead Post Concussion Symptoms Questionnaire J Neurol 2006 253 1603 1614 Publisher Full Text 18 Asselstine J Kristman VL Armstrong JJ et al The Rivermead PostConcussion Questionnaire score is associated with disability and self reported recovery six months after mild traumatic brain injury in older adults Brain Inj 2020 34 195 202 PubMed Abstract Publisher Full Text 19 Agha RA Borrelli MR Vella Baldacchino M et al The STROCSS statement Strengthening the Reporting of Cohort Studies in 7 Shinomiya N Molecular Mechanisms of Hyperbaric Oxygen Therapy In Hyperb Oxyg Ther Singapore Springer Singapore 2020 3 20 Publisher Full Text 8 Raveenthiraraja T Manoharan S Hyperbaric oxygen therapy A review Int J Pharm Pharm Sci 2013 5 52 54 9 Oley MH Oley MC Tjandra DE et al Hyperbaric oxygen therapy in the healing process of foot ulcers in diabetic type 2 patients marked by interleukin 6 vascular endothelial growth factor and PEDIS score A randomized controlled trial study Int J Surg Open 2020 27 154 161 Publisher Full Text

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MEDICAL GAS RESEARCH Hyperbaric oxygen therapy 1 5 ATA in treating sports related TBI CTE two case reports Stoller Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 5 July 2011

Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 CASE REPORT MEDICAL GAS RESEARCH Open Access Hyperbaric oxygen therapy 1 5 ATA in treating sports related TBI CTE two case reports Kenneth P Stoller Abstract Despite adequate evidence including randomized controlled trials hyperbaric oxygen is not yet recognized as efficacious for treating various forms of brain injury specifically traumatic brain injury Political economic issues have kept this benign therapy from being widely adopted despite the lack of viable alternatives Two football players with TBI CTE are herewith shown to benefit from being treated with hyperbaric oxygen as documented by neurocognitive examinations and functional brain imaging in one case treatment commenced decades after the brain injury Perhaps the interest in HBOT by those participating in high risk sports will help expand this orphan therapy into mainstream medicine Background Many more concussions were being reported in the National Football League NFL in the 2010 season A total of 154 concussions including practices and games were reported from the start of the preseason through the eighth week of the 2010 regular season That is an increase of 21 percent over the 127 concussions during the same span in 2009 and a 34 percent jump from the 115 reported through the eighth week of the 2008 season Associated Press Dec 13 2010 This either means better reporting is taking place or the game is getting more violent or some combination of the above What hasn t changed is a lack of treatment The lack of appropriate treatment for traumatic brain injury TBI and chronic traumatic encephalopathy CTE is not exclusive to the NFL by any means but football players both young high school students and old retired NFL veterans are beginning to ask for Hyperbaric Oxygen Therapy HBOT to help them recover from their injuries Is this a new fad for a therapy looking for a disease after all if HBOT were efficacious in treating TBI CTE wouldn t everyone in the medical profession know about it and wouldn t there be chambers connected to every trauma center HBOT has been treating brain injuries as far back as 1963 when it was first found effective in treating carbon Correspondence info hbotnm com 404 Brunn School Rd D Santa Fe NM 87505 USA monoxide poisoning 1 2 Although the misconception that HBOT is only treating carboxy hemoglobin persists to this day 3 Brain injuries caused by decompression sickness and arterial gas emboli began being treated by HBOT using Navy Treatment Table six 4 5 Delayed treatment 3 months of an ischemic stroke with HBOT was reported by the US Navy in 1969 6 Subsequently successful treatment with HBOT for late treatment of a stroke diabetic encephalopathy and near drowning global anoxia was also reported 7 with the additional evidence of pre and post functional brain imaging SPECT The medical literature continues to grow showing HBOT is efficacious in treated old carbon monoxide poisoning COP also known as delayed neuropsychiatric syndrome DNS of COP 8 9 While the historic literature points to HBOT as being efficacious for many conditions HBOT is an orphan therapy that falls outside of the medical paradigm where drugs and interventions are selectively fed into the standard ofcare reimbursement complex by corporate interests that control both medical information and clinical practice trends A therapy that exists outside our dysfunctional medical paradigm tends to be but a footnote that is passed over buried or ignored completely Despite controlled randomized trials demonstrating HBOT s efficacy for treating TBI ignoring HBOT for treating brain injuries seems to have became codified even though other forms of intervention have fallen short High quality clinical trials demonstrating the efficacy of HBOT in brain injury eventually get buried or 2011 Stoller licensee BioMed Central Ltd This is an Open Access article distributed under the terms of the Creative Commons Attribution License http creativecommons org licenses by 2 0 which permits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 forgotten Sometimes those who don t understand the nuance of oxygen dose or timing perform studies and resulting misinterpreted results further marginalize this important tool for treating brain injury 10 Recent clinical research has demonstrated the efficacy of HBOT at 1 5 ata for traumatic brain injury even when administered years after injury While in acute severe TBI HBOT has been shown to be effective in reducing mortality 11 Harch et al demonstrated consistent SPECT brain imaging improvements showing improved brain blood flow in chronic TBI patients treated with HBOT 1 5 12 15 Since the original work of Drs Neubauer and Harch the efficacy of HBOT 1 5 in a chronic stable TBI has been well documented 16 17 Patients with abnormal functional brain scans secondary to TBI show consistent improvement after HBOT 1 5 Recently Harch et al reported dramatic improvement in a series of 15 patients treated with HBOT 1 5 in a clinical trial of military acquired TBI 18 Functional brain scans continue to document results for HBOT 1 5 for combat related blast related TBI 19 In two airmen with pre injury neuropsychiatric testing and chronic stable TBI symptoms HBOT 1 5 resulted in resolution of symptoms as well as a return to the pre injury values for testing 20 In a randomized controlled trial of stable severe TBI treated with HBOT 1 5 Lin et al demonstrated improvement in the Glasgow Coma Scale Page 2 of 5 21 Rockswold has demonstrated improvement in the Glasgow Coma Scale and reduced mortality in acute TBI patients undergoing HBOT with minimal risk 22 23 HBOT 1 5 in this group of acute patients appears safe and does not produce oxygen toxicity 24 Other individual trials also have demonstrated the efficacy of HBOT 1 5 for chronic stable TBI 25 A 310 patient Chinese trial demonstrated improvement clinically in neuropsychiatric testing as well as in functional brain imaging after HBOT 1 5 26 In a randomized controlled trial of 21 brain injured adults HBOT 1 5 resulted in improved neuropsychiatric testing for the treated group 27 Presented here are two case reports of football induced TBI CTE that responded to HBOT Case presentations Case 1 A retired NFL player now in his early 50 s was first hospitalized with a TBI resulting in loss of consciousness during a tackling drill playing pop Warner football Numerous minor concussions took place over the years but he experienced his second major concussion during the first play for his NFL team San Francisco 49ers early in the first quarter and went through 25 to 30 smelling salts in order to finish the game He had no recollection of participating in that game but was sent back on the practice field the very next day Figure 1 Source MicroCog Assessment Independent Evaluation by Amen Clinic The lower the score on reaction time the better the result

Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 Page 3 of 5 A B Figure 2 Pre and post HBOT SPECT imaging showing overall improved blood flow in the post HBOT scan per Daniel Amen MD Early in his second season 1981 he developed hydrocephalus and underwent emergency shunt brain surgery Four months after his team won Super Bowl XVI his shunt failed and he had back to back emergency brain surgeries and was given last rites By 1990 he had nine more shunt revisions He was treated with HBOT 1 5 times 40 for 60 minutes per treatment one treatment per day 100 oxygen He was independently followed with neurocognitive evaluations and SPECT brain imaging by the Amen Clinic in California Figure 1 shows a marked improvement in 5 out of 6 indices on the MicroCog assessment done after

Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 40 HBOT sessions Figure 2 shows the SPECT scan of this former football player Page 4 of 5 100 90 Case 2 A high school football player age 15 years received two concussions two weeks apart After the first concussion 1st scrimmage game of the year he became nauseous very emotional broke down after the game and wanted to quit the team A week later he received two head to head contacts without loss of consciousness he was not able to attend school the week following this concussion because of headaches A CAT scan was negative and his neurologist diagnosed him with migraines He lost his ability to read of over the next two weeks and was diagnosed with post concussion syndrome PCS and treated with steroids without clinical response and then narcotics along with a black boxed anti depressant Three months later he took the IMPACT neurocognitive test a test originally developed to evaluate sports concussions at the University of Pittsburgh Medical Center UPMC for their Sports Medicine Concussion Program 28 29 The IMPACT software evaluates and documents multiple aspects of neurocognitive functioning including memory brain processing speed reaction time and post concussive symptoms Furthermore unlike standard neurocognitive testing modalities the immediate post concussion assessment and cognitive testing has shown itself to be a reliable evaluation tool with virtually no practical effect on score stability 30 He was treated with the identical protocol his retired NFL counterpart was treated with i e HBOT 1 5 60 minutes once a day for 40 sessions of 100 oxygen Figure 3 shows the results of the IMPACT neurocgnitve test on this young football player Without a baseline test prior to the season it is impossible to know what this player s normal verbal and visual memory scores might have been All that can be said is that after 40 treatment sessions he is almost matching High school mean scores His visual motor speed improved 35 and his reaction time improved 25 which place him better than and at high school mean respectively Subjectively he experienced an 80 reduction in frequency and severity of headaches and a complete resolution of nausea Conclusions HBOT is the only non hormonal treatment approved by the FDA for the repair and regeneration of human tissue Six of the 13 approved indications are directly related to brain injury and wound repair relevant to treating TBI The USA Olympic team has now brought 80 70 60 50 Verbal Memory Visual Memory 40 30 20 10 0 Pre HBOT 20 HBOT 40 HBOT High School Mean Figure 3 IMPACT test results for verbal and visual memory pre HBOT at 20 treatments and at 40 treatments in HBOT to treat sports related injuries as part of their armamentarium The treatment of traumatic brain injury with hyperbaric oxygen is evolving rapidly A plethora of cellular studies demonstrate the mechanisms of favorable action of HBOT and animal studies have also been able to add to the clinical rationale and utility for treating a variety of traumatic and ischemic brain injuries Controlled randomized clinical trials have demonstrated efficacy of HBOT for traumatic brain injury Therefore the time has come for this orphan therapy to be adopted and for it to take its place as standard practice for treating both acute and chronic TBI High profile athletes who are willing to share their response to HBOT for TBI CTE may accelerate the day when this benign humanitarian and non invasive therapy is recognized as a quintessential tool for treating brain injury after all oxygen is what the brain fundamentally feeds upon Consent Written informed consent was obtained from the patients or their legal guardians for publication of this Case report and any accompanying images A copy of the written consent is available for review by the Editorin Chief of this journal Abbreviations ATA atmospheres absolute COP carbon monoxide poisoning CTE chronic traumatic encephalopathy DNS delayed neuropsychiatric syndrome HBOT hyperbaric oxygen therapy IMPACT immediate post concussion assessment and cognitive testing NFL National Football League PCS post concussion syndrome SPECT single photon emission computed tomography TBI traumatic brain injury Acknowledgements The two patients in this case report were treated at the Hyperbaric Oxygen Clinic in Sacramento with the assistance and dedication of Mike Greenhalgh and Bryan Burke

Stoller Medical Gas Research 2011 1 17 http www medicalgasresearch com content 1 1 17 Authors contributions KPS supervised the hyperbaric oxygen treatments and in case two supervised the administration of the IMPACT test KPS also drafted the manuscript Authors information KPS is an Adjunct Assistant Professor AT Still University SOMA Competing interests Dr Stoller is President of the International Hyperbaric Medical Association and a principal investigator on the International Hyperbaric Medical Foundation s National Brain Injury Rescue Rehabilitation Project He is medical director of the New Hope Clinic Santa Monica San Francisco Institute for Hyperbaric Medicine the Hyperbaric Oxygen Clinic of Sacramento and the Hyperbaric Medical Center of New Mexico Received 25 March 2011 Accepted 5 July 2011 Published 5 July 2011 References 1 Sluiter ME The treatment of carbon monoxide poisoning by the administration of oxygen at high atmospheric pressure Proc R Soc Med 1963 56 1002 8 2 Smith G Sharp GR Lancet 1960 2 905 3 Stoller KP Hyperbaric oxygen and carbon monoxide poisoning a critical review Neurological Research 2007 29 146 155 4 Duff JH Shibata HR Vanschaik L Usher R Wigmore RA MacLean LD Hyperbaric oxygen a review of treatment in eighty three patients Can Med Assoc J 1967 97 10 510 5 5 Workman RD Oxygen decompression following air dives for use in hyperbaric oxygen therapy Res Rep 2 64 Rep US Navy Exp Diving Unit 1964 Dec 15 1 10 6 Hart GB Thompson RE The treatment of cerebral ischemia with hyperbaric oxygen OHP Stroke 1971 2 3 247 50 7 Neubauer RA James P Cerebral oxygenation and the recoverable brain Neurol Res 1998 20 Suppl 1 S33 6 8 Chang DC Lee JT Lo CP Fan YM Huang KL Kang BH Hsieh HL Chen SY Hyperbaric oxygen ameliorates delayed neuropsychiatric syndrome of carbon monoxide poisoning Undersea Hyperb Med 2010 37 1 23 33 9 Senol MG Yildiz S Ersanli D Uzun G Gumus T Narin Y Ozkan S Ayata A Carbon monoxide induced cortical visual loss treatment with hyperbaric oxygen four years later Med Princ Pract 2009 18 1 67 9 10 Stoller KP Hyperbaric oxygen and carbon monoxide poisoning a critical review Neurological Research 2007 29 146 155 11 Clifton GL Hypothermia and hyperbaric oxygen as treatment modalities for severe head injury New Horiz 1995 3 3 474 8 Review 12 Harch PG Van Meter KW Gottlieb SF Staab P HMPAO SPECT brain imaging and low pressure HBOT in the diagnosis and treatment of chronic traumatic ischemic hypoxic and anoxic encephalopathies Undersea and Hyperbaric Medicine 1994 21 Suppl 30 13 Harch PG Van Meter KW Neubauer RA Gottlieb SF Use of HMPAO SPECT for assessment of response to HBO in ischemic hypoxic encephalopathies In Textbook of Hyperbaric Medicine Appendix 2 edition Edited by Jain KK Seattle WA Hogrefe 1996 480 491 14 Harch PG Neubauer RA Hyperbaric oxygen therapy in global cerebral ischemia anoxia and coma In Textbook of Hyperbaric Medicine Volume Chapter 18 3 edition Edited by Jain KK Hogrefe 1999 319 345 15 Harch PG Neubauer RA Hyperbaric oxygen therapy in global cerebral ischemia anoxia and coma In Textbook of Hyperbaric Medicine Volume Chapter 18 3 edition Edited by Jain KK Hogrefe 1999 319 345 16 Neubauer RA Gottlieb SF Pevsner NH Hyperbaric Oxygen for Treatment of Closed Head Injury Southern Medical Journal 1994 87 9 933 936 17 Barrett KF Masel BE Harch PG Ingram F Corson KP Mader JT Cerebral blood flow changes and cognitive improvement in chronic stable traumatic brain injuries treated with hyperbaric oxygen therapy Neurol 1998 Suppl A178 A179 18 Harch PG Andrews SR Fogarty E Lucarini J Aubrey C Staab PK Van Meter KW Hyperbaric Oxygen Therapy Treatment of Chronic MildModerate Blast Induced Traumatic Brain Injury Post Concussion Syndrome with Post Traumatic Stress Disorder Pilot Trial Presented at 8th World Congress on Brain Injury Washington DC 2010 Page 5 of 5 19 Harch PG Fogarty EF Staab PK Van Meter K Low pressure hyperbaric oxygen therapy and SPECT brain imaging in the treatment of blastinduced chronic traumatic brain injury post concussion syndrome and post traumatic stress disorder a case report Cases J 2009 2 6538 20 Wright JK Zant E Groom K Schlegel RE Gilliland K Case report Treatment of mild traumatic brain injury with hyperbaric oxygen Undersea Hyperb Med 2009 36 6 391 9 21 Lin JW Tsai JT Lee LM Lin CM Hung CC Hung KS Chen WY Wei L Ko CP Su YK Chiu WT Effect of hyperbaric oxygen on patients with traumatic brain injury Acta Neurochir Suppl 2008 101 145 9 22 Rockswold SB Rockswold GL Zaun DA Zhang X Cerra CE Bergman TA Liu J A prospective randomized clinical trial to compare the effect of hyperbaric to normobaric hyperoxia on cerebral metabolism intracranial pressure and oxygen toxicity in severe traumatic brain injury J Neurosurg 2010 112 5 1080 94 23 Rockswold SB Rockswold GL Defillo A Hyperbaric oxygen in traumatic brain injury Neurol Res 2007 29 2 162 72 Review 24 Rockswold SB Rockswold GL Defillo A Hyperbaric oxygen in traumatic brain injury Neurol Res 2007 29 2 162 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ImPACT following sport concussion Clin Neuropsychol 2003 17 460 467 doi 10 1186 2045 9912 1 17 Cite this article as Stoller Hyperbaric oxygen therapy 1 5 ATA in treating sports related TBI CTE two case reports Medical Gas Research 2011 1 17 Submit your next manuscript to BioMed Central and take full advantage of Convenient online submission Thorough peer review No space constraints or color figure charges Immediate publication on acceptance Inclusion in PubMed CAS Scopus and Google Scholar Research which is freely available for redistribution Submit your manuscript at www biomedcentral com submit

www nature com scientificreports Persistent post concussion syndrome PPCS is a common and significant morbidity among the pediatric population following traumatic brain injury TBI 1 Mild TBI mTBI accounts for over 90 of all TBI in children affecting more than 10 of children under the age of 162 3 Although most of the children recover completely in a short period of time about 10 30 suffer from PPCS lasting from months to years4 PPCS has a significant effect on a child s quality of life school performance family function and activities5 6 A recent survey confirmed that PPCS is underdiagnosed as over 25 of the children coming to the ED due to mTBI will suffer from PPCS7 Guidelines for early management of acute concussion recommend limited periods of rest followed by a gradual return to school and other activities8 9 Early sub symptom threshold aerobic exercise has shown efficacy and speeds recovery in the first month post injury10 11 In contrast the existing evidence for effective PPCS treatments remains limited12 Treatment usually include physical therapy neurocognitive therapies among others Graded exercise programs cervicovestibular rehabilitation and multimodal collaborative care have been suggested in small studies13 16 The beneficial effects of hyperbaric oxygen therapy HBOT has been recently shown in adults with chronic TBI and PPCS17 21 The combined action of hyperoxia and hyperbaric pressure leads to significant improvements Israel Israel 2022 12 15233

www nature com scientificreports Figure 1 Patient flowchart in tissue oxygenation while targeting both oxygen and pressure sensitive genes17 Preclinical models and clinical studies in TBI survivors have demonstrated the effect of HBOT through several mechanisms including antiinflammatory mitochondrial function restoration increased perfusion via angiogenesis and induction of proliferation and migration of stem cells17 18 22 23 Importantly previous randomized controlled studies conducted on adults with PPCS showed that HBOT can be effective even years after injury20 24 However the effects of HBOT on PPCS in the pediatric population has never been evaluated The aim of the study was to evaluate the effects of HBOT on a pediatric population suffering from PPCS after TBI with ongoing symptoms for at least six months after injury in a randomized sham control double blind clinical trial Out of 52 children that were assessed for eligibility 15 declined to participate and 12 did not meet the screening inclusion criteria Study recruitment was stopped prior to reaching the required sample size since parents of eligible children refused to participate in a sham controlled study A total of 25 included children completed baseline assessments and randomized to either HBOT or sham arms Eight children four of the HBOT and four of the sham group were under ten and did not complete the pretreatment Neurotrax cognitive evaluation All other included children were over ten and completed all cognitive assessments Fig 1 The baseline characteristics of the cohort are provided in Table 1 The mean age of patients at inclusion and at injury were 11 6 2 32 and 6 7 3 18 respectively and 80 were males There were no significant differences in demographics TBI characteristics time from injury or PPCS symptoms between the two arms Patient blinding was found to be reliable where 11 15 73 3 of the HBOT group and 7 10 70 of the sham group perceived they were treated by HBOT p 1 Cognitive function Results of the cognitive function evaluations are summarized in Table 2 The two groups had similar baseline scores in all cognitive domains evaluated by the Neurotrax computerized battery The general cognitive score was significantly improved in the HBOT 90 2 10 3 to 96 1 10 3 p 0 01 compared to the sham group 95 1 7 5 to 94 9 7 7 p 0 96 with a medium effect size of 0 598 see Fig 2 The most striking change was found in the memory domain with a medium effect size d 0 480 and a mean change of 11 5 12 5 p 0 017 following HBOT compared to 3 4 8 8 p 0 39 see Fig 2 There were no statistically significant changes in other cognitive domains Compared to the sham group there were significant changes in the WISC IV digit span test HBOT 2 5 2 3 vs sham 0 7 1 3 p 0 17 d 0 891 and cancellation test HBOT 9 5 14 2 p 0 02 vs sham 7 2 12 5 p 0 12 following HBOT There were significant differences in the baseline scores of the WISC IV symbol search between the groups In the RAVLT test there was a significant increase in the RAVLT total learning following HBOT HBOT 9 1 11 9 p 0 01 vs sham 5 3 8 5 p 0 09 with no change in the learning rate Phonemic fluency was significantly improved in the HBOT group 5 0 8 1 p 0 031 compared to the sham group 2 0 9 7 p 0 55 with a large effect size of 0 806 2022 12 15233 Vol 1234567890

www nature com scientificreports N HBOT group Control group 15 10 P value Age 11 99 2 32 11 00 2 32 0 308 Adolescents 7 46 7 3 30 0 0 678 Gender 12 80 0 8 80 0 1 000 Number of siblings 3 40 2 75 3 00 0 94 0 663 Age at injury 7 10 3 52 6 15 2 70 0 477 Time after injury 4 89 2 23 4 85 1 78 0 965 Mechanism of injury Motor vehicle collision 2 13 3 4 40 0 0 175 Bicycle accident 2 13 3 1 10 0 1 000 Fall 8 53 3 4 40 0 0 688 Struck by object blow 3 20 0 1 10 0 0 626 Arrival by EMS 12 80 0 7 70 0 0 653 Skull fractures 5 33 3 1 10 0 0 345 Abnormal imaging finding 5 33 3 4 40 0 1 000 Hospitalization 7 46 7 5 50 0 1 000 Symptoms LOC 5 33 3 2 20 0 0 659 Amnesia 3 20 0 3 30 0 0 653 Vomiting 6 40 0 3 30 0 0 691 Severity score Mild 11 73 3 7 70 0 1 000 Moderate 2 13 3 3 30 0 0 358 Severe 2 13 3 0 0 0 0 500 Presence of persistent post concussion symptoms Learning difficulties 15 100 0 8 80 0 0 150 Poor concentration 14 93 3 8 80 0 0 543 Social difficulties 5 33 3 6 60 0 0 241 Forgetfulness memory 8 53 3 7 70 0 0 678 Chronic headache 7 46 7 3 30 0 0 678 Depression anxiety 7 46 7 6 60 0 0 688 Current ADHD medications 4 26 7 3 30 0 1 000 Table 1 Baseline characteristics Data presented as n continuous data mean SD In the 5PT test both groups improved significantly at the one minute score HBOT 3 7 3 4 p 0 001 sham 1 6 1 1 p 0 003 However after two minutes only the HBOT group had a significant change HBOT 4 4 4 8 p 0 004 sham 1 1 3 8 p 0 43 with a medium effect size of 0 739 No significant differences were found in the TOMAL2 test for both groups There were significant differences in the baseline scores in semantic fluency NEPSY animal sorting and the trail making test A Results of the cognitive function evaluations are summarized in Table 2 PPCS symptoms Compared to the sham group the HBOT group had significant improvements in the HBI total score 7 9 10 7 p 0 02 d 0 171 cognitive score 12 6 19 1 p 0 035 d 0 269 and somatic score 12 3 18 5 p 0 034 d 0 02 The HBI emotional and fatigue scores did not change p 0 05 In the BC PSI questionnaire both groups had significant increases in their emotional score with a large negative effect size HBOT 6 9 11 1 p 0 04 sham 15 7 16 2 p 0 04 d 0 676 The HBOT group had a significant increase in BC PSI cognitive score 10 3 11 4 p 0 007 with a small negative effect size d 0 17 The quality of life questionnaire PedsQL showed significant improvements in both the psychosocial health summary score 9 2 13 2 p 0 027 d 0 167 and the school functioning 12 7 17 5 p 0 023 d 0 217 following HBOT Results of PPCS symptoms questionnaires are summarized in Table 3 Behavioral assessment The Conners 3 parents questionnaire showed significant improvements with small to medium effect sizes post HBOT in the following domains of the Conners 3 tool hyperactivity score 6 6 9 8 p 0 031 d 0 244 executive score 11 2 11 6 p 0 004 d 0 153 and learning problem score 5 7 9 2 p 0 044 d 0 59 In the BRIEF questionnaire following HBOT there were significant changes in the global executive composite score 7 0 6 2 p 0 001 d 0 528 planning organizing score 7 7 8 5 p 0 007 d 1 09 initiation 2022 12 15233 Vol 0123456789

www nature com scientificreports HBOT Pre Control Post Change Three months P value Pre Post Change Three months P value P value baseline Net effect size Computerized tests N 10 General 90 2 10 3 Memory 90 75 19 9 Executive function 93 2 11 0 Attention Information processing speed Motor skills 6 0 010 95 2 7 6 95 0 7 7 0 20 9 6 0 960 0 320 0 60 0 017 93 0 11 8 96 4 14 8 3 4 8 9 0 390 0 800 0 48 92 3 12 2 0 89 12 5 0 830 92 8 16 7 89 9 11 2 2 9 20 8 0 740 0 960 0 13 88 5 10 7 95 8 15 0 7 3 17 9 0 230 93 23 15 9 97 8 8 9 4 6 15 4 0 500 0 490 0 20 90 2 9 9 90 6 11 0 0 38 7 5 0 880 84 6 15 1 86 9 13 9 2 1 13 7 0 770 0 440 0 11 88 7 21 1 96 2 17 7 7 6 12 2 0 080 103 0 9 9 100 8 12 1 2 2 7 0 0 470 0 140 0 59 96 1 10 3 5 9 5 8 102 3 9 8 11 5 12 5 Pen and paper tests N 15 WISC 4 digit span 13 3 2 9 15 8 3 2 2 5 2 3 0 001 13 0 1 9 13 7 2 2 0 7 1 3 0 169 0 769 0 89 WISC 4 coding 43 7 11 9 47 2 11 7 3 5 7 4 0 086 34 6 10 3 35 9 12 5 1 3 9 1 0 672 0 081 0 27 WISC 4 symbol search 24 1 6 3 25 7 5 0 0 155 17 1 4 8 21 9 6 7 4 8 4 2 0 010 0 012 0 76 WISC 4 cancellation 84 1 17 0 93 7 18 2 9 5 14 2 0 021 74 2 18 9 81 4 15 7 7 2 12 5 0 121 0 217 0 17 7 3 2 7 1 9 2 4 0 064 0 025 1 13 48 1 11 7 5 3 8 5 0 097 0 946 0 35 6 8 1 8 0 7 2 7 0 486 0 323 0 33 NEPSY animal 7 6 2 1 sorting RAVLT total learning 42 5 9 2 RAVLT learning 7 3 2 5 rate 10 1 6 4 1 7 1 1 4 0 5 1 9 0 346 5 1 2 7 51 5 8 2 9 1 11 9 0 010 42 8 11 8 7 0 1 9 0 3 2 9 0 728 6 1 2 8 Phonemic fluency 17 8 7 3 22 8 8 3 5 0 8 1 0 031 17 9 11 4 15 9 7 6 2 0 9 7 0 553 0 983 0 81 Semantic fluency 34 4 8 2 39 3 7 4 4 9 6 9 0 016 27 3 5 9 35 0 5 5 7 7 9 2 0 038 0 041 0 36 TOMAL2 abstract visual memory 17 9 8 9 22 9 9 4 5 0 11 5 0 115 15 9 7 7 16 3 11 7 0 4 9 2 0 888 0 601 0 43 TOMAL2 memory for location 11 1 5 3 13 7 6 6 2 5 5 9 0 116 7 2 3 9 9 6 4 8 2 3 4 2 0 138 0 078 0 04 TOMAL2 manual imitation 33 0 10 3 33 3 8 6 0 3 12 4 0 935 26 6 12 0 25 6 9 7 1 0 13 0 0 823 0 195 0 10 Trail making test A TMT 35 9 8 9 35 5 9 6 0 3 8 9 0 887 55 9 27 6 49 9 23 8 6 1 34 0 0 608 0 021 0 26 Trail making test B TMT 134 2 77 4 95 7 39 9 38 5 58 3 0 023 174 2 95 3 140 7 76 2 33 6 71 1 0 194 0 294 0 08 Five points 1 min 12 3 5 5 16 1 4 8 3 7 3 4 0 001 9 8 4 2 11 4 4 5 1 6 1 1 0 003 0 283 0 75 Five points 2 min 19 8 8 5 23 2 6 5 4 4 4 8 0 004 15 4 7 6 16 5 6 7 1 1 3 8 0 434 0 254 0 74 Table 2 Cognitive function changes Baseline comparison of p values test the null hypothesis of equal means of the two groups at the baseline using an unpaired t test three months comparison of p values test the null hypothesis of equal means of each group pre post intervention HBOT sham respectively using a paired t test bold P 0 05 Net effect size is the subtraction of Cohen s D effect size of the control group from the HBOT group Cohen s D effect size Neurotrax scores are normalized to age 7 0 7 1 p 0 004 d 0 781 inhibition score 8 4 8 9 p 0 005 d 0 586 working memory 6 1 9 3 p 0 037 d 0 258 and the metacognition index 6 4 5 8 p 0 002 d 0 706 with medium to large effect sizes There was a significant improvement in the shift parameter in the sham group with a small negative effect size d 0 428 p 0 02 Both groups had significant improvement in the behavioral regulation BR index d 0 202 p 0 01 Results of the behavioral assessment data are summarized in Table 4 and Figs 3 and 4 Neuro physical evaluation The two groups had similar non pathological baseline BESS scores total score 12 p 0 05 Following HBOT there was a positive trend in the firm surface total score 4 7 3 0 to 3 4 2 9 2022 12 15233 Vol 1234567890

www nature com scientificreports Figure 2 Primary endpoint changes boxplot Both memory and the general cognitive scores were significantly increased after HBOT with no changes in the sham group Data shown as absolute mean change per cognitive domain The central mark indicates the median and the bottom and top edges of the box indicate the 25th and 75th percentiles respectively Red symbols indicate outliers P 0 05 p 0 054 with a large effect size of 0 99 compared to the sham group 4 2 3 0 to 5 1 2 9 p 0 20 Results of the balance evaluations are summarized in Table 5 Brain imaging Nine patients were excluded due to having the pre and post scans in different MRI machines and head coils due to replacement of the hospital s MRI machine A total of eight subjects of the HBOT group and eight of the sham group were included in the analysis Voxel based analysis demonstrated significant mean diffusivity MD decreases in the HBOT group compared to the sham group in the following regions right fusiform BA37 right lingual gyri BA18 right and left insula BA13 left supramarginal gyrus BA40 and the right and left inferior frontal gyri BA47 Table 6 and Fig 5 There were strong correlations between MD changes and cognitive scores changes phonemic fluency change correlated with MD changes in both the right lingual gyrus r 0 817 p 0 02 right and left insula r 0 636 0 686 p 0 0001 and left supramarginal gyrus r 0 611 p 0 02 The right insula MD changes correlated with both the WISV IV digit span r 0 591 p 0 026 and the 5PT 2 min changes r 0 540 p 0 046 Safety Nine out of the 10 90 sham patients and 13 15 86 7 patients from the HBOT group had side effects p 1 Five out of the nine 55 6 sham patients and 2 13 15 3 of the HBOT group suffered from otalgia Additionally 4 9 44 4 sham patients and 9 13 69 2 HBOT patients had signs of mild moderate barotrauma p 1 All cases were treated conservatively However due to repeated events two patients one HBOT one sham required tympanic tube insertion to continue protocol sessions Lastly two HBOT treated patients complained of headaches which were treated conservatively This is the first randomized sham controlled trial evaluating HBOT s effect on pediatric PPCS patients in a prospective manner We found that HBOT induces significant improvements in cognitive function PPCS behavioral symptoms and quality of life The main improved cognitive domains include memory in both Neurotrax WISC IV RAVLT and verbal fluency and executive function in 5PT and trail making Behavioral symptom changes included executive functioning and attention difficulties These findings were associated with brain microstructural changes in the frontal temporal and the paralimbic regions associated with cognitive roles Following TBIs children and adolescents experience changes in both cognitive and behavioral functioning The most common cognitive function deficits are working memory executive functioning and attention These can significantly affect a child s learning capabilities and quality of life25 28 Moreover the attention and executive functioning deficits impact progress in academics and psychosocial development and may have a negative 2022 12 15233 Vol 0123456789

www nature com scientificreports HBOT Pre N Control Post Change Three months P value 13 Pre Post Change Three months P value P value baseline Net effect size 7 HBI Total HBI score 60 7 16 2 68 6 15 9 7 9 10 7 0 020 58 2 20 9 68 1 17 2 9 9 12 8 0 086 0 784 0 17 Cognitive score 40 6 22 2 53 2 20 4 12 6 19 1 0 035 35 3 23 7 42 5 16 8 7 1 22 5 0 433 0 644 0 27 Somatic score 62 7 30 7 74 9 24 1 12 3 18 5 0 035 71 4 21 0 84 1 15 3 12 7 23 1 0 196 0 531 0 02 Emotional score 70 9 19 2 73 5 17 3 2 6 21 6 0 676 67 7 27 1 74 1 25 0 6 3 14 9 0 303 0 774 0 19 Behavior score 57 8 25 8 64 7 25 0 6 8 17 8 0 190 51 9 28 6 64 0 25 9 12 2 15 1 0 077 0 657 0 32 BC PSI physical score 72 0 26 6 81 9 18 7 9 9 17 9 0 069 68 4 24 1 79 6 12 9 11 2 20 6 0 199 0 780 0 07 BC PSI cognitive score 64 1 18 3 74 4 15 5 10 3 11 4 0 007 50 0 27 5 63 1 12 5 13 1 24 0 0 199 0 210 0 17 BC PSI emotional 71 5 17 9 score 78 5 16 1 6 9 11 1 0 044 62 9 24 9 78 6 24 2 15 7 16 2 0 042 0 406 0 68 BC PSI fatigue Score 60 6 29 8 69 2 24 3 8 7 24 1 0 221 66 1 22 9 82 1 19 9 16 1 22 5 0 108 0 691 0 31 Psychosocial health summary score 67 1 13 3 76 3 16 3 9 2 13 2 0 027 56 4 26 1 63 6 21 5 7 1 10 8 0 132 0 268 0 17 Physical functioning 77 4 21 4 80 3 20 8 2 9 9 4 0 291 70 5 20 2 79 0 15 2 8 5 10 5 0 076 0 517 0 57 Emotional functioning 64 6 18 0 72 7 23 3 8 1 23 5 0 239 60 7 30 4 67 1 31 6 6 4 12 5 0 222 0 737 0 08 BC PSI PedsQL Social functioning 84 6 13 7 91 5 10 3 6 9 14 1 0 102 69 3 36 1 75 7 34 3 6 4 14 6 0 289 0 213 0 04 School functioning 64 6 27 3 12 7 17 5 0 023 39 3 21 8 47 9 15 3 8 6 21 7 0 337 0 293 0 22 51 9 24 5 Table 3 PPCS symptoms changes Baseline comparison of p values test the null hypothesis of equal means of the two groups at the baseline using an unpaired t test three months comparison of p values test the null hypothesis of equal means of each group pre post intervention HBOT sham respectively using a paired t test bold P 0 05 net effect size is the subtraction of Cohen s D effect size of the control group from the HBOT group Cohen s D effect size influence on family functioning29 30 Working memory is a temporary capacity limited memory holding store This memory is critical for executive function that matures during puberty adolescence and early adulthood27 We found a significant increase in working memory in both computerized battery and traditional pen and paper based tests Executive functioning is a term used to describe a variety of abilities allowing purposeful goal directed problem solving behavior including behavioral regulation planning and organizational skills and self monitoring In our study we found that the HBOT group significantly improved in both the objective 5PT test as well as in the parents subjective scores in the BRIEF questionnaire Behavioral symptoms of attention difficulties are observed as inattention hyperactivity and impulsivity which are the primary symptoms associated with attention deficit hyperactivity disorder ADHD 31 These symptoms are overrepresented in children with TBI with 20 of children meeting ADHD criteria prior to the injury and an additional 20 of children experiencing new attention difficulties by two years post injury32 Interestingly no changes in attention measures were seen in cognitive tests in both groups However we found a significant improvement in all behavioral symptoms of attention difficulties in the Conners 3 questionnaire In order to evaluate intervention effect on both questionnaires and psychometric measures previous studies have used change from baseline However most of these measures do not have a minimal clinical important difference MCID value which somewhat complicates clinical value Thus a more valuable value to consider clinical effect is using Cohen s D effect size see Tables where an effect size of 0 2 is considered small 0 5 moderate and 0 8 large An effect size of 0 2 can be considered as MCID33 In our clinical perspective we consider an effect size of over 0 5 to be a meaningful clinically significant improvement Thus most of the significant statistical changes found in the HBOT compared to sham group can be considered as clinically significant The significant improvement in brain microstructure MD induced by HBOT in the current study was in certain cortical regions which correlate with the significant changes in cognitive function including executive function working memory and verbal fluency These regions have been previously associated with PPCS symptoms and cognitive dysfunction in PPCS patients The fusiform gyrus has been found to correlate with PPCS symptoms specifically executive function and emotional functioning34 35 Both the insula inferior frontal cortex and the supramarginal gyrus have been associated with working memory36 Regarding safety the relatively high rate of adverse events 86 90 of the patients is to be expected when treating children as our previous work has found a higher risk of 2 7 compared to adults37 Moreover children with cognitive dysfunction as well as ADHD provide additional challenges However all side effects were 2022 12 15233 Vol 1234567890

www nature com scientificreports HBOT Pre N Control Post Change Three months P value 13 Pre Post Change Three months P value P value baseline Net effect size 7 Conners 3 Inattention score 70 5 15 5 66 0 13 7 4 5 13 8 0 2666 71 0 13 6 68 1 11 6 2 9 5 9 0 2474 0 9424 0 136 Hyperactivity score 66 2 16 9 59 5 12 2 6 6 9 8 0 0319 66 9 18 6 62 6 14 8 4 3 8 9 0 2498 0 9361 0 244 Learning problems score 62 7 12 3 57 0 9 1 5 7 9 2 0 0446 71 4 15 6 71 0 12 5 0 4 8 4 0 8976 0 2080 0 59 Executive score 64 3 13 8 53 1 8 1 11 2 11 6 0 0044 68 6 16 5 59 3 11 7 9 3 14 8 0 1479 0 5675 0 153 Aggression score 63 7 19 1 55 2 14 5 8 5 14 7 0 0577 62 4 13 8 61 7 17 3 0 7 9 4 0 8478 0 8848 0 595 Peer relations score 67 1 18 4 65 2 17 2 1 9 9 9 0 4954 68 6 17 2 61 9 18 2 6 7 9 1 0 0973 0 8681 0 499 Inhibit 60 2 13 0 51 8 10 2 8 4 8 9 0 0052 63 4 17 4 59 6 16 7 3 9 4 7 0 0732 0 6567 0 586 Shift 62 9 15 6 58 2 13 1 4 7 9 9 0 1119 66 3 14 5 57 7 14 2 8 6 7 2 0 0197 0 6597 0 428 Self monitor 56 4 13 6 51 7 11 9 4 7 7 8 0 0504 54 6 13 7 52 4 15 4 2 1 4 8 0 2863 0 7908 0 367 Initiate 63 5 10 4 56 5 9 7 7 0 7 1 0 0039 63 0 9 9 62 6 13 4 0 4 10 6 0 9184 0 9164 0 781 Working memory 69 1 11 1 63 0 10 4 6 1 9 3 0 0371 71 9 9 0 68 0 6 5 3 9 6 8 0 1864 0 5957 0 258 Plan organize 65 3 13 9 57 6 12 7 7 7 8 5 0 0067 66 6 9 1 68 0 9 6 1 4 8 0 0 6548 0 8396 1 094 Or of materials 51 7 11 0 48 9 9 5 2 8 6 4 0 145 56 4 11 5 56 9 9 9 0 4 6 1 0 8594 0 4043 0 506 Task monitoring 56 2 13 9 52 6 11 1 3 5 7 9 0 1342 63 9 14 3 55 7 14 0 8 1 9 5 0 0637 0 2809 0 542 BRI 60 6 14 8 53 9 12 2 6 7 7 8 0 0093 62 3 17 0 57 0 17 4 5 3 4 9 0 028 0 8312 0 202 MI 63 8 12 2 57 4 11 8 6 4 5 8 0 0019 67 1 9 4 65 0 9 5 2 1 6 4 0 409 0 5522 0 706 GEC 63 3 13 3 56 3 12 1 7 0 6 2 0 0015 66 3 12 3 62 4 12 5 3 9 5 5 0 114 0 6475 0 528 BRIEF Table 4 Behavioral symptoms changes Baseline comparison of p values test the null hypothesis of equal means of the two groups at baseline using an unpaired t test three month comparison of p values test the null hypothesis of equal means of each group pre post intervention HBOT sham respectively using a paired t test bold P 0 05 net effect size is the subtraction of Cohen s D effect size of the control group from the HBOT group Cohen s D effect size BRI behavioral regulation index MI metacognition GEC global executive composite Figure 3 Conners scale for assessing ADHD questionnaire domains change Following HBOT the Conners 3 questionnaire showed significant improvements in the hyperactivity score executive score learning problem score and negative impression Normal range 40 59 P 0 05 P 0 01 See Table 4 for full statistical information 2022 12 15233 Vol 0123456789

www nature com scientificreports Figure 4 Behavior rating inventory of executive function questionnaire domains change Following HBOT the BRIEF questionnaire showed significant improvements in the inhibit initiate working memory planning behavioral regulation index metacognition and global executive composite scores Normal range 49 64 BRIEF behavior rating inventory of executive function P 0 05 P 0 01 See Table 4 for full statistical information HBOT Pre Control Post Change Three months P value Pre Post Change Three months P value P value baseline Net effect size 0 9 2 1 0 2042 0 6959 0 997 0 1 2 6 0 9074 0 3278 0 125 0 8321 0 5796 0 082 N 14 10 FIRM 4 7 3 0 3 4 2 9 1 3 2 3 0 0537 4 2 3 0 5 1 2 9 FOAM 5 6 3 2 6 1 4 2 0 5 5 8 0 7533 7 4 5 0 7 3 5 5 BESS 12 10 4 4 6 9 6 7 0 0 8 6 8 0 6714 11 9 8 2 11 6 8 1 0 3 4 3 Table 5 Neuro physical changes Baseline comparison of p values test the null hypothesis of equal means of the two groups at baseline using an unpaired t test three month comparison of p values test the null hypothesis of equal means of each group pre post intervention HBOT sham respectively using a paired t test bold P 0 05 net effect size is the subtraction of Cohen s D effect size of the control group from the HBOT group Cohen s D effect size MNI coordinates Anatomical location BA X Y Z t value Fusiform R 37 48 66 2 3 89 Cluster size 50 0 00028 P value Lingual R 18 11 67 0 3 52 122 0 00073 Insula L 13 42 2 7 3 03 44 0 00121 Supra marginal L 40 51 44 48 3 31 29 0 00128 Inferior frontal L 47 42 23 0 3 31 76 0 00128 Insula R 13 40 4 13 3 07 58 0 00238 Inferior frontal R 47 39 37 14 2 96 23 0 00307 Table 6 Brain regions with significant mean diffusivity MD changes The table reports each brain region found significant in a time by group repeated measures ANOVA comparing the two groups HBOT N 8 sham N 8 The results are in specific Montreal Neurological Institute MNI coordinates X sagittal Y coronal Z axial refer to MNI BA Brodmann area All coordinates emerged at a threshold of p 0 005 uncorrected Minimum cluster size 20 R right L left 2022 12 15233 Vol 1234567890

www nature com scientificreports Figure 5 Brain regions with significant post HBOT changes in mean diffusivity MD Group by time interaction ANOVA model in A DTI mean diffusivity MD in gray matter p 0 005 uncorrected B Significant correlation between change in phonemic fluency and the right lingual MD BA 18 and right insula MD BA 13 r is Pearson s correlation coefficient The 95 prediction interval is presented in the shaded area R right L left BA Brodmann area Brain images were created using BrainNet Viewer Software http www nitrc org projects bnv considered mild and handled conservatively without any long term sequala Notably the similar rate of side effects barotrauma in particular in the sham group provides validation to our sham protocol and blinding methods The majority of pediatric guidelines and previous studies focused on acute concussion management Early sub symptom threshold aerobic exercise has shown efficacy and speeds recovery in the first month post injury10 11 However the literature on persistent pediatrics PPCS remains limited with few randomized controlled trials and without an effective modality Both cognitive behavioral treatment and melatonin have been shown effective for sleep disturbance in pediatric PPCS but did not help in behavior or cognition38 39 Perceptual cognitive training did not show any benefit40 and transcranial direct current stimulation has yet to show positive effects41 Collaborative care intervention including cognitive behavioral treatment medications and care management have shown to reduce PPCS symptoms with a small effect size42 In the current study we found that HBOT s net effect size of global cognitive improvement was 0 598 medium effect size and the behavioral changes were at magnitudes of 0 5 1 medium large effect sizes These effects exceed any other suggested intervention and these effects were seen even after several years following the acute injury Evidence for HBOT s effectiveness in adult with PPCS has been accumulating in the past decade In our original adult RCT20 56 civilians with PPCS 1 6 years after the acute injury were included in a crossover design study The HBOT group which received 40 daily sessions at 1 5 ATA showed significant improvements in all cognitive functions memory executive functions attention and information processing speed The control group which received standard treatment had no significant change in any of the parameters Several Department of Defense DoD funded clinical trials followed with inconclusive findings due different methodological flaws43 45 Weaver et al 23 recently completed a trial funded by the DoD Va which included 71 active military servicemen and veterans who suffered from PCS more than three months to five years after mild TBI Participants were divided into two groups 40 daily HBOT sessions at 1 5 ATA or 1 2 ATA breathing air which cannot be considered placebo or inert given throughout 12 weeks The HBOT group had significant improvements in their PPCS symptoms questionnaires as well as the cognitive processing speed and sleep measures after 13 weeks but these results showed regression six months later Harch et al 24 randomized 50 military and civilian patients suffering from PPCS following mTBI to either 40 HBOT sessions at 1 5 ATA in eight weeks or an equivalent no treatment control period similar to the design by Boussi Gross et al 20 HBOT subjects experienced significant improvements in PPCS symptoms and cognitive function as well as depression anxiety sleep and quality of life metrics The current study is the first to evaluate the application of HBOT in pediatric PPCS Due to our accumulated experience with PPCS patients who further benefitted from 60 sessions21 our pediatric protocol included 60 sessions rather than 40 sessions Notably standing out from the above trials our study is the first to include a true inert sham intervention without repeated high pressure exposures The therapeutic mechanisms of HBOT in PPCS have been shown in both animal models and humans These include neuroplasticity by stimulating cell proliferation46 promoting neurogenesis of endogenous neural stem 2022 12 15233 Vol 0123456789

www nature com scientificreports cells47 regenerating axonal white matter48 improving maturation and myelination of injured neural fibers49 50 and stimulating axonal growth thus increasing the ability of neurons to function and communicate with each other51 52 This effect on neuronal microstructure has been demonstrated by our group using MRI DTI in adults with PPCS19 On the cellular level HBOT can improve cellular metabolism reduce apoptosis alleviate oxidative stress and restore mitochondrial function in both neurons and glial cells 47 53 54 In addition to neuroplasticity HBOT induces angiogenesis the creation of new blood vessels By inducing angiogenesis HBOT improves the cerebral vascular blood flow necessary for neurogenesis and synaptogenesis55 56 We have demonstrated this effect on cerebral blood flow in adults with PPCS using MRI DSC57 The study has several limitations and strengths worth mentioning First the sample size is considerably limited and underpowered as we were unable to recruit the needed number of eligible patients in four years The main reason was parents refusal to have their child participate in a sham controlled study Parents mentioned that after reading both the literature and study s consent form containing available data on HBOT effects in adults with PPCS they did not like the 50 chance of sham treatment This may serve as a possible setback to any future sham controlled study design in adult or pediatric PPCS However the presence of significant changes seen in a small group suggests the high potency of the intervention Second up to a third of patients used ADHD related drugs which were not an exclusion criterion as long as the medications were unchanged However these medications can play a role in both cognitive function and behavior symptoms Due the low number of patients a post hoc sensitivity analysis is underpowered to draw conclusions Third the duration of the effect is yet to be determined in long term follow ups Fourth the current protocol included one hour daily sessions without air breaks similar to our previous study in adults suffering from PPCS The use of longer daily sessions including air breaks which were shown to be beneficial in different adult populations remains to be determined in future studies Along with these limitations several strengths should be emphasized This was a first of its kind sham double blinded controlled study with a true inert control protocol and effective blinding of patients Secondly In addition to the subjective questionnaires filled by the parents we used objective evaluations with reliable test retest validity In summary the study suggests that HBOT improves cognitive and behavioral function PPCS and quality of life in children suffering from PPCS even years after injury Additional data is needed to optimize the protocol and to characterize the children who can benefit the most from this treatment Patients were 8 to 15 years old with mTBI loss of consciousness with duration of 0 30 min posttraumatic amnesia with duration of less than 24 h Glasgow Coma Scale GCS grade of 13 15 or moderate TBI GCS of 9 12 as well as additional criteria such as CT abnormalities within six months to 10 years prior to the inclusion in the study suffering from at least two post concussive syndrome symptoms PCS as measured by the post concussion symptom inventory for at least three months Included patients had no change in cognitive or behavioral functions for at least a month prior to the beginning of the study Patients were excluded if they had any dynamic neurologic improvement or worsening during the past month had been treated with HBOT for any other reason prior to their inclusion suffered from any chest pathology incompatible with pressure changes including asthma inner ear disease claustrophobia previous neurologic conditions e g epilepsy neuromuscular diseases metabolic diseases etc brain tumors skull base fractures active malignancy or encephalomalacia Patients who underwent intracranial surgery including ventricular drainage subdural hematomas drainage epidural hematomas drainage and intracerebral hemorrhage evacuation were excluded as well Lastly patients who were unable to perform awake brain MRIs were excluded After signing the informed consent by the parents patients underwent screening cognitive assessments where children over the age of ten underwent a computerized battery Neurotrax see below and children younger than 10 underwent pen and paper based tests Patients were included in the study with a decrease of at least one standard deviation SD in at least one of the following domains memory attention information processing speed executive function than expected per age and education level A prospective randomized double blind sham controlled trial was conducted from December 1st 2017 to November 1st 2021 at Shamir Medical Center SMC Israel Eligible patients were randomized to either HBOT or sham control groups in a 1 1 ratio according to a computerized randomization table supervised by a blinded researcher To evaluate participant masking patients were questioned after the first session on their perception regarding the treatment they received An evaluation procedure was done at baseline and 1 3 weeks after the last HBOT or sham session All evaluators were blinded to the patients group allocation The study was approved by SMC s Institutional Review Board IRB No 120 16 ASF and all participants parents signed an informed consent prior to their inclusion All research was performed according to the relevant guidelines and regulations This study was registered with ClinicalTrials gov number NCT03339037 on 13 11 2017 Both HBOT and sham protocols were administrated in a multi place Starmed 2700 chamber HAUX Germany The protocol comprised of 60 one day sessions five sessions per week within a three month period The HBOT protocol included breathing 100 oxygen by mask at 1 5 ATA for 60 min with no air breaks Compression decompression rates were 1 0 m minute The sham protocol included breathing 21 oxygen by mask at 1 03 ATA for 60 min To mask the controls the chamber pressure was raised up to 1 1 ATA during the first five minutes of the session along with circulating air noise followed by decompression 0 4 m minute to 1 03 ATA during the next five minutes 2022 12 15233 Vol 1234567890

www nature com scientificreports Cognitive function Computerized cognitive health assessments were evaluated by the NeuroTrax computerized cognitive testing battery NeuroTrax Corporation Bellaire TX 58 59 This assessment comprises of several cognitive tests that evaluate various aspects of brain capabilities including memory executive function attention information processing speed and motor skills The cognitive indices were based on scores of six cognitive tests verbal memory non verbal memory go no go test Stroop test staged information processing test and the catch game The cognitive domains are a combination the several sub tests where the global score is the mean value of the cognitive domains Outcome parameters were calculated using custom software blind to diagnosis or the testing site59 Domain scores were normalized for age and gender Additional information is also available on the NeuroTrax website http www neurotrax com Trail making test The trail making test children s version parts A and B60 measures mental set shifting attention and cognitive processing speed The test can provide information about visual search speed scanning speed of processing mental flexibility as well as executive functioning The task requires a subject to connect a sequence of 25 consecutive targets on a sheet of paper There are two parts to the test In the first part the targets are all numbers from 1 to 25 and the test taker needs to connect them in sequential order In the second part the dots go from 1 to 13 and include letters from A to L in Hebrew script Aleph to Lamed As in the first part the patient must connect the dots in order while alternating letters and numbers in the shortest time possible without lifting the pen from the paper Scoring is based on time taken to complete the test with lower scores being better The Wechsler Intelligence Scale for Children 4th Edition WISC IV Integrated The WISC IV61 is a psychometric measure of intelligence in children who are between 6 and 16 years of age We performed five subsets out of the 10 the WMI which included the digit span and letter number sequencing subtests the PSI which includes the coding and symbol search subtests and the cancellation The digit span and coding subtests are reliable and valid measures of short term memory working memory and speed of information processing in children and adults The digit span subtest measures the participant s ability to repeat numbers read aloud by the examiner in a forward and backward order The coding subtest requires the participant to reproduce symbols in a sequence as quickly as possible Test of memory and learning TOMAL The TOMAL62 is a memory battery that is designed for children 5 to 19 years old It is composed of 10 core subtests memory for stories word selective reminding object recall digits forward paired recall facial memory visual selective reminding abstract visual memory visual sequential memory and memory for location and four supplemental subtests letters forward digits backward letters backward and manual imitation The core subtests combine to produce a number of composite indices and the verbal memory index nonverbal memory index and attention concentration index were used in this study The Rey auditory verbal learning test RAVLT The RAVLT is a neuropsychological assessment designed to evaluate verbal memory in patients63 Five presentations of a 15 word list are given each followed by an attempted recall This is followed by a second 15 word list list B followed by recalling of list A Delayed recall and recognition are also tested A key feature of the RAVLT is that it affords the opportunity to measure rates of learning as opposed to recall of a single stimulus or series of stimuli The five points test 5PT The 5PT64 is a structured and standardized test that assesses figural fluency functions which are associated with executive functioning A participant is asked to generate as many unique designs as possible in a certain time limit The number of completed correct designs are counted after one and two minutes have elapsed Verbal fluency Verbal fluency is a cognitive function that facilitates information retrieval from memory Successful retrieval requires executive control over cognitive process such as selective attention mental set shifting internal response generation and self monitoring The FAS test65 measures both semantic and phonemic word fluency It assesses phonemic fluency by requesting an individual to orally produce as many words as possible that begin with specific letters within a prescribed time frame usually one minute The semantic fluency test requires a subject to generate as many words as possible from a given semantic category e g fruits within a limited time usually one minute The NEPSY Second Edition NEPSY II The NEPSY II is a standardized neuropsychological battery for children aged 3 16 years that assesses functioning across six domains executive functioning and attention memory and learning sensorimotor functioning social perception language and visuospatial processing66 The NEPSY II yields both raw and standardized scores by age and has adequate to high reliability including internal consistency and test retest reliability validity and interscorer agreement For the purpose of this study we administered the animal sorting subtest from the NEPSY II battery used to test the domains of attention and executive functioning Each subtest used in this study includes a teaching example or learning trial in which the examiner is able to correct the child s errors and to ensure that the child understands the task The subsets yield a mean standard score of 10 with a standard deviation of three Post concussion symptoms The British Columbia post concussion symptom inventory BC PSI The BC PSI is a 16 item measure designed to assess the presence and severity of PCS symptoms Parents were asked to rate the frequency and intensity of 13 symptoms over the past two weeks including that day The symptoms included items such as headache poor sleep temper problems and poor concentration The psychological symptoms were rated on a Likert type scale measuring frequency from 0 not at all to 5 constantly and the intensity ratings 2022 12 15233 Vol 0123456789

www nature com scientificreports ranged from 0 not at all to 5 very severe problem 67 Scoring was calculated by multiplying frequency by intensity to create a single score for each item and this score was then converted to a score on a scale of 0 4 The produced score of 0 1 was considered 0 2 3 1 4 6 2 8 12 3 15 4 These scores represent severity whereby 0 none 1 2 mild and 3 moderate or greater Similarly total score representations are 0 low 1 9 normal 10 14 unusually high and 15 extremely high relative to healthy adults The internal consistency reliability for the items comprising the total score is r 0 8267 68 Health and behaviour inventory HBI The HBI is a 20 item scale that measures the frequency of different symptoms that are associated with concussion69 Parents completed the child version of the HBI by rating the frequency of symptoms experienced on a 4 point Likert scale as 0 never 1 rarely 2 sometimes and 3 often In addition to individual symptom ratings a total symptom score was calculated as the sum of all items in which higher scores indicate more frequent symptoms Quality of life Paediatric quality of life inventory PEDSQL The PedsQL 4 0 is a reliable and valid measure of health related quality of life HRQoL in healthy children and those with acute and or chronic health conditions70 Parents were asked to complete the parents versions which exist for children aged 2 18 years in four age groups Items are calculated and transformed into an overall score with a range of 0 to 100 points with more points indicating better HRQoL Secondary outcomes included the PedsQL 4 0 subscale scores physical emotional social and school The MCID is estimated at 4 88 6 2771 Behavioural assessment Conner s Rating Scale The Conner s Rating Scale is a reliable and valid parent questionnaire assessing their child s functioning in emotional social cognitive and behavioral domains The emphasis of this measure is attentional in nature72 and is useful in monitoring change in behavior across the day73 The Conner s rating scale contains four component subscales ADHD index hyperactivity cognition and appositional The ADHD index subscale contains 12 items three of which are also used in the 6 item cognition problems inattentive subscale74 The hyperactivity cognition and appositional subscales each contain six items The ADHD index subscale provides an indicator of children likely to have attentional problems The hyperactivity scale assesses both the hyperactivity and impulsivity domains of ADHD symptoms The appositional subscale assesses appositional behaviours The cognition problems inattention subscale assesses traits that may lead to difficulties in school academic performance74 Compared with clinicians and teachers parents have more opportunities throughout the day to observe the impact of ADHD on quality of life during activities that are school related such as homework and non school related such as chores or play 74 Behavioural rating inventory of executive function BRIEF BRIEF is a reliable and valid parent questionnaire assessing their child s executive functioning75 The questionnaire was designed for parents of school aged children that enables professionals to assess executive function behaviours at home It contains 86 items within eight theoretically and empirically derived clinical scales that measure the different aspects of EF inhibition shift emotional control initiate working memory planning organization organization of materials and monitor75 Neuro physical evaluation The balance error scoring system BESS is a useful means of measuring postural stability which frequently exhibits deficits after a concussion76 Patients were tested in three positions feet touching side by side a single leg stance on the nondominant leg and a heel to toe stance with the dominant foot in front for 20 s each on a firm surface and then on a 50 cm 41 cm 6 cm thick foam pad The dominant leg was identified by asking subjects which foot they would use to kick a ball Patients were instructed to close their eyes and place hands on hips for 20 s Subjects were also told that the clock would not stop running during testing so if they ever came out of the starting position they should make any necessary adjustments and return to the position as quickly as possible One error point was given every time the subject moved hands off hips opened eyes stepped stumbled abducted or flexed the hip greater than 30 lifted the forefoot or heel from the testing surface or remained out of the testing position for more than five seconds The maximum score for any one trial was 10 points If a subject could not maintain one of the positions for a minimum of five seconds at any point during a trial we assigned the maximum score of 10 for that trial We calculated the total BESS score sum of scores from all six trials the firm surface score sum of scores from the three firm surface trials and foam surface score sum of scores from the three foam surface trials Brain imaging Brain imaging MRI scans were performed on a MAGNETOM Skyra 3 T scanner configured with 20 channel receiver head coils Siemens Healthcare Erlangen Germany The MRI protocol included T2 weighted 3D fluid attenuated inversion recovery FLAIR susceptibility weighted imaging SWI pre and post contrast high resolution MPRAGE 3D T1 weighted and diffusion tensor imaging DTI Whole brain diffusion weighted images were acquired with the following parameters 50 axial slices slice thickness 2 2 mm voxel size 1 5 1 5 mm TR 9300 ms TE 91 ms and matrix 128 128 mm 2022 12 15233 Vol 1234567890

www nature com scientificreports Diffusion gradients were applied along 30 noncollinear directions b 1000 s mm2 and one volume without diffusion weighting Preprocessing of DTI images was preformed using the SPM software version 12 UCL London UK and included motion correction co registration with MPRAGE T1 images spatial normalization and spatial smoothing with kernel size of 6 mm full width half maximum FWHM Diffusion brain volume denoising was performed using the joint anisotropic LMMSE filter for stationary rician noise removal77 and calculation of DTI FA fractional anisotropy and MD mean diffusivity maps were performed using in house software written in MATLAB R2021b MathWorks Natick MA Safety Participants were monitored for adverse events including barotraumas either ear or sinuses and oxygen toxicity pulmonary and central nervous system Continuous data are expressed as means standard deviations SD Two tailed independent t tests were performed to compare variables between groups when a normality assumption held according to a Kolmogorov Smirnov test Net effect sizes were evaluated using Cohen s d method defined as the improvement from baseline after sham intervention was subtracted from the improvement after HBOT divided by the pooled standard deviation of the composite score Categorical data are expressed in numbers and percentages compared by chi square Fisher s exact tests A value of p 0 05 was considered significant Imaging data analysis was performed on the normalized FA and MD maps using the voxel based method to generate statistical parametric maps A gray matter mask was applied on the MD maps and a white matter mask on the FA maps using a threshold of 0 2 A within subject repeated measure ANOVA model was used to test the main interaction effect between time and group implemented in SPM software version 12 UCL London UK Correlations between MD changes and cognitive scores were performed using Pearson s correlation Data analysis was performed using MATLAB R2021b MathWorks Natick MA Statistics Toolbox Sample size The estimated sample size was calculated based on test reliability in healthy volunteers78 A NeuroTrax global cognitive score improvement of 2 in the placebo and 12 in the treatment arm with a mean score of 100 and standard deviation of 14 to demonstrate a 10 effect Assuming a power of 80 and 5 twosided level of significance a total of 62 participants would be required 31 participants in each arm Considering a dropout rate of 15 the total sample size required is 70 The datasets analyzed during the current study are available from the corresponding author on reasonable request Received 13 June 2022 Accepted 29 August 2022 1 Barlow K M et al Epidemiology of postconcussion syndrome in pediatric mild traumatic brain injury Pediatrics 126 2 e374 e381 2010 2 Langlois J A Rutland Brown W Thomas K E The incidence of traumatic brain injury among children in the United States Differences by race J Head Trauma Rehabil 20 3 229 238 2005 3 McKinlay A et al Prevalence of traumatic brain injury among children adolescents and young adults Prospective evidence from a birth cohort Brain Inj 22 2 175 181 2008 4 Zemek R L Yeates K O Rates of persistent postconcussive symptoms JAMA 317 13 1375 1376 2017 5 Ganesalingam K et al Family burden and parental distress following mild traumatic brain injury in children and its relationship to post concussive symptoms J Pediatr Psychol 33 6 621 629 2008 6 Novak Z et al Association of persistent postconcussion symptoms with pediatric quality of life JAMA Pediatr 170 12 e162900 2016 7 Fried E et al Persistent post concussive syndrome in children after mild traumatic brain injury is prevalent and vastly underdiagnosed Sci Rep 12 1 4364 2022 8 McCrory P et al Consensus statement on concussion in sport the 5 th international conference on concussion in sport held in Berlin October 2016 Br J Sports Med 51 11 838 847 2017 9 Grool A M et al Association between early participation in physical activity following acute concussion and persistent postconcussive symptoms in children and adolescents JAMA 316 23 2504 2514 2016 10 Leddy J J et al Early subthreshold aerobic exercise for sport related concussion A randomized clinical trial JAMA Pediatr 173 4 319 325 2019 11 Leddy J J et al Early targeted heart rate aerobic exercise versus placebo stretching for sport related concussion in adolescents A randomised controlled trial Lancet Child Adolesc Health 5 11 792 799 2021 12 Barlow K M Postconcussion syndrome A review J Child Neurol 31 1 57 67 2016 13 Schneider K J et al Cervicovestibular rehabilitation in sport related concussion A randomised controlled trial Br J Sports Med 48 17 1294 1298 2014 14 Leddy J J et al Exercise treatment for postconcussion syndrome a pilot study of changes in functional magnetic resonance imaging activation physiology and symptoms J Head Trauma Rehabil 28 4 241 249 2013 15 Gagnon I Galli C Friedman D Grilli L Iverson G L Active rehabilitation for children who are slow to recover following sport related concussion Brain Inj 23 12 956 964 2009 16 Barlow K M et al Efficacy of melatonin in children with postconcussive symptoms A randomized clinical trial Pediatrics 145 4 1 2020 17 Hadanny A Efrati S The hyperoxic hypoxic paradox Biomolecules 10 6 1 2020 18 Efrati S Ben Jacob E Reflections on the neurotherapeutic effects of hyperbaric oxygen Expert Rev Neurother 14 3 233 236 2014 2022 12 15233 Vol 0123456789

www nature com scientificreports 19 Tal S Hadanny A Sasson E Suzin G Efrati S Hyperbaric oxygen therapy can induce angiogenesis and regeneration of nerve fibers in traumatic brain injury patients Front Hum Neurosci 11 508 2017 20 Boussi Gross R et al Hyperbaric oxygen therapy can improve post concussion syndrome years after mild traumatic brain injury randomized prospective trial PLoS ONE 8 11 e79995 2013 21 Hadanny A Abbott S Suzin G Bechor Y Efrati S Effect of hyperbaric oxygen therapy on chronic neurocognitive deficits of post traumatic brain injury patients Retrospective analysis BMJ Open 8 9 e023387 2018 22 Gottfried I Schottlender N Ashery U Hyperbaric oxygen treatment from mechanisms to cognitive improvement Biomolecules 11 10 1 2021 23 Weaver L K et al Hyperbaric oxygen for post concussive symptoms in United States military service members A randomized clinical trial Undersea Hyperb Med 45 2 129 156 2018 24 Harch P G et al Hyperbaric oxygen therapy for mild traumatic brain injury persistent postconcussion syndrome A randomized controlled trial Med Gas Res 10 1 8 20 2020 25 McAllister T W Flashman L A McDonald B C Saykin A J Mechanisms of working memory dysfunction after mild and moderate TBI Evidence from functional MRI and neurogenetics J Neurotrauma 23 10 1450 1467 2006 26 Kumar S Rao S L Chandramouli B A Pillai S Reduced contribution of executive functions in impaired working memory performance in mild traumatic brain injury patients Clin Neurol Neurosurg 115 8 1326 1332 2013 27 Toledo E et al The young brain and concussion Imaging as a biomarker for diagnosis and prognosis Neurosci Biobehav Rev 36 6 1510 1531 2012 28 Muscara F Catroppa C Anderson V The impact of injury severity on executive function 7 10 years following pediatric traumatic brain injury Dev Neuropsychol 33 5 623 636 2008 29 Mangeot S Armstrong K Colvin A N Yeates K O Taylor H G Long term executive function deficits in children with traumatic brain injuries Assessment using the Behavior Rating Inventory of Executive Function BRIEF Child Neuropsychol 8 4 271 284 2002 30 Wilkinson A A et al Brain biomarkers and pre injury cognition are associated with long term cognitive outcome in children with traumatic brain injury BMC Pediatr 17 1 173 2017 31 Yeates K O et al Long term attention problems in children with traumatic brain injury J Am Acad Child Adolesc Psychiatry 44 6 574 584 2005 32 Gerring J P et al Premorbid prevalence of ADHD and development of secondary ADHD after closed head injury J Am Acad Child Adolesc Psychiatry 37 6 647 654 1998 33 Copay A G Subach B R Glassman S D Polly D W Jr Schuler T C Understanding the minimum clinically important difference A review of concepts and methods Spine J 7 5 541 546 2007 34 Mortaheb S et al Neurophysiological biomarkers of persistent post concussive symptoms A scoping review Front Neurol 12 687197 2021 35 Killgore W D S et al Gray matter volume and executive functioning correlate with time since injury following mild traumatic brain injury Neurosci Lett 612 238 244 2016 36 Smits M et al Postconcussion syndrome after minor head injury Brain activation of working memory and attention Hum Brain Mapp 30 9 2789 2803 2009 37 hadanny A Meir O Bechor Y Fishlev G Bergan J Efrati S The safety of hyperbaric oxygen treatment retrospective analysis in 2334 patients Undersea Hyperb Med 43 2 113 122 2016 38 Barlow K M et al Efficacy of melatonin for sleep disturbance in children with persistent post concussion symptoms secondary analysis of a randomized controlled trial J Neurotrauma 38 8 950 959 2021 39 Tomfohr Madsen L et al A pilot randomized controlled trial of cognitive behavioral therapy for insomnia in adolescents with persistent postconcussion symptoms J Head Trauma Rehabil 35 2 E103 E112 2020 40 Teel E Brossard Racine M Corbin Berrigan L A Gagnon I Perceptual cognitive training does not improve clinical outcomes at 4 and 12 weeks following concussion in children and adolescents A randomized controlled trial J Head Trauma Rehabil 36 2 E97 E107 2021 41 Quinn de Launay K Cheung S T Riggs L Reed N Beal D S The effect of transcranial direct current stimulation on cognitive performance in youth with persistent cognitive symptoms following concussion a controlled pilot study Brain Inj 36 1 39 51 2022 42 McCarty C A et al Collaborative care for adolescents with persistent postconcussive symptoms A randomized trial Pediatrics 138 4 1 2016 43 Cifu D X et al Effects of hyperbaric oxygen on eye tracking abnormalities in males after mild traumatic brain injury J Rehabil Res Dev 51 7 1047 1056 2014 44 Cifu D X Hart B B West S L Walker W Carne W The effect of hyperbaric oxygen on persistent postconcussion symptoms J Head Trauma Rehabil 29 1 11 20 2014 45 Miller R S et al Effects of hyperbaric oxygen on symptoms and quality of life among service members with persistent postconcussion symptoms A randomized clinical trial JAMA Intern Med 175 1 43 52 2015 46 Mu J et al Delayed hyperbaric oxygen therapy induces cell proliferation through stabilization of cAMP responsive element binding protein in the rat model of MCAo induced ischemic brain injury Neurobiol Dis 51 133 143 2013 47 Yang Y J et al Hyperbaric oxygen induces endogenous neural stem cells to proliferate and differentiate in hypoxic ischemic brain damage in neonatal rats J Undersea Hyperbaric Med Soc 35 2 113 129 2008 48 Chang C C et al Damage of white matter tract correlated with neuropsychological deficits in carbon monoxide intoxication after hyperbaric oxygen therapy J Neurotrauma 26 8 1263 1270 2009 49 Vilela D S Lazarini P R Da Silva C F Effects of hyperbaric oxygen therapy on facial nerve regeneration Acta Otolaryngol 128 9 1048 1052 2008 50 Haapaniemi T Nylander G Kanje M Dahlin L Hyperbaric oxygen treatment enhances regeneration of the rat sciatic nerve Exp Neurol 149 2 433 438 1998 51 Bradshaw P O Nelson A G Fanton J W Yates T Kagan Hallet K S Effect of hyperbaric oxygenation on peripheral nerve regeneration in adult male rabbits Undersea Hyperbaric Med J Undersea Hyperbaric Med Soc 23 2 107 113 1996 52 Mukoyama M Iida M Sobue I Hyperbaric oxygen therapy for peripheral nerve damage induced in rabbits with clioquinol Exp Neurol 47 3 371 380 1975 53 Zhang J H Lo T Mychaskiw G Colohan A Mechanisms of hyperbaric oxygen and neuroprotection in stroke Pathophysiology 12 1 63 77 2005 54 Calvert J W Cahill J Zhang J H Hyperbaric oxygen and cerebral physiology Neurol Res 29 2 132 141 2007 55 Chen J et al Intravenous administration of human bone marrow stromal cells induces angiogenesis in the ischemic boundary zone after stroke in rats Circ Res 92 6 692 699 2003 56 Jiang Q et al Investigation of neural progenitor cell induced angiogenesis after embolic stroke in rat using MRI Neuroimage 28 3 698 707 2005 57 Tal S et al Hyperbaric oxygen may induce angiogenesis in patients suffering from prolonged post concussion syndrome due to traumatic brain injury Restor Neurol Neurosci 33 6 943 951 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www nature com scientificreports 58 Doniger GM Mindstreams Computerized Cognitive Tests Test Descriptions Available http www mirror upsite co il uploaded files 1383_e7d7d3d98c924f036d3123733419149d pdf Accessed 05 July 2013 2007 Available from http www mirror upsite co il uploaded files 1383_e7d7d3d98c924f036d3123733419149d pdf 59 Doniger GM Guide to MindStreams Normative Data Available http www mirror upsite co il uploaded files 1383_b44d4786c9 1058be301cb09a94ba70f4 pdf Accessed 05 July 2013 2012 Available from http www mirror upsite co il uploaded files 1383_ b44d4786c91058be301cb09a94ba70f4 pdf 60 Espy K A Cwik M F The development of a trial making test in young children The TRAILS P Clin Neuropsychol 18 3 411 422 2004 61 Grizzle R Wechsler intelligence scale for children Fourth Edition In Encyclopedia of child behavior and development eds Goldstein S Naglieri J A 1553 1555 Springer US 2011 62 Ritchie D Nierenberg B Test of Memory and Learning In Encyclopedia of child behavior and development eds Goldstein S Naglieri J A 1479 1480 Springer US 2011 63 Markel Fox S Hooper S R The Rey auditory verbal learning test RAVLT Preliminary normative data for children ages 6 to 12 Arch Clin Neuropsychol 7 4 347 1992 64 Tucha L Aschenbrenner S Koerts J Lange K W The five point test Reliability validity and normative data for children and adults PLoS ONE 7 9 e46080 2012 65 Barry D Bates M E Labouvie E FAS and CFL forms of verbal fluency differ in difficulty A meta analytic study Appl Neuropsychol 15 2 97 106 2008 66 Holcomb M J Davis A S NEPSY II In Encyclopedia of child behavior and development eds Goldstein S Naglieri J A 1006 1008 Springer US 2011 67 Iverson G L Lange R T Examination of postconcussion like symptoms in a healthy sample Appl Neuropsychol 10 3 137 144 2003 68 Sady M D Vaughan C G Gioia G A Psychometric characteristics of the postconcussion symptom inventory in children and adolescents Arch Clin Neuropsychol 29 4 348 363 2014 69 Levant R F Wimer D J Williams C M An evaluation of the Health Behavior Inventory 20 HBI 20 and its relationships to masculinity and attitudes towards seeking psychological help among college men Psychol Men Masculin 12 1 26 41 2011 70 Desai A D et al Validity and responsiveness of the pediatric quality of life inventory PedsQL 4 0 generic core scales in the pediatric inpatient setting JAMA Pediatr 168 12 1114 1121 2014 71 Jayadevappa R Cook R Chhatre S Minimal important difference to infer changes in health related quality of life a systematic review J Clin Epidemiol 89 188 198 2017 72 Conners C Pitkanen J Rzepa SJEoCNNY NY Springer Conners 3rd edition conners 3 conners 2008 675 678 2011 73 Conners C K Sitarenios G Parker J D Epstein J N The revised Conners Parent Rating Scale CPRS R Factor structure reliability and criterion validity J Abnorm Child Psychol 26 4 257 268 1998 74 Mueller F Brozovich R Johnson C B Conners rating scales revised CRS R 24 1 4 83 97 1999 75 Gioia G A Isquith P K Guy S C Kenworthy L J C N Test review behavior rating inventory of executive function 6 3 235 238 2000 76 Bell D R Guskiewicz K M Clark M A Padua D A Systematic review of the balance error scoring system Sports Health 3 3 287 295 2011 77 Trist n Vega A Aja Fern ndez S DWI filtering using joint information for DTI and HARDI Med Image Anal 14 2 205 218 2010 78 Dwolatzky T et al Validity of a novel computerized cognitive battery for mild cognitive impairment BMC Geriatr 3 1 4 2003 We would like to thank Dr Mechael Kanovsky for his editing of this manuscript All authors contributed substantially to the preparation of this manuscript A H S E were responsible for protocol design A H S Y O S Y Z Y B and S E were responsible for patient recruitment A H S Y O S L R A S G S E L S F N P G F R T M A R E G Y Z Y B were responsible for data acquisition A H M C E Sa A S and E S were responsible for data analysis All authors interpreted the data A H M C and E S wrote the manuscript All authors revised and finalized the manuscript The study was supported by the research grant from Ettie Gad Propper AH ES SG BY ZY work for AVIV Scientific LTD ES is a shareholder at AVIV Scientific LTD Other authors don t have any conflict of interest Correspondence and requests for materials should be addressed to A H Reprints and permissions information is available at www nature com reprints Publisher s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations 2022 12 15233 Vol 0123456789

www nature com scientificreports Open Access This article is licensed under a Creative Commons Attribution 4 0 International License which permits use sharing adaptation distribution and reproduction in any medium or format as long as you give appropriate credit to the original author s and the source provide a link to the Creative Commons licence and indicate if changes were made The images or other third party material in this article are included in the article s Creative Commons licence unless indicated otherwise in a credit line to the material If material is not included in the article s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use you will need to obtain permission directly from the copyright holder To view a copy of this licence visit http creativecommons org licenses by 4 0 The Author s 2022 2022 12 15233 Vol 1234567890

Editorial Reflections on the neurotherapeutic effects of hyperbaric oxygen Expert Rev Neurother 14 3 233 236 2014 Shai Efrati Author for correspondence The Institute of Hyperbaric Medicine Assaf Harofeh Medical Center Zerifin Israel and Research and Development Unit Assaf Harofeh Medical Center Zerifin Israel and Sackler School of Medicine Tel Aviv University Tel Aviv Israel and Sagol School of Neuroscience Tel Aviv University Tel Aviv Israel Tel 972 089 779 3935 Fax 972 089 204 989 efratishai 013 net Eshel Ben Jacob Author for correspondence The Institute of Hyperbaric Medicine Assaf Harofeh Medical Center Zerifin Israel and Research and Development Unit Assaf Harofeh Medical Center Zerifin Israel and Sagol School of Neuroscience Tel Aviv University Tel Aviv Israel and The Raymond and Beverly Sackler Faculty of Exact Sciences School of Physics and Astronomy Tel Aviv University Tel Aviv Israel and Center for Theoretical Biological Physics Rice University Houston TX USA Tel 972 036 407 845 Fax 001 713 247 8162 eshel rice edu Traumatic brain injury TBI and stroke are the major causes of brain damage and chronic neurological impairments There is no agreed upon effective metabolic intervention for TBI and stroke patients with chronic neurological dysfunction Clinical studies published this year present convincing evidence that hyperbaric oxygen therapy HBOT might be the coveted neurotherapeutic method for brain repair Here we discuss the multi faceted role of HBOT in neurotherapeutics in light of recent persuasive evidence for HBOT efficacy in brain repair and the new understanding of brain energy management and response to damage We discuss optimal timing of treatment dosage suitable candidates and promising future directions The challenge Traumatic brain injury TBI stroke and age related metabolic brain disorders are the major causes of brain damage and chronic neurological impairments Today there is no agreed upon effective metabolic treatment intervention in the routine clinical practice for TBI and stroke patients with chronic neurological dysfunction Intensive therapy and rehabilitation programs are valuable for improving quality of life right after brain injury but often provide only partial relief and leave the patients chronically disabled With the aging of the population the severity of the problem posed by stroke TBI mainly due to home accidents and metabolic disorders which can lead to dementia and Alzheimer s disease is expected to increase Experts agree that novel neurotherapeutic methods to repair and protect the brain from damage caused by these insults are needed more than ever before New hope Clinical studies published this year present convincing evidence that hyperbaric oxygen therapy HBOT might be the coveted neurotherapeutic method for brain repair 1 2 HBOT is a treatment in which oxygen enriched air up to 100 is administrated to patients in a chamber where the pressure is elevated above one atmosphere absolute 1ATA which is the ambient atmospheric pressure It is becoming widely acknowledged that the combined action of hyperoxia and hyperbaric pressure leads to significant improvement in tissue oxygenation while targeting both oxygenand pressure sensitive genes 3 6 resulting in improved mitochondrial metabolism with anti apoptotic and anti inflammatory effects 7 12 Here we reflect on the multifaceted role of HBOT in neurotherapeutics in light of recent persuasive evidence for HBOT efficacy in brain repair and of new understanding of brain energy management and response to damage We discuss the optimal timing of treatment optimal dose response curve oxygenpressure levels suitable candidates and promising future directions A generation of debate The idea that HBOT can provide a valuable tool for brain repair was first proposed almost half a century ago and has been considered anecdotal ever since Interest was renewed in the mid 90s 8 13 but the results were either ignored or seriously questioned by the medical KEYWORDS hyperbaric oxygen neuroplasticity neurotherapeutic stroke traumatic brain injury informahealthcare com 10 1586 14737175 2014 884928 2014 Informa UK Ltd ISSN 1473 7175 233

Editorial Efrati Ben Jacob community Huang and Obenaus in their 2011 review presented an objective summary of the clinical trials and associated debate till 2011 and a thoughtful description of the animal trials and their implications 8 They reasoned that the HBOTinduced neuroprotection in animal model is due to the observed improved tissue oxidation improved mitochondrial redox preservation of mitochondria integrity hindering of mitochondria associated apoptotic pathways as well as antiinflammatory effects 8 Until 2011 all human HBOT studies involved severe TBI patients While mortality was decreased in those studies there was no significant change in the quality of life This combined with knowledge from a wealth of animal studies indicated that the time of treatment and dose response curve should be reassessed Most importantly it led to the understanding that HBOT should be practiced on mild tomoderate TBI patients who are more apt to achieve clinically meaningful recovery Pre and post treatment SPECT imaging showed that HBOT led to restoration of neuronal activity in stunned areas A second randomized prospective trial published earlier this year used a similar crossover approach and presented equally persuasive evidence that HBOT can also revitalize chronically impaired brain function and significantly improve the quality of life of post stroke patients even years after the event 1 The participants in this study suffered a stroke 6 36 months prior to the trial They were also randomly divided into treated and control group went through brain function and quality evaluations These stroke patients were also treated with 40 HBOT daily sessions However while the mTBI patients were treated at a lower pressure of 1 5ATA since they all had an intact macrovascular bed the stroke patients were given 90 min sessions at 2 0ATA and 100 oxygen The results of SPECT imaging were well correlated with clinical improvements and revealed restored activity mostly in stunned areas in the surroundings of necrotic foci Persuasive new evidence Convincing evidence that HBOT can revitalize chronically impaired brain functions and significantly improve the quality of life of mild TBI mTBI patients with prolonged postconcussion syndrome at late chronic stage even years after injury are presented in a new randomized prospective trial published this year 2 A crossover approach was adopted in order to overcome the HBOT inherent sham control constraint discussed further below The participants who had suffered mTBI 1 5 years prior to the trial were randomly divided into two groups trial and control The trial group patients received 2 months of HBOT while the control group went without treatment in those 2 months The latter were given the same treatment as the trial group 2 months later The advantage of the crossover approach is the opportunity for a triple comparison between treatments of two groups between treatment and no treatment periods of the same group and between treatment and no treatment in different groups It also overcomes the problem posed by the impossibility of making people believe they are exposed to high pressure when they are actually not The treatment consisted of 40 daily sessions lasting 1 h at pressure of 1 5ATA and breathing 100 oxygen The patients cognitive functions and quality of life were assessed by detailed computerized evaluations and compared for all patients with single photon emission computed tomography SPECT scans HBOT sessions led to similar significant improvements in tests of cognitive function and quality of life in both groups No significant improvements occurred by the end of the nontreatment period in the control group Analysis of brain imaging showed significantly increased neuronal activity after a 2 month period of HBOT compared with the control period What makes the results particularly persuasive is the remarkable agreement between the cognitive function restoration and the changes in brain functionality as detected by the SPECT scans The diffuse nature of the mTBI injury renders the pathological damage hard to detect by common neuroimaging methods such as computed tomography and MRI 234 HBOT can activate cerebral plasticity revitalize chronically impaired brain functions The new trials provide convincing evidence that HBOT can induce cerebral plasticity leading to repair of chronically impaired brain functions and improved quality of life in poststroke patients and mTBI patients with prolonged postconcussion syndrome even years after the brain insult 1 2 The term cerebral plasticity is used here as an umbrella term that encompasses both neuroplasticity as commonly used in neuroscience and beyond synaptic changes such as myelinization regeneration of axonal white matter angiogenesis and changes in the glial fabric The observed restoration of neuronal activity in the metabolically dysfunctional stunned areas indicate HBOT as a potent means of delivering to the brain sufficient oxygen needed for activation of neuroplasticty and restoration of impaired functions These are entailed via assortment of intricate mechanisms some of which are mentioned below Underlying repair mechanisms Brain insults may result in a variety of brain injuries including impairment of microvascular integrity and cerebral perfusion These lead to reduced metabolism and neuronal activity which in turn lead to loss of synapses and tampered neuronal connectivity 2 8 9 The stunned areas mentioned earlier are characterized by anaerobic metabolism and ATP depletion culminating in stagnation and shortage of energy for the healing processes and they may persist like this dysfunctional but alive for years after injury 2 8 9 The decreased oxygen level not only causes reduction in neuronal activity but also prevents the generation of new synaptic connections and angiogenesis HBOT can initiate vascular repair and improve cerebral vascular flow induce regeneration of axonal white matter stimulate axonal growth promote blood brain barrier integrity and reduce inflammatory reactions as well as brain edema 7 12 At the cellular level HBOT can improve cellular metabolism reduce apoptosis alleviate oxidative stress and increase Expert Rev Neurother 14 3 2014

Reflections on the neurotherapeutic effects of hyperbaric oxygen levels of neurotrophins and nitric oxide through enhancement of mitochondrial function in both neurons and glial cells and may even promote neurogenesis of endogenous neural stem cells 7 12 14 Regarding mitochondria it is important to note that stroke and TBI engender depolarization of the mitochondria membrane and induction of mitochondrial permeability transition pore which reduces the efficiency of energy production and elevate the level of reactive oxygen species 15 HBOT can inhibit mitochondrial permeability transition pore and thus has the potential to reverse this abnormality 8 However it must be applied carefully to ascertain that the increased tissue oxygen does not cause cellular toxicity due to overly high reactive oxygen species levels Time of treatment Many innate repair mechanisms each with a different characteristic time are activated following the onset of acute brain injury and some may be negatively affected by premature application of HBOT HBOT procedure can begin either at the degenerative or at the regenerative stage At the degenerative stage it must be administered with great care to avoid toxicity On the other hand elevated oxygen levels during the regenerative stage would supply the energy needs for the innate brain repair processes While it is not possible to mark a clear line between the regenerative and the degenerative phases 16 it is quite clear that more than 1 month after the acute event in a stable patient the degenerative process has ended The differences in initiation times and protocols of HBOT may explain contradictive results in previous studies where HBOT timing was not taken into consideration 1 2 17 21 Dose response curve treatment duration The minimal effective dosages of the active ingredients in HBOT pressure and oxygen concentration are still unknown and future studies are needed to test this issue by evaluating the optimal case specific dose response curves For example in the previously described trials 2 0ATA was used for post stroke patients and 1 5ATA was used for mTBI 1 2 There are many case reports illustrating significant effects with even small increases in air pressure including effects on the brain 22 The dose response curve is related to the inherent difficulties in handling the sham control and is a source of misinterpretation of clinical studies the minimal elevated pressure a patient can sense is 1 3 atmosphere which can induce more than 50 elevation in tissue oxygenation Since such oxygenation can have significant physiological effects 1 2 treatment with room air at 1 3ATA is not an ineffectual treatment as is required from a proper sham control At the same time over oxygenation in response to pressure above 2 0ATA can have an inhibitory effect or even focal toxicity It is conceivable that HBOT above two atmospheres can be less effective than 1 3ATA explaining the unexpected improvements in control groups when 1 3ATA was used for sham control informahealthcare com Editorial The duration of treatment is also an unresolved issue It is quite clear that weeks to months would be necessary for brain tissue regeneration and angiogenesis but the upper time limit from which no further improvement is expected is still unknown More studies are needed to determine the minimal effective dosage and the duration for a specific brain injury Non invasive in chamber measurements are currently being developed specifically EEG and diffusion tensor imaging and may shed some light on this important question Clearly there is an urgent need for additional larger scale multi center clinical studies to further confirm the findings and determine the most effective and personalized treatment protocols To guarantee effective and well designed clinical studies wide scale biomedical research is required Such research will also provide validation of the clinical findings crucial aid in interpretation of the results and important clues to additional applications of HBOT Optimal candidates for HBOT Since hyperbaric oxygen therapy is the only treatment proven to significantly benefit post stroke and mTBI patients without limiting side effects it is reasonable to allow the millions of these patients to benefit from it right away and not wait for rigorous studies The classical candidate for HBOT is a patient with unrecovered brain injury where tissue hypoxia is the limiting factor for the regeneration possesses In this patient HBOT may induce neuroplasticity in the stunned regions where there is a brain anatomy physiology e g SPECT computed tomography mismatch 1 2 The anatomical physiological imaging should serve as part of the basic evaluation of every HBOT candidate just like transcutaneous oximetry at the ulcer bed serves as a basic evaluation for patients suffering from peripheral non healing wounds 23 24 Looking ahead Based on the aforementioned rationale one can surmise that HBOT could also be effective in early stages of vascular dementia Alzheimer s disease and other conditions where the clinical presentation could not be fully explained by anatomical imaging However this will require novel basic biomedical research to understand the HBOT effect on the recently discovered mitochondria associated cellular response to hypoxia 7 12 which is a common denominator of stroke TBI dementia and aging 25 26 Financial competing interests disclosure The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript This includes employment consultancies honoraria stock ownership or options expert testimony grants or patents received or pending or royalties No writing assistance was utilized in the production of this manuscript 235

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