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The effects of intermittent exposure to hyperbaric oxygen for the treatment of an acute soft tissue injury Babul, Shelina 2001-12-31

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The effects of intermittent exposure to hyperbaric oxygen for the treatment of an acute soft tissue injury by S H E L I N A BABUL B S c , T h e University of British C o l u m b i a ,  1990  A T H E S I S S U B M I T T E D IN P A R T I A L F U L F I L L M E N T O F THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in THE FACULTY OF GRADUATE STUDIES S c h o o l of H u m a n Kinetics W e accept this thesis a s conforming to the required standard  T H E UNIVERSITY O F BRITISH C O L U M B I A  October 2001 © S h e l i n a B a b u l , 2001  In presenting this thesis in partial fulfilment of the  requirements for an advanced  degree at the University of British Columbia, I agree that the Library shall make it freely available for reference and study. I further agree that permission for extensive copying of this thesis for scholarly purposes may be granted by the head of my department  or  by  his  or  her  representatives.  It  is  understood  that  copying  or  publication of this thesis for financial gain shall not be allowed without my written permission.  Department The University of British Columbia Vancouver, Canada o  DE-6 (2/88)  ABSTRACT  T h i s study e x a m i n e d the effects of intermittent e x p o s u r e to hyperbaric o x y g e n therapy ( H B O ) for the treatment of d e l a y e d onset m u s c l e s o r e n e s s ( D O M S ) . It is apparent in the literature that a great deal of controversy exists in using this form of therapy to treat tissue injuries. It w a s h y p o t h e s i z e d that subjects e x p o s e d to hyperbaric o x y g e n would recover from D O M S faster than subjects e x p o s e d to normoxic air.  Sixteen  sedentary, female university students participated in the study a n d w e r e randomly a s s i g n e d to either an experimental or control group. All subjects performed m a x i m a l voluntary eccentric contractions (30 sets of 10 repetitions/minute)  300  of their  non-dominant leg (110° - 35° of knee flexion) at a slow s p e e d (30° per s e c o n d ) on the K i n C o m D y n a m o m e t e r , to elicit m u s c l e d a m a g e a n d injury. H B O treatments c o n s i s t e d of 1 0 0 % o x y g e n for 60 minutes at 2.0 A T A while the control group received 2 1 % o x y g e n at 1.2 A T A for the s a m e amount of time. Both groups received treatment immediately after the induction of D O M S a n d e a c h d a y after for a period of 4 d a y s [day 2 post-exercise thru day 5 post-exercise].  D e p e n d e n t variables  (perceived  m u s c l e s o r e n e s s , isokinetic strength, q u a d r i c e p s c i r c u m f e r e n c e , creatine k i n a s e ( C K ) , interleukin-6  (IL-6)  and  malondialdehyde  (MDA)  were  assessed  baseline  (pre-  e x e r c i s e , day 1), 4 hours post-exercise (day 2), 2 4 hours post-exercise (day 3), 4 8 hours post-exercise (day 4) a n d 72 hours post-exercise (day 5). MRI [T2 relaxation time/STIR]) w a s a s s e s s e d b a s e l i n e (day 1), 2 4 hours post-exercise (day 3) a n d 7 2 hours post-exercise (day 5). Isokinetic strength (p<0.05) a n d p e r c e i v e d s o r e n e s s (p<0.05) indicated significance for injury to the quadricep m u s c l e for both groups but no difference w a s s e e n between groups (p=0.102, p=0.571 respectively). Q u a d r i c e p circumference w a s m e a s u r e d at the 10 a n d 2 0 c m reference point a b o v e the superior portion of the patella. T h e 1 0 c m girth m e a s u r e m e n t indicated significance (p<0.05) for muscle  injury but there  w a s no difference  between  groups  (p=0.815); 2 0  cm  m e a s u r e m e n t s h o w e d no significance (p<0.05) for both within a n d b e t w e e n groups (p=0.677). N o significance w a s evident for s e r u m C K (p<0.05), both within a n d between groups (p=0.647).  M D A a n a l y s i s revealed no significance (p<0.05) both  within a n d between groups (p=0.580). A n a l y s i s of IL-6 demonstrated no significance  (p<0.05) for both within a n d b e t w e e n groups (p=0.111). Finally, MRI a n a l y s i s for T 2 weighted imaging of the rectus femoris, v a s t u s m e d i u s a n d v a s t u s lateralis s h o w e d no statistical significance (p<0.05) between groups for treatment effects  (p=0.800,  p=0.361, a n d p=0.806 respectively). Similarly, a n a l y s i s of the S T I R i m a g e s indicated no statistical significance (p<0.05) for the s a m e three m u s c l e s (p=0.796,  p=0.580,  a n d p=0.265 respectively). T h e findings of this study s u g g e s t that hyperbaric o x y g e n therapy w a s not effective  in the treatment  indicated by the markers e v a l u a t e d .  of e x e r c i s e - i n d u c e d m u s c l e injury a s  TABLE OF CONTENTS  Abstract  ii  T a b l e of C o n t e n t s  iv  List of T a b l e s  vi  List of F i g u r e s  viii  Acknowledgments  x  Dedication  xi  Preface  xii  Chapter 1 - General Introduction 1.1 Introduction to Hyperbaric O x y g e n  1  1.2 C o m b i n i n g Hyperbaric O x y g e n ( H B O ) with D e l a y e d O n s e t  6  M u s c l e S o r e n e s s ( D O M S ) : Putting the P i e c e s of the P u z z l e Together 1.3 H y p o t h e s i s  7  1.4 A s s u m p t i o n s  7  1.5 Limitations  8  1.6 Delimitations  8  Chapter 2 - Review of the Literature 2.1 Effects of Hyperbaric O x y g e n In W o u n d Healing & T i s s u e Survival  10  2.2 O x y g e n Toxicity R e l a t e d to Hyperbaric O x y g e n T h e r a p y  13  2.3 P r e v i o u s S t u d i e s E x a m i n i n g Hyperbaric O x y g e n a n d T i s s u e Injuries  18  2.4 T h e R o l e of Hyperbaric O x y g e n in Sport & E x e r c i s e M e d i c i n e  25  2.5 A c u t e Soft T i s s u e Injury: D e l a y e d - O n s e t M u s c l e S o r e n e s s  28  2.6 T h e R o l e of M a g n e t i c R e s o n a n c e Imaging in the Detection of D O M S  46  2.7 P e r c e i v e d M u s c l e S o r e n e s s and the V i s u a l A n a l o g S c a l e ( V A S )  48  Chapter 3 - Methodology 3.1 Experimental D e s i g n  50  3.2 Subjects  52  3.3 P r o c e d u r e  52  IV  3.4 Statistical A n a l y s i s  61  3.5 Statistical P o w e r  61  Chapter 4-  Results  62  Chapter 5 -  Discussion  82  Chapter 6 -  Conclusions & Recommendations  100  References  103  Appendix  Appendix A - Consent Form  121  Appendix B - Visual Analog Scale  124  A p p e n d i x C - G r a p h i c a l Representation of R a w D a t a  125  A p p e n d i x D - S T I R Images for all subjects (days 1, 3, 5)  152  V  LIST OF TABLES  Table #  Title of Tables  Page #  T a b l e 1.  P r e s s u r e equivalents for o x y g e n c o n s u m p t i o n  4  T a b l e 2.  T h e r a p e u t i c u s e s of hyperbaric o x y g e n  4  T a b l e 3.  S e l e c t l a n d m a r k s in the history a n d d e v e l o p m e n t of hyperbaric medicine, dating back to the 1660's  5  Table 4.  C e l l u l a r a n d b i o c h e m i c a l benefits of hyperbaric o x y g e n  13  T a b l e 5.  Contraindications to H B O therapy  17  T a b l e 6.  Patient preparation prior to H B O for safety a n d fire prevention  17  T a b l e 7.  Benefits of H B O in sports injuries  28  T a b l e 8.  P r o p o s e d m e c h a n i s m of action in sports injuries  28  T a b l e 9.  M e c h a n i s m by which o x y g e n g e n e r a t e s free radicals  44  T a b l e 10.  P h y s i c a l characteristics for both groups (age, height a n d weight).  62  Table 11.  A v e r a g e ratings of perceived s o r e n e s s for the q u a d r i c e p m u s c l e of the non dominant leg before (baseline) a n d after (days 2, 3, 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  63  T a b l e 12.  A v e r a g e maximal isokinetic eccentric torque of the q u a d r i c e p m u s c l e before (baseline) a n d after (days 2, 3, 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  64  T a b l e 13.  A v e r a g e q u a d r i c e p circumference m e a s u r e d at both the 10 a n d 2 0 c m point a b o v e the superior portion of the patella. M e a s u r e m e n t s w e r e taken before (baseline) a n d after (days 2, 3, 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  66  T a b l e 14.  M e a n s e r u m creatine kinase v a l u e s before (baseline) a n d after (days 2, 3, 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  68  T a b l e 15.  M e a n M a l o n d i a l d e h y d e v a l u e s before (baseline) a n d after (days 2, 3, 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  70  T a b l e 16.  M e a n interleukin-6 v a l u e s before (baseline) a n d after (days 2, 3, 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  71  T a b l e 17.  A v e r a g e T 2 relaxation times of the rectus femoris m u s c l e t a k e n before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  73  T a b l e 18.  A v e r a g e T 2 relaxation times of the v a s t u s intermedius m u s c l e t a k e n before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  74  T a b l e 19.  A v e r a g e T 2 relaxation times of the v a s t u s lateralis m u s c l e t a k e n before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  76  Table 20.  A v e r a g e signal intensity ratio for S T I R image of the rectus femoris m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  77  Table 21.  A v e r a g e signal intensity ratio for S T I R i m a g e of the v a s t u s intermedius m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  79  Table 22.  A v e r a g e signal intensity ratio for S T I R i m a g e of the v a s t u s lateralis m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e .  80  vii  LIST OF FIGURES Figure #  Title of Figures  Page #  F i g u r e 1.  P r e s s u r e - d u r a t i o n relationship for effects of o x y g e n toxicity w h e n using hyperbaric o x y g e n therapy.  14  Figure 2.  S e q u e n c e of events a s s o c i a t e d with d e l a y e d onset m u s c l e s o r e n e s s , including m e c h a n i c a l a n d b i o c h e m i c a l processes.  33  Figure 3.  Theoretical m o d e l s h o w i n g role of cytokines a n d neutrophils during e x e r c i s e inducing d a m a g e to skeletal muscle.  41  Figure 4.  B i o c h e m i c a l m e c h a n i s m for oxygen-free radical formation resulting in skeletal m u s c l e d a m a g e a n d inflammation during e x e r c i s e .  45  Figure 5.  A v e r a g e rating of perceived s o r e n e s s for the q u a d r i c e p m u s c l e of the non-dominant leg, a c c o r d i n g to the visual a n a l o g s c a l e (range 1-10), before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e .  63  Figure 6.  A v e r a g e m a x i m a l eccentric torque for the quadricep m u s c l e , before the e x e r c i s e protocol (baseline) a n d after hyperbaric/normoxic e x p o s u r e .  65  Figure 7.  A v e r a g e quadricep circumference (10 c m location), before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic exposure .  67  Figure 8.  A v e r a g e q u a d r i c e p circumference (20 c m location), before e c c e n t r i c e x e r c i s e (baseline) and after hyperbaric/normoxic exposure.  67  F i g u r e 9.  A v e r a g e creatine kinase ( C K ) levels, before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e .  69  Figure 10.  A v e r a g e m a l o n d i a l d e h y d e ( M D A ) levels, before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e .  70  Figure 1 1 .  A v e r a g e interleukin-6 (IL-6) levels, before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e .  72  Figure 12.  M e a n T 2 relaxation times (msec) before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e (days 3, 5) for the rectus femoris m u s c l e .  73  Figure 13.  M e a n T 2 relaxation times (msec) before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e (days 3, 5) for the v a s t u s intermedius m u s c l e .  75  Figure 14.  M e a n T 2 relaxation times (msec) before eccentric e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e (days 3, 5) for the v a s t u s lateralis m u s c l e .  76  Figure 15.  M e a n Short Tip Inversion R e c o v e r y ( S T I R ) ratios before eccentric e x e r c i s e (baseline) and after hyperbaric/normoxic e x p o s u r e (days 3, 5) for the rectus femoris m u s c l e .  78  Figure 16.  M e a n Short Tip Inversion R e c o v e r y ( S T I R ) ratios before eccentric e x e r c i s e (baseline) a n d after hyperbaric/ normoxic e x p o s u r e (days 3, 5) for the v a s t u s intermedius m u s c l e .  79  Figure 17.  M e a n Short Tip Inversion R e c o v e r y ( S T I R ) ratios before e c c e n t r i c e x e r c i s e (baseline) a n d after hyperbaric/normoxic e x p o s u r e (days 3, 5) for the v a s t u s lateralis m u s c l e .  81  ix  /  ACKNOWLEDGMENTS  I w o u l d like to thank my family for their continued support a n d e n c o u r a g e m e n t during the c o u r s e of my graduate studies, my c l o s e friends for all their patience  and  understanding a n d P a u l for his motivation, words of w i s d o m a n d being s o supportive, e s p e c i a l l y w h e n I n e e d e d it the most.  I w o u l d a l s o like to thank the following people for their efforts a n d contributions to the study, b e c a u s e without their a s s i s t a n c e , this study would n e v e r h a v e b e e n c o m p l e t e d : Dr.  Dennis  Jansen,  Dr.  Urs  Steinbrecher,  Clyde  Smith,  Orca  Bay  Sports  &  Entertainment, F a h r a R a j a b a l i , T e r e s a Liu, N a b e e l a P o p a t i a , a s well a s the technical staff at St. P a u l ' s Hospital for their help. Furthermore, I w o u l d a l s o like to extend my appreciation to the B C Injury R e s e a r c h & Prevention Unit ( B C I R P U ) . P a r m i n d e r , your support, a d v i c e a n d flexibility in work allowed m e to complete the study a n d my P h D program. T h a n k Y o u !  Lastly, I would like to e x p r e s s my most s i n c e r e gratitude to my t h e s i s advisor a n d committee m e m b e r s : Dr. T e d R h o d e s , Dr. J a c k Taunton & Dr. M i k e L e p a w s k y . It w a s , without  a  doubt,  their  guidance,  patience,  understanding,  enthusiasm  and  e n c o u r a g e m e n t that h e l p e d m e to complete my graduate studies. Throughout  the  c o u r s e of my thesis, I e n c o u n t e r e d n u m e r o u s stumbling b l o c k s a n d pitfalls a n d it w a s with their continued support that I w a s able to p e r s e v e r e a n d continue the study to its entirety.  X  DEDICATION One individual who played an enormous part in my life and educational pursuits was my mother whose laughter, love and continuous support made a world of difference in my life. This is for you mom!  xi  PREFACE During this past d e c a d e , society h a s s e e n a growing n u m b e r of individuals participating in sport a n d recreational activities. H o w e v e r , the n u m b e r of injuries a s a result of this i n c r e a s e in activity h a s a l s o risen. Unfortunately, a primary cost related to injury recovery is the time-lost from participating in a n d resuming normal functional activity. T h i s h a s c o m p e l l e d health c a r e p r o f e s s i o n a l s to s e e k more efficient a n d effective therapeutic interventions in treating s u c h injuries. Hyperbaric o x y g e n therapy  may  serve to provide a m e a n s of therapy to facilitate a s p e e d i e r resumption to pre-injury activity levels a s well a s improve both the short a n d long-term p r o g n o s i s of the injury.  Although a growing interest in sports a n d e x e r c i s e medicine is b e c o m i n g evident in the literature, the u s e of hyperbaric o x y g e n a s an intervention controversial.  in this field h a s b e e n  T o date, n u m e r o u s professional athletic t e a m s , ranging from h o c k e y  (NHL), football ( N F L ) , basketball ( N B A ) a n d s o c c e r ( M L S , E u r o p e a n L e a g u e ) , utilize a n d rely on the u s e of hyperbaric o x y g e n a s adjuvant therapy for n u m e r o u s sportsrelated injuries acquired from playing competitive sports. H o w e v e r , to date, there is a paucity of r e s e a r c h on the application benefits of hyperbaric therapy a n d sports injuries.  Further r e s e a r c h n e e d s to be c o n d u c t e d s u g g e s t i n g a n d validating  the  significant effects of this treatment modality a n d further grounding its importance in sports a n d e x e r c i s e m e d i c i n e .  xii  C H A P T E R 1: G E N E R A L  INTRODUCTION  1.1 Introduction to Hyperbaric Oxygen Although the  roots of  modern  hyperbaric  o x y g e n therapy  date  back  three  centuries, it is only in the last few d e c a d e s that the scientific foundation h a s b e e n laid recognizing the benefits of inspiring 1 0 0 % o x y g e n at  greater-than-ambient  p r e s s u r e s [186]. Hyperbaric o x y g e n h a s b e e n u s e d in a variety of m e d i c a l conditions a n d clinical settings but this form of therapy to treat soft tissue injuries remains a controversial i s s u e .  Hyperbaric o x y g e n ( H B O ) therapy refers to the therapeutic procedure  where  patients inspire 1 0 0 % o x y g e n while their entire b o d i e s are subjected to p r e s s u r e s greater than ambient barometric p r e s s u r e at s e a level (the patient e n c o u n t e r s p r e s s u r e s greater than o n e a t m o s p h e r e absolute [1 A T A ] or 7 6 0 m m H g ) . O x y g e n is administered at a pressure a b o v e normal atmospheric to i n c r e a s e o x y g e n delivery to i s c h e m i c or hypoxic t i s s u e s . Barometric pressure c h a n g e s during hyperbaric o x y g e n therapy are often e x p r e s s e d in multiples of absolute (ATA) [3].  atmospheric  A c h a n g e of 1 A T A is equivalent to 14.7 p o u n d s per s q u a r e  inch (PSI), 7 6 0 m m H g or 3 3 feet of s e a w a t e r ( F S W ) [3]. Therefore, a n individual c o m p r e s s e d to 2 or 3 A T A is equivalent to being 3 3 - 6 6 feet below the surface of the o c e a n [Table 1].  G a i n i n g m e d i c a l reputation in the last 3 5 y e a r s , the efficacy of H B O therapy h a s widely b e e n a c c e p t e d a n d utilized a s the primary treatment of d e c o m p r e s s i o n s i c k n e s s (illness resulting from rapid c h a n g e s in p r e s s u r e by divers or aviators), air e m b o l i s m s (introduction of air into the circulation s y s t e m , often involuntarily by medical professionals) a n d carbon m o n o x i d e poisoning. a s a s u c c e s s f u l adjunct traumatic  ischemias,  in c r u s h injuries,  refractory  wounds,  It h a s a l s o b e e n u s e d  compartment  necrotizing  soft  syndromes, tissue  burns,  infections,  osteomyelitis, radiation tissue d a m a g e , c o m p r o m i s e d skin grafts a n d flaps a n d c a s e s of extreme blood loss [3,9,34-35] [Table 2].  1  T h e principle of hyperbaric therapy is b a s e d on the following two c o m p o n e n t s : 1) the physiologic  effects of h y p e r o x e m i a (increase in partial p r e s s u r e of o x y g e n  d i s s o l v e d in arterial blood) a n d 2) the mechanical  effects of i n c r e a s e d p r e s s u r e  [3,39]. T h r o u g h t h e s e two c o m p o n e n t s , e n o u g h o x y g e n is d i s s o l v e d in the p l a s m a to meet the metabolic n e e d s of injured t i s s u e .  T h i s is of  extreme  importance in soft tissue injuries where the environment is hypoxic. T o date, hyperbaric o x y g e n h a s b e e n r e c o g n i z e d a s a significant adjunct in the treatment of a variety of w o u n d s .  A m p l e e v i d e n c e exists that s u g g e s t s inspiration  of  o x y g e n a b o v e 2 1 % to injured body tissue c a n h a v e n u m e r o u s beneficial effects [5-13]. It h a s b e e n demonstrated that increasing the partial p r e s s u r e of o x y g e n c a n counter local tissue hypoxia (under-oxygenation of tissue) [5-9], promote peripheral vasoconstriction [5-9], d e c r e a s e blood flow to the a r e a of insult [5,811], promote healing of d a m a g e d tissue a n d prevent infection by inhibition of the growth of a n a e r o b i c m i c r o o r g a n i s m s [7,11-13].  T h e application of hyperbaric o x y g e n ( H B O ) for the treatment of sports injuries h a s recently b e e n grounded in the scientific literature a s a modality of therapy. H o w e v e r , the c o n c e p t of using c o m p r e s s e d g a s for m e d i c a l p u r p o s e s h a s a rich history, with the origins a n d development of hyperbaric m e d i c i n e being c l o s e l y tied to the history of diving medicine [9, 31-32] [Table 3]. It w a s noticed that residents living at high altitudes had w o u n d s that h e a l e d more slowly than at s e a level a s c o m p a r e d to people living in u n d e r s e a habitats at hyperbaric p r e s s u r e s (deep s e a divers) w h o s e w o u n d s h e a l e d faster at the e n h a n c e d  pressure  environment [8, 33].  Essentially, clinical hyperbaric medicine c a n be v i e w e d a s a relatively  new  application of an old established technology to help resolve s e l e c t e d m e d i c a l problems  [3].  C o n s e q u e n t l y , with this  growing  popularity,  the  number  of  hyperbaric units in the United States h a s grown from 3 4 to 2 6 0 facilities, from 1 9 7 7 - 1 9 9 8 , with over 3 5 0 single-occupant (monoplace) c h a m b e r s [36-38].  2  Currently, hyperbaric o x y g e n h a s s e v e r a l clinical indications for which it is the primary treatment modality: d e c o m p r e s s i o n s i c k n e s s , air e m b o l i s m a n d c a r b o n m o n o x i d e poisoning [14,15]. O t h e r conditions s u c h a s burns, c r u s h injuries, compartment s y n d r o m e s a n d osteomyelitis h a v e a l s o b e e n s h o w n to heal more rapidly with the application of hyperbaric o x y g e n therapy w h e n u s e d a s a n adjunct [14,16-20]. T h e r e h a s a l s o b e e n clinical investigation into the beneficial effects of hyperbaric o x y g e n a n d brain injuries [21,22]. Further r e s e a r c h is underway e x a m i n i n g its application for the treatment of multiple s c l e r o s i s [23-26] a n d its u s e is presently being investigated in children with cerebral palsy to determine whether it i m p r o v e s the quality of life for t h e s e children [27-29]. Recently, H B O h a s a l s o b e e n u s e d in treating patients with H I V / A I D S , a s it h a s b e e n demonstrated to reduce the severity of, a n d s e c o n d a r y complications arising from, opportunistic infections [194].  Competitive sports have taken a whole  n e w m e a n i n g in the  Competition is fierce a s athletes strive to be best in their field.  last d e c a d e . H o w e v e r , this  competitive nature in athletes h a s invoked a higher incidence of injuries in t h e s e players. T h e s e injuries, ranging from broken b o n e s , torn m u s c l e s , t e n d o n s a n d ligaments, m a y be a result of acute impact f o r c e s in contact sports or the everyday rigors of training a n d conditioning [30]. T h i s is where the field of sports a n d e x e r c i s e medicine plays a crucial role in the rehabilitation of t h e s e athletes of all levels. P h y s i c i a n s are frequently c h a l l e n g e d by athletes, c o a c h e s a n d trainers to provide n e w treatments to facilitate a s p e e d i e r recovery [30]. O n e s u c h treatment is the u s e of hyperbaric o x y g e n to a c c e l e r a t e the recovery p r o c e s s a n d allow the injured athlete to return to competition faster than the normal c o u r s e of rehabilitation.  j  V  3  T a b l e 1: P r e s s u r e E q u i v a l e n t s for O x y g e n C o m p r e s s i o n  ATA  mmHg  FSW  (atmosphere absolute)  (feet of sea water)  1  760  0  2  1520  33  3  2280  66  6  4560  165  T a b l e 2: Therapeutic U s e s of Hyperbaric O x y g e n *  Strong Scientific Evidence Maltreatment Decompression sickness Arterial g a s e m b o l i s m v S e v e r e c a r b o n m o n o x i d e poisoning a n d s m o k e inhalation Adjunctive treatment Prevention and treatment of osteoradionecrosis Improved skin graft a n d flap healing Clostridial m y o n e c r o s i s  Suggestive Scientific Evidence Adjunctive treatment Refractive osteomyelitis Radiation induced injury A c u t e traumatic i s c h e m i c injury P r o l o n g e d failure of w o u n d healing E x c e p t i o n a l a n e m i a from blood l o s s  * adapted from Leach RM et al  40  T a b l e 3: S e l e c t landmarks in the history a n d development of hyperbaric m e d i c i n e , dating back to the 1 6 0 0 ' s  Date  Key Landmark  1662  J . H e n s h a w u s e d c o m p r e s s e d air for the treatment of a variety of diseases D i s c o v e r y of o x y g e n by J . Priestley T. B e d d o e s a n d J . Watt wrote first book on m e d i c a l application of o x y g e n C . G . P r a v a z of F r a n c e constructed largest hyperbaric c h a m b e r of that time to treat a variety of ailments ^ First hyperbaric c h a m b e r on North A m e r i c a n continent in O s h a w a , Canada J . S . H a l d a n e s h o w e d that a m o u s e p l a c e d in a jar containing o x y g e n at 2.0 A T A failed to d e v e l o p C O poisoning O . J . C u n n i n g h a m built a hyperbaric c h a m b e r in L a w r e n c e , K a n s a s u s e d to treat a variety of ailments O . J . C u n n i n g h a m builds the largest c h a m b e r in the world in Cleveland A . R . B e n k e a n d L.A. S h a w first u s e d H B O for treatment of decompression sickness 1. B o e r e m a , father of m o d e r n hyperbaric m e d i c i n e , performed ,„ cardiac surgery in a hyperbaric c h a m b e r First International C o n g r e s s on Hyperbaric M e d i c i n e in Amsterdam U n d e r s e a M e d i c a l S o c i e t y f o u n d e d in the U S A . N o w known a s the U n d e r s e a a n d Hyperbaric M e d i c a l Society. Extensive e x p a n s i o n of hyperbaric facilities in J a p a n a n d U S S R Formation of the A m e r i c a n C o l l e g e of Hyperbaric M e d i c i n e K.K. J a i n demonstrated H B O integration with p h y s i c a l therapy Formation of the International S o c i e t y of Hyperbaric M e d i c i n e 1  1775 1796 1837 1860 1895 1921 1928 1937 1956 1963 1967 1970s 1983 1987 1988  5  1.2 Combining Hyperbaric Oxygen with Delayed Onset Muscle Soreness: Putting the Pieces of the Puzzle Together  T h e d e l a y e d - o n s e t m u s c l e s o r e n e s s ( D O M S ) m o d e l will elicit m u s c l e injury a n d tissue inflammation in h u m a n s a n d s e r v e a s a n excellent m o d e l to a s s e s s the efficacy of using this form of treatment in the rehabilitation of exercise-related injuries.  D O M S , c h a r a c t e r i z e d by high intensity eccentric contractions, clinically presents a s i n c r e a s e d stiffness, a d e c r e a s e d range of motion, t e n d e r n e s s , decline in force, swelling, electrically silent m u s c l e shortening a n d a release of m u s c l e e n z y m e s in the blood  [119, 182].  R o d e n b u r g et al [224] h a s stated that " D O M S ultimately  arises from a ' s e q u e n c e of events' occurring after eccentric e x e r c i s e , including myofibrillar  disruption, i n c r e a s e d permeability  of the s a r c o l e m m a to  muscle  proteins, free radical release a n d inflammatory p r o c e s s e s , with the latter possibly leading to D O M S . T h i s ' s e q u e n c e of e v e n t s ' is initiated by m e c h a n i c a l stress on the m u s c l e fibres, metabolic overload or a combination of both".  D u e to the lack of scientific e v i d e n c e supporting the efficacy of using hyperbaric o x y g e n therapy in sport a n d exercise-related injuries, the effects of using this treatment modality w a s looked at in a n acute soft tissue inflammatory condition, namely D O M S .  T h i s premise is b a s e d on the m e c h a n i s m of action a s s o c i a t e d  with H B O , a n d the m e c h a n i c a l a n d b i o c h e m i c a l p r o c e s s e s that o c c u r during a bout of D O M S . M u s c l e d a m a g e , m u s c l e s o r e n e s s a n d loss of m u s c l e function will be  determined  by  assessing  isokinetic  strength  decrements,  associated  p e r c e i v e d m u s c l e s o r e n e s s , elevation in blood e n z y m e levels of creatine k i n a s e , interleukin 6 a n d m a l o n d i a l d e h y d e , a s well a s magnetic r e s o n a n c e imaging of e d e m a t o u s tissue.  E a c h piece of the " p u z z l e " c a n be put together to determine whether, in fact, hyperbaric o x y g e n d o e s have an important  role in the future of sports a n d  6  e x e r c i s e m e d i c i n e . If it d o e s , further r e s e a r c h c a n then be c o n d u c t e d on a wide array of other injuries acquired in competitive a n d recreational activities a s well a s other clinical disorders  1.3 Hypothesis T h e p u r p o s e of this study w a s to determine whether intermittent e x p o s u r e to hyperbaric o x y g e n in the treatment group, c o m p a r e d to the control group, would i n c r e a s e the rate of recovery from D O M S , thus:  1.  R e d u c i n g p e r c e i v e d m u s c l e s o r e n e s s over the five-day testing period.  2.  Significantly improving  eccentric m u s c l e strength  during the  recovery  of  DOMS. 3.  D e c r e a s i n g e l e v a t e d creatine k i n a s e ( C K ) levels a s s o c i a t e d with skeletal m u s c l e d a m a g e over the five d a y s , thereby bringing C K levels c l o s e to pree x e r c i s e levels by day 5.  4.  D e c r e a s i n g e l e v a t e d interleukin-6 (IL-6) levels a s s o c i a t e d with e x e r c i s e induced m u s c l e injury over the five d a y s , thereby bringing IL-6 levels c l o s e to baseline by d a y 5.  5.  D e c r e a s i n g elevated m a l o n d i a l d e h y d e  ( M D A ) levels a s s o c i a t e d with  lipid  peroxidation a s a result of m u s c l e injury over the five d a y s , thereby bringing M D A levels c l o s e to pre-exercise levels by day 5. 6.  R e d u c i n g e d e m a in the quadricep m u s c l e of the non-dominant leg over the five-day testing period, a s e v i d e n c e d by magnetic r e s o n a n c e imaging (MRI).  1.4  Assumptions  T h e following a s s u m p t i o n s were m a d e w h e n designing the study: 1.  All subjects will r e s p o n d honestly, to the best of their knowledge, regarding the amount of activity that they perform on a weekly b a s i s a s well a s their involvement in competitive sporting activities.  2.  All subjects will report accurate pain s c o r e s , to the best of their ability, w h e n filling out the visual a n a l o g s c a l e .  7  3. Performing 3 0 0 eccentric contractions will create m u s c l e injury a n d tissue inflammation ( D O M S ) in the quadricep m u s c l e of the non-dominant leg.  1.5 Limitations T h e study, at the present time, w a s limited by the following: 1. T h e severity of s o r e n e s s , a s p e r c e i v e d by individual subjects, m a y be prone to inter-subject variability. 2. Subject recruitment: It w a s extremely difficult to recruit subjects for the study d u e to the time commitment required over the 5 d a y s of treatment a n d the travel d i s t a n c e s required by the subjects (Allan M c G a v i n S p o r t s M e d i c i n e Clinic, B u c h a n a n E x e r c i s e S c i e n c e Laboratory, U B C , St. P a u l ' s Hospital a n d MRI V a n c o u v e r ) .  1.6 Delimitations 1. S a m p l e s e l e c t i o n ; sedentary or relatively s e d e n t a r y individuals, b e t w e e n the a g e s of 18-40 y e a r s (N=16; n=8 per group), w e r e recruited to m a x i m i z e the extent of D O M S s i n c e their quadricep m u s c l e will not be a d a p t e d to eccentric loading. 2. T h e Kinetic C o m m u n i c a t o r ( K i n C o m ) D y n a m o m e t e r w a s u s e d to test the subject's m u s c l e strength through a specific range of motion while performing isolated knee e x t e n s i o n s . 3. T h e D O M S protocol (eccentric e x e r c i s e protocol) allowed the investigator to control the insult of the injury. 4. T h e c i r c u m f e r e n c e m e a s u r e m e n t of the quadricep m u s c l e e v a l u a t e d the girth of the thigh, including s k i n , fat, m u s c l e , b o n e a n d p o s s i b l e e d e m a occurring within the musculature. 5. IL-6 m e a s u r e m e n t s taken at baseline a n d repeated throughout the fiveday testing period demonstrated the cytokine r e s p o n s e during inflammation a n d D O M S .  8  M a l o n d i a l d e h y d e m e a s u r e m e n t s taken at b a s e l i n e a n d repeated throughout recovery w e r e indicative of lipid peroxidation during m u s c l e injury.  CHAPTER 2: REVIEW OF THE LITERATURE 2.1 Effects of Hyperbaric Oxygen in Wound Healing and Tissue Survival During a soft tissue injury, a disruption of cells a n d blood v e s s e l s o c c u r within the t i s s u e , resulting in h y p o x i a . T h i s is followed by a s u b s e q u e n t aggregation of platelets a n d c o l l a g e n to the extracellular  fluid  and  a r e a of  injury.  v a s c u l a r dilation  In addition, a n  o c c u r s , followed  by  i n c r e a s e in an  influx  of  neutrophils, m a c r o p h a g e s , fibroblasts, smooth m u s c l e cells a n d endothelial cells to c l e a n s e a n d reconstruct the insult.  Lactate, h y p o x i a a n d the production of  cytokines further c a u s e s growth stimulation, which l e a d s to a n g i o g e n e s i s a n d the production of c o l l a g e n .  T h e w o u n d is essentially c o n s i d e r e d h e a l e d after the  o c c u r r e n c e of this c a s c a d e of e v e n t s [41-42, 50].  T i s s u e d a m a g e c a n lead to e d e m a , complications with blood flow a n d eventual tissue death, including i s c h e m i a  [8, 41-42].  T h e i n c r e a s e in extracellular fluid  a n d v a s c u l a r dilation impairs o x y g e n delivery from capillaries to cells b e c a u s e of a n i n c r e a s e in the diffusion distance [9]. Resulting d e c r e a s e s in o x y g e n t e n s i o n s (e.g. below 30 m m H g ) m a k e the cells more susceptible to infection  [7, 43] a n d  the efficacy of leukocytes in reducing invading o r g a n i s m s b e c o m e s defective [12, 44]. Furthermore, host repair p r o c e s s e s are also impaired during this time [7].  O x y g e n , therefore, plays a critical role in the w o u n d healing p r o c e s s . It s e r v e s a s a catalyst a n d energy s o u r c e for m a i n t e n a n c e , metabolism a n d repair In w o u n d  healing, o x y g e n s e r v e s to  [45, 46].  provide the additional energy s o u r c e  n e c e s s a r y for the reparative p r o c e s s . Early in the repair of w o u n d s , fibroblasts begin to migrate, divide a n d p r o d u c e c o l l a g e n , which is an essential matrix for w o u n d healing. O x y g e n must a l s o be present in sufficient quantities for fibroblast proliferation a n d c o l l a g e n production to occur. A d e q u a t e a m o u n t s of proline a n d lysine (two amino a c i d s incorporated by oxygen) must be hydroxylated with o x y g e n for collagen to be s y n t h e s i z e d by fibroblasts [47].  O x y g e n must a l s o be  present in a d e q u a t e a m o u n t s during the repair p r o c e s s to provide energy for protein synthesis.  It h a s b e e n demonstrated that raising o x y g e n tension in  10  tissues increases the ratio of R N A / D N A [48].  Pal et al [49] have stated that an  increase of 150% above the normal physiologic range for oxygen (PO =40 2  mmHg) increases the rate of collagen production by seven times.  An adequate delivery of oxygen via an extensive capillary network is essential since the diffusion of oxygen through tissues is limited  [46].  Disruption of the  capillaries from trauma produces hypoxia and the release of hormonal mediators. Macrophages release an angiogenesis factor, which is a potent stimulus for endothelial cell activity [50].  Polymorphonuclear cells (PMN) which locate,  identify, phagocytose, kill and digest microorganisms, require oxygen to kill organisms by producing superoxide, hydrogen peroxide, singlet oxygen and [  other products via the respiratory burst  [51].  Detoxifying free radicals by  superoxide dismutase, catalase and glutathione protect the P M N s .  Therefore,  the degree of P M N cell function in killing of bacteria is directly dependent on oxygen tensions [44, 52].  Hyperbaric Oxygen has two components in wound healing and tissue survival: acting mechanically, due to its pressure component and physiologically,  due to its  oxygen component [39]. The oxygen content is determined by a combination of oxygen that is bound to hemoglobin plus the amount of oxygen that is dissolved in the plasma.  [Total 0  2  = h e m o g l o b i n b o u n d o x y g e n + o x y g e n d i s s o l v e d in p l a s m a ]  The application of hyperbaric oxygen increases the amount of available oxygen to the hypoxic area of injury, increasing oxygen tensions that make host repair processes functional [8, 9]. Increased availability causes a shift in the oxygen cascade (a gradient from the partial pressure of oxygen in the ambient air to that available immediately to the tissues on a cellular level).  This effect of  hyperoxygenation is based on a combination of two laws, namely Henry and Dalton's Laws. Henry's Law states that as the P 0 increases during compression, 2  11  the amount of o x y g e n d i s s o l v e d directly into the p l a s m a i n c r e a s e s [3, 53]. i n c r e a s i n g the partial p r e s s u r e of o x y g e n in the air, a significant amount o x y g e n b e c o m e s d i s s o l v e d in the blood p l a s m a .  By of  Furthermore, Dalton's L a w  states that air is a mixture of g a s e s a n d that the total p r e s s u r e exerted is the s u m of the partial p r e s s u r e s of e a c h of the g a s e s in the mixture [53]. T h e s e two laws c o m b i n e d p r o d u c e the effect of h y p e r o x e m i a , which allows e n o u g h o x y g e n to d i s s o l v e in the p l a s m a to meet metabolic n e e d s . At s e a level in room air, there is 0.32 ml of o x y g e n d i s s o l v e d in e a c h 100 ml of whole blood (0.32 vol.%).  When  breathing 1 0 0 % o x y g e n , e a c h additional a t m o s p h e r e of p r e s s u r e p r o d u c e s a n additional 2.3 v o l . % o x y g e n d i s s o l v e d in p l a s m a . At 2.0 A T A , the blood o x y g e n content  i n c r e a s e s 2 . 3 % while p l a s m a a n d tissue o x y g e n t e n s i o n s i n c r e a s e  tenfold  (1000%)  [8,9,11,22,54].  Consequently,  sufficient  oxygen  becomes  physically d i s s o l v e d in the p l a s m a to k e e p t i s s u e s alive despite the inability of h e m o g l o b i n - b o u n d o x y g e n to reach the insulted a r e a [17, 55].  At 3.0 A T A , the  partial p r e s s u r e of o x y g e n in the blood c a n i n c r e a s e to a s m u c h a s 2 2 0 0 m m H g ( p l a s m a contains 6.8 v o l . % oxygen). T h i s elevated o x y g e n p r e s s u r e i n c r e a s e s the o x y g e n diffusion gradient a n d improves o x y g e n delivery to relatively i s c h e m i c t i s s u e s [53-56]. At this p r e s s u r e , e n o u g h o x y g e n c a n be d i s s o l v e d in the p l a s m a to sustain life temporarily without a n y red blood cells [57].  S e c o n d a r y effects of hyperbaric o x y g e n therapy include vasoconstriction, g a s volume  reduction, inhibition  of a n a e r o b i c o r g a n i s m s a n d n e o v a s c u l a r i z a t i o n .  V a s o c o n s t r i c t i o n c a u s e s a d e c r e a s e in e d e m a at the w o u n d site leading to decreased  tissue  perfusion  without  sacrificing  oxygenation  [53].  Sufficient  a m o u n t s of o x y g e n are d i s s o l v e d in the blood p l a s m a to adequately c o m p e n s a t e for the d e c r e a s e in blood flow to the a r e a of injury [9, 58]. T h i s reduction in blood flow p r o d u c e s a s u b s e q u e n t 2 0 % reduction in post-traumatic v a s o g e n i c e d e m a [59]. Hyperbaric o x y g e n therapy a l s o c a u s e s a reduction in g a s v o l u m e reducing the s i z e of the g a s b u b b l e s .  by  T h i s physiological effect is b a s e d on  B o y l e ' s law stating, "the v o l u m e of a g a s is inversely proportional to its p r e s s u r e at a constant temperature".  Therefore, a s the p r e s s u r e i n c r e a s e s , the v o l u m e of  12  a g a s d e c r e a s e s , reducing the size of g a s b u b b l e s that i m p e d e circulation [53]. B e c a u s e of this effect, H B O is u s e d a s the primary treatment in air e m b o l i s m s a n d d e c o m p r e s s i o n s i c k n e s s [60].  A l t h o u g h aerobic o r g a n i s m s continue to thrive w h e n given i n c r e a s e d a m o u n t s of o x y g e n , their growth m a y b e inhibited w h e n the partial p r e s s u r e of o x y g e n e x c e e d s 1.3 A T A [57, 61]. H o w e v e r , inhibition of aerobic o r g a n i s m s is best a c h i e v e d w h e n they a r e superficial [62] (e.g. a superficial ulcer or burn). Hyperbaric o x y g e n therapy u s e d a s a n adjunct h a s a l s o b e e n d e m o n s t r a t e d to b e beneficial in treating a n a e r o b i c infections s u c h a s g a s g a n g r e n e a n d inhibition of a l p h a toxin production [62] [Table 4].  T a b l e 4 : C e l l u l a r a n d B i o c h e m i c a l Benefits of Hyperbaric O x y g e n * • • • • • •  P r o m o t e s a n g i o g e n e s i s a n d w o u n d healing Kills certain a n a e r o b e s P r e v e n t s growth of s p e c i e s s u c h a s P s e u d o m o n a s P r e v e n t s production of clostridial a l p h a toxin R e s t o r e s neutrophil mediated bacterial killing in previously hypoxic t i s s u e s R e d u c e s leukocyte a d h e s i o n in reperfusion injury, preventing r e l e a s e of p r o t e a s e s a n d free radicals which c a u s e vasoconstriction a n d cellular damage  *adapted from Leach RM  2.2 Oxygen Toxicity Related to Hyperbaric Oxygen Therapy O x y g e n in large a m o u n t s , like most therapeutic modalities, c a n b e toxic a n d life threatening.  H o w e v e r , it h a s widely b e e n reported that toxicity is related to both  the duration of e x p o s u r e a n d p r e s s u r e [63] (Figure 1). O x y g e n toxicity affects both central a n d pulmonary n e r v o u s s y s t e m s . O x y g e n toxicity involving the central nervous s y s t e m is termed the Paul Bet effect a n d results in s e i z u r e s [205]. O x y g e n toxicity affecting the pulmonary s y s t e m is termed the Lorrain-Smith effect, a n d c a u s e s e d e m a in the lungs a n d s u b s e q u e n t alveolar c o l l a p s e [206].  13  Figure 1: P r e s s u r e - d u r a t i o n relationship for effects of o x y g e n toxicity w h e n using hyperbaric o x y g e n therapy [63]  0.5 ATM 3  6  12  15  18  21  24  Exposure Duration (hours)  Central n e r v o u s s y s t e m a s s o c i a t e d grand mal s e i z u r e s have b e e n d o c u m e n t e d at a p r e s s u r e of 3.0 A T A , [62, 63] while pulmonary e d e m a h a s b e e n detected at 2.0 A T A [54,  64-66].  H o w e v e r , both toxic effects  e x p o s u r e s of three or more hours  [9, 6 5 , 66, 68].  occurred at  prolonged  D a v i s [67] estimated the  incidence of o x y g e n s e i z u r e s to be one in 11,000 treatments.  Thorn  [207]  reported that the likelihood of central nervous s y s t e m agitation is approximately 0 . 0 0 0 9 % . F a c t o r s contributing to a seizure are fever, e x e r c i s e , a p p r e h e n s i o n a n d C O 2 buildup [57]. T y p i c a l warning signs m a y include tunnel vision, s h o r t n e s s of breath, tinnitus, n a u s e a a n d extreme a p p r e h e n s i o n . If a seizure o c c u r s during treatment, the patient s h o u l d be r e m o v e d from the c h a m b e r but not until the seizure h a s s t o p p e d . T o d e c o m p r e s s the patient during the tonic p h a s e of the s e i z u r e c a n put the patient at risk of air or o x y g e n e m b o l i s m [57]. S e i z u r e s , if promptly treated, have no permanent s e q u e l a e .  P n e u m o t h o r a x c a n o c c u r under hyperbaric conditions due to p r e d i s p o s i n g lung pathology s u c h a s g a s trapping in a localized portion of the lung [57]. T h e patient m a y e x p e r i e n c e respiratory distress, s u d d e n stabbing chest pain, a shift of the t r a c h e a toward the unaffected side, lack of chest m o v e m e n t on the affected side, d e c r e a s e d breath s o u n d s on the affected side a n d i n c r e a s e d tympany When  a multiplace  c h a m b e r is u s e d ,  treatment  of  pneumothorax  [57].  can  14  be  m a n a g e d by inserting a n e e d l e or chest tube into the affected side. F o r treatment in a m o n o p l a c e c h a m b e r , the patient must be d e c o m p r e s s e d before treatment. T h i s m a y c a u s e the p n e u m o t h o r a x to double or triple in v o l u m e but respiratory distress m a y s u b s i d e w h e n r e c o m p r e s s i o n o c c u r s .  P u l m o n a r y o x y g e n toxicity m a y be a consideration in patients maintained on an inspired o x y g e n fraction  of  greater than  4 0 % between  hyperbaric  oxygen  s e s s i o n s [208]. In this instance, the clinician on h a n d must be a w a r e of the hyperbaric o x y g e n treatment to monitor for toxicity a n d intervene if the n e e d arises.  T h e u s e s of aspirin, insulin, steroids, epinephrine a n d norepinephrine h a v e all b e e n noted to i n c r e a s e the onset of o x y g e n toxicity  [9, 34]. Concurrent therapy  with doxorubicin ( A d r i a m y c i n ) , cis-platinum a n d disulfiram ( A n t a b u s e ) are a l s o R  incompatible with H B O [3]. p r o d u c e d a high  mortality  R  W h e n c o m b i n e d with H B O therapy, doxorubicin h a s rate  in a n i m a l s . Furthermore  cis-platinum  used  concurrently with H B O therapy d e c r e a s e s the strength of healing w o u n d s a n d disulfiram blocks production of s u p e r o x i d e d i s m u t a s e , the body's major protection against o x y g e n toxicity [3]. C o n c e r n s h a v e a l s o b e e n e x p r e s s e d a s to whether physical e x e r c i s e p r e d i s p o s e s patients to  o x y g e n toxicity  [9]  but  a  study  c o n d u c t e d by S t e v e n s et al [66] c o n c l u d e d that there were no a d v e r s e effects of o x y g e n toxicity with e x e r c i s e .  C o n v e r s e l y , antioxidants s u c h a s vitamin C ,  vitamin E a n d m e x a m i n e , lithium, a n d m a g n e s i u m h a v e b e e n u s e d to prevent or delay the onset of toxicity [202-204].  T h e best way, however, to  prevent  prolonged o x y g e n toxicity is to take periodic b r e a k s , inspiring normal air ( 2 1 % oxygen)  [9].  A five y e a r prospective study e x p o s i n g 12 4 6 8 patients to 1 0 0 %  o x y g e n at 2.0, 2.2 a n d 2.4 A T A s h o w e d no oxygen-related complications or s e i z u r e s at 2.0 A T A  [68].  Mild aural barotrauma w a s the only complication  o b s e r v e d in this study. B a r o t r a u m a is defined a s any injury to structures s u c h a s the e a r due to differences between a t m o s p h e r i c a n d intratympanic p r e s s u r e s (commonly referred to a s " s q u e e z e " ) [57].  D u e to p r e s s u r e c h a n g e s a s s o c i a t e d i  15  with hyperbaric treatment, the patient must be taught to e q u a l i z e the pressure in the e a r by swallowing, y a w n i n g or using the v a l s a l v a m a n o e u v e r . T h e primary s y m p t o m of " s q u e e z e " is pain in the e a r while the c h a m b e r is being c o m p r e s s e d [57]. B a r o t r a u m a c a n a l s o o c c u r in the s i n u s cavity, primarily the frontal s i n u s . D e c o n g e s t a n t s are usually p r e s c r i b e d w h e n this o c c u r s . In a s e r i e s of studies c o n d u c t e d , D a v i e s reported that o n e in 2 7 0 c a s e s h a d barotrauma significant e n o u g h to interrupt treatment [67].  Others h a v e stated complications s u c h a s  n a u s e a , tooth a n d s i n u s pain a n d blurred vision [9].  Therefore, it c a n be  c o n c l u d e d that no toxic effects are s e e n within the current therapeutic range of 2.0 A T A at a 6 0 - 9 0 minute e x p o s u r e time, a c c o m p a n i e d by frequent air b r e a k s during treatment.  P r o p e r s c r e e n i n g is e s s e n t i a l before hyperbaric treatment c a n be administered. Contraindications for H B O therapy include upper respiratory tract infections, diabetes, p r e g n a n c y , confinement s y n d r o m e , pneumothorax, sinusitis a n d fever [Table 5].  Fire in the 1 0 0 % 0  2  environment of a mono/multiplace c h a m b e r is, although rare,  a c o n c e r n that s h o u l d be a d d r e s s e d . All efforts s h o u l d be maintained to minimize the risk, including the e x c l u s i o n of items that c o u l d be a s s o c i a t e d with heat or flame. T h e s e include velcro, glycerin (including hair products a n d c o s m e t i c s ) , perfumes, lotions a n d non-cotton fabrics (should be 1 0 0 % cotton to reduce static) . [Table 6].  O x y g e n toxicity is a s e r i o u s c o n s e q u e n c e of treatment with hyperbaric o x y g e n . H o w e v e r , this c a n be controlled with proper s c r e e n i n g of the patient prior to administration of treatment a n d appropriate control over length of e x p o s u r e a n d p r e s s u r e levels. Ideally, it c a n be c o n c l u d e d that treatments for 6 0 - 9 0 minute durations are within a r e a s o n a b l e level'to avoid complications that m a y arise. Furthermore, treatment s h o u l d be administered at" 2.0-2.5 A T A , a c c o m p a n i e d by frequent air b r e a k s .  16  T a b l e 5: Contraindications to H B O T h e r a p y * • • • • • • • • • • •  Pneumothorax Uncontrolled high fevers S e v e r e chronic obstructive pulmonary d i s e a s e Optic neuritis A c u t e viral infection Congenital spherocytosis U p p e r respiratory tract infections Pregnancy Psychiatric p r o b l e m s Prior thoracic or e a r surgery A c u t e seizure disorders  *adapted from Foster J  T a b l e 6: Patient Preparation Prior to H B O for Safety a n d Fire Prevention  • •  • •  T h e patient s h o u l d s h o w e r or bathe daily a n d hair s h o u l d be / s h a m p o o e d if oily before H B O B o d y oils, lotions, talc, petroleum products, m a k e - u p , c o l o g n e , perfume, deodorants, nail polish, hair oils a n d s p r a y are not to be u s e d before treatment. O n l y all cotton clothing a n d linens are allowed in the c h a m b e r to prevent development of static electricity. W i g s or hairpieces, hearing aids, jewelry a n d contact l e n s e s are not to be worn in the c h a m b e r .  *adapted from Curtis et al  17  2.3 Previous Studies Examining HBO and Tissue Injuries Burns  A vast amount of r e s e a r c h exists on the effects of H B O on burns, e s p e c i a l l y thermal burns.  First d o c u m e n t e d in the 1970's, patients e x p o s e d to H B O found  that burns dried sooner, h a d fewer infections a n d h e a l e d m o r e quickly  [18, 70].  H e a l i n g time w a s s h o w n to d e c r e a s e by 3 0 % in more recent controlled studies [10, 71-73]. rates  of  Other authors e x a m i n i n g s e c o n d d e g r e e burns report i n c r e a s e d  epithelialization,  decreased  fluid  microcirculation, d e c r e a s e d c o n v e r s i o n s from  requirements,  preservation  partial to full t h i c k n e s s  of  injury,  reduction in e d e m a a n d inflammatory r e s p o n s e a n d d e c r e a s e s in grafting a n d surgical p r o c e d u r e s , hospital time a n d relevant a s s o c i a t e d c o s t s [3, 5, 72-74].  In  a m o d e l s c a l d burn, N y l a n d e r et al [76] demonstrated a d e c r e a s e in global e d e m a following a n e a r burn. Stewart et al [75] e x a m i n e d A T P , p h o s p h o c r e a t i n e a n d c o l l a g e n s y n t h e s i s in burn w o u n d s of rats.  T h e y c o n c l u d e d that a s an  adjunct, H B O could preserve or e n h a n c e energy rich p h o s p h a t e c o m p o u n d s a n d c o l l a g e n s y n t h e s i s in model burn w o u n d s .  Although large a m o u n t s of r e s e a r c h exist demonstrating the healing potential a n d beneficial  effects  on  epithelialization,  some  d i s c r e p a n c y d o e s exist  literature with respect to the efficacy of H B O therapy a s a n adjunct O t h e r controlled, r a n d o m i z e d h u m a n studies s h o w no effect  in  the  treatment.  in the  healing  p r o c e s s [43, 76]. T h i s d i s c r e p a n c y m a y be attributable to the time lag from injury to the initiation of treatment, possibly 2 4 - 4 8 hours post-injury.  Infections  T h e role of o x y g e n in fighting infections h a s also b e e n e s t a b l i s h e d . O x y g e n acts a s a potent antibiotic, improving the ability of s p e c i a l s c a v e n g e r white blood cells (phagocytes) to rid the body of bacteria a n d other foreign proteins.  Hyperbaric  o x y g e n h a s a distinct antimicrobial effect that is equal to or better than that of n u m e r o u s antibiotics [29]. T h i s form of treatment m a y a l s o e n h a n c e the actions of certain antibiotics a n d thereby i n c r e a s e their effectiveness in o v e r c o m i n g  18  infections [29].  B e c a u s e of its i m m u n e s y s t e m - e n h a n c i n g effects, hyperbaric  o x y g e n therapy  helps fight all m i c r o o r g a n i s m s , both a n a e r o b i c a n d aerobic  o r g a n i s m s . T h e literature h a s reported over 4 , 0 0 0 c a s e s of g a s g a n g r e n e treated with hyperbaric o x y g e n therapy. O n e study reported a survival rate of 8 8 . 3 % a m o n g 2 4 8 patients w h o received treatment [209]. A n o t h e r study of 139 patients reported a survival rate of 70 percent, with 8 0 % of the survivors a b l e to avoid amputations [210]. Ellis a n d M a n d a l a n a l y z e d 58 patients w h o failed to r e s p o n d to  conventional  treatment  of  antibiotics  and  surgery.  After  treatment  with  hyperbaric o x y g e n , there w a s a survival rate of 8 4 % [211].  Wound  Healing  A w o u n d is any disruption in the b o d y ' s t i s s u e s . It is often a s s o c i a t e d with the loss of skin (and underlying tissue), m u s c l e or bone. M a n y w o u n d s r e s p o n d to conventional  m e d i c i n e while  others  do  not.  Primarily,  a wound  lacks  the  n e c e s s a r y o x y g e n required for healing to take place. T h i s m a y be d u e to a blood clot, which interferes with the circulation. A s a result, the o x y g e n supply to the a r e a of insult is diminished a n d toxic materials a c c u m u l a t e s i n c e w a s t e products cannot be r e m o v e d . T h i s starts a c y c l e of d a m a g e to affected t i s s u e s . W o u n d s are a l s o prone to infection s i n c e the lack of o x y g e n in the t i s s u e s c a n reduce the injured p e r s o n ' s d e f e n s e by d e c r e a s i n g the activity of infection-fighting  white  blood cells [12]. Underoxygenation c a n a l s o deactivate the cells that produce granulation tissue a n d interfere with c o l l a g e n production [194]. Therefore, it c a n be c o n c l u d e d that a lack of o x y g e n in the w o u n d e d tissue c a n interfere with the entire w o u n d healing p r o c e s s .  In the treatment of o p e n w o u n d s , H B O plays a vital role in c o l l a g e n synthesis, hydroxylation of the collagen m o l e c u l e a n d intracellular A T P production, thus promoting granulation  tissue repair a n d growth tissue  formation,  [7, 8]. T o s u m m a r i z e , H B O  revascularization,  epithelialization,  promotes enhanced  fibroblastic activity a n d leukocyte killing, fibroblast migration, a n g i o g e n e s i s a s well a s capillary budding [3, 3 6 , 78-80].  A m p l e clinical investigations  have  19  d e m o n s t r a t e d that in hypoxic w o u n d s (PO2 = 5-20 m m H g ) , tissue healing o c c u r s with sufficient tissue oxygenation, which is e n h a n c e d by H B O [81].  In a study of  patients with chronic diabetic foot lesions, Doctor et al [184] o b s e r v e d a better control of infection a n d l e s s n e e d for amputation  in the group treated with  conventional m a n a g e m e n t a n d s e s s i o n s of H B O .  Bone  Healing  and  Hyperbaric  Oxygen  Therapy  B o n e is c o m p o s e d of three layers: a s p o n g y inner layer, a rigid middle layer a n d a tough outer layer. W h e n bone is injured, s p e c i a l cells called o s t e o c l a s t s work to repair the d a m a g e . T h e s e cells carve paths through the bone t i s s u e a r o u n d the brake a n d c a u s e d e a d b o n e to be r e a b s o r b e d by the body. O s t e o b l a s t s , another group of cells, then create new b o n e [29]. T h e s e o s t e o c l a s t s d e p e n d a n d utilize o x y g e n for proper function. Therefore, hyperbaric o x y g e n m a y facilitate healing by stimulating both o s t e o c l a s t s a n d osteoblasts [7, 8].  bone  It m a y a l s o  stimulate the production of new blood v e s s e l s , s o that the growing b o n e r e c e i v e s a s t e a d y supply of nutrients, including o x y g e n . T h i s b l o o d - v e s s e l network helps support the function of the o s t e o c l a s t s a n d brings infection-fighting white blood cells to the a r e a of injury [29].  Osteomyelitis  Osteomyelitis is a bacterial infection that usually o c c u r s on the outer layers of b o n e a s well a s the inner bone marrow. S t a p h y l o c o c c i bacteria are  primarily  implicated in this type of infection. T h e g e r m s that c a u s e this infection c a n enter the bone during a n injury or surgery. Furthermore, it m a y a l s o reach the bone directly  from  a  nearby  infection  or  indirectly  through  the  bloodstream.  Osteomyelitis m a y be either acute or chronic, with s e v e r e pain, swelling a n d r e d n e s s at the site of infection in the acute p h a s e of infection. High fever is a l s o prominent in patients with osteomyelitis. In the chronic stage, s y m p t o m s include bone pain, t e n d e r n e s s a n d local m u s c l e s p a s m .  20  C o n v e n t i o n a l treatment for osteomyelitis includes antibiotics a n d s e v e r a l w e e k s of b e d rest. Surgery m a y be required to take out d e a d bone a n d soft t i s s u e , fill h o l e s a n d impjant artificial d e v i c e s d e s i g n e d to k e e p the d i s e a s e d b o n e s a n d joints from moving [29]. It is imperative to treat the bone infection promptly a n d vigorously in order to prevent it from s p r e a d i n g to other parts of the body.  Hyperbaric o x y g e n h a s three main functions in treating osteomyelitis. First it helps strengthen the b o n e cells (osteoclasts) that reabsorb d e a d b o n e , removing bony debris more effectively. S e c o n d , it e n h a n c e s the function of the i m m u n e s y s t e m ' s white blood cells, s i n c e they d e p e n d on o x y g e n . T h i s is e s p e c i a l l y effective w h e n u s e d with antibiotics [29]. Lastly, hyperbaric o x y g e n h e l p s the body to create new blood v e s s e l s . T h r o u g h t h e s e m e c h a n i s m s , hyperbaric o x y g e n e n a b l e s the body to get rid of the d i s e a s e d bone a n d replace it with healthy b o n e [29].  A n i m a l r e s e a r c h on the effectiveness of hyperbaric o x y g e n for the treatment of osteomyelitis dates b a c k to the late 6 0 ' s . T h e s e investigators d e m o n s t r a t e d that a n i m a l s were c u r e d of bone infections with the u s e of hyperbaric o x y g e n a l o n e (i.e. no antibiotics or surgery).  Furthermore, they t r e a t e d another group of rats  prior to inducing infection but found that it did not prevent the infection taking hold. T h e authors c o n c l u d e d that hyperbaric o x y g e n w a s a n  from  effective  treatment modality in treating osteomyelitis b e c a u s e it e n h a n c e s the host's own immune s y s t e m a n d not b e c a u s e it kills the bacteria directly [212].  The  benefits  of  hyperbaric  oxygen  h a v e a l s o b e e n e s t a b l i s h e d in  human  treatments. N e u b a u e r et al [29] reported that the overall s u c c e s s rate in various investigations using hyperbaric o x y g e n therapy on osteomyelitis r a n g e s from 60 to 8 5 % , with a lower rate of recurrence. D a v i s [213] demonstrated in a five y e a r follow-up study, that in 136 patients with refractory chronic osteomyelitis ( c a s e s which failed to heal with antibiotic treatment or surgery) of the s p i n e , extremities, pelvis, skull a n d chest wall, over half of t h e s e patients h a d their infection clear up.  21  A n o t h e r study demonstrated a cure rate of 8 5 % in a two-year follow-up study of 4 0 patients with chronic osteomyelitis. Hyperbaric o x y g e n in this study w a s u s e d a s a n adjunct to surgery a n d antibiotics [214].  Aseptic  Bone  Necrosis  A s e p t i c b o n e n e c r o s i s ( A B N ) o c c u r s - w h e n b o n e b e c o m e s inflamed without being infected. It usually o c c u r s a s a complication of d e c o m p r e s s i o n s i c k n e s s but c a n o c c u r in diabetes, hepatitis, rheumatoid arthritis a n d sickle cell a n e m i a . It m a y also result a s a s i d e effect of various therapeutic p r o c e d u r e s s u c h a s radiation therapy a n d steroid treatment [29]. A B N c a n a l s o arise s p o n t a n e o u s l y in the general population, especially in children a g e s eight to fourteen [215-216].  A B N is essentially a blood supply problem [217]. T h i s blood v e s s e l disruption results in i s c h e m i a or a r e d u c e d blood supply resulting in b o n e not  getting  e n o u g h nutrients a n d o x y g e n . In addition, cellular w a s t e s a c c u m u l a t e [217].  A s e p t i c b o n e n e c r o s i s should be treated properly a s multiple joints are often involved a n d m a y lead to permanent disability. W i d e s p r e a d A B N requires a great deal of joint-replacement surgery, c a u s i n g a large amount of e x p e n s e , pain a n d disability [29].  T h i s is where hyperbaric o x y g e n therapy fits in. It s e e m s logical that increasing ^ the amount of o x y g e n in the affected t i s s u e s m a y halt deterioration a n d promote healing.  N e u b a u e r a n d c o l l e a g u e s [29] have c o n c l u d e d that hyperbaric o x y g e n  d o e s indeed help patients with A B N , h o w e v e r treatment should be long-term. Short-term treatment d o e s relieve pain a n d disability but no lasting cure t a k e s place [218-219].  Fractures  Clinical investigation  on the u s e of H B O on fractures h a s b e e n promising,  especially where non-union a n d c o m p l i c a t e d fractures with i n c r e a s e d c h a n c e of  22  infection are involved [82]. RNA,  F i n d i n g s s u c h a s i n c r e a s e s in osteoblastic D N A a n d  b o n e mineralization, h e m a t o m a a n d c a l l u s formation,  callus  nitrogen  content, capacity for protein s y n t h e s i s , alteration of the homeostatic environment, reduction  in  fibroblastic  osteoblast  proliferation  formation,  capillary  budding, the  c o n c l u s i o n that H B O s p e e d s recovery in fractures a n d r e d u c e s healing time  [75,  greater  and  collagen  a n d o s t e o g e n e s i s have all led to  8 3 , 85].  activity  formation,  R e s e a r c h e r s have c o n c l u d e d that hyperbaric o x y g e n therapy l e a d s to cartilage  production  and  bone  formation  [220].  It  has  also  been  demonstrated to help in bone grafts [221]. In a n a n i m a l study of 4 8 7 fractures in a n d around the joints, r e s e a r c h e r s c o n c l u d e d that hyperbaric o x y g e n therapy u s e d a s an adjunct to conventional orthopedic m e t h o d s , shortened the p r o c e s s of b o n e regeneration a n d w o u n d healing by ten to twelve d a y s [222].  A g a i n , contradiction d o e s exist in the literature w h e r e b y other authors o b s e r v e no d e c r e a s e s in healing time  [86, 87].  T h i s m a y be d u e to the time of onset of  treatment from the actual injury time (12 hours or more after the injury).  Crush  Injuries  / Traumas  / Compartment  Syndrome  C r u s h injuries involve diffuse blood l o s s resulting in tissue hypoxia, which severely affects cellular function [19]. in e d e m a a c c o m p a n i e s the insult.  In addition, i n c r e a s e d blood flow resulting  T h i s e d e m a c a u s e s a n i n c r e a s e d diffusion  distance for o x y g e n to travel to the capillaries  [9].  circulation to the injured a r e a is diminished [88]. reduction  in  blood  flow  in  addition  m a c r o p h a g e to function [44, 45].  to  In c l o s e d c r u s h injuries, all H B O s e r v e s to c a u s e a 2 0 %  providing  a  sufficient  medium  for  Clinical trials have d e m o n s t r a t e d beneficial  effects s u c h a s 1) countering w o u n d infection a c c o m p a n y i n g o p e n traumas of the extremities 2) accelerating the recovery of neutrophil prevention  of  limb  amputation  and  4)  healing  the  phagocytic activity open  suppuration (dead skin with a distinct line of demarcation)  fracture  [13, 66].  3)  without However,  the a b o v e studies failed to c o m p a r e the H B O treated patients with those treated more conservatively, thus not illustrating a c l e a r difference in healing patterns. v. 23  In  a study demonstrating the effects of H B O on the m a n a g e m e n t of s e v e r e t r a u m a of the limbs in older patients with grade III soft-tissue injuries, B o u c h a r d et al [89] found  H B O to  improve  wound  healing a n d  reduce the  repetitive  surgery  n e c e s s a r y in c a s e s of aggravation of crushing t i s s u e d a m a g e . N y l a n d e r et al [90] e x p o s e d rat hind limbs to 3 hours of temporary i s c h e m i a followed by 4 5 minutes of H B O s e s s i o n s at 2.5 A T A . T h e results s h o w e d a significant reduction in posti s c h e m i c e d e m a a n d thus c o n c l u d e d that H B O w a s a useful adjunct in the treatment of acute i s c h e m i c conditions w h e n surgery couldn't be attempted or failed to reverse i s c h e m i a [76, 9 0 , 91]. J o n e s et al [92] c o n d u c t e d a preliminary study on the effects of H B O o n ten patients with c o m p r e s s i v e lesions of the spinal c o r d leading to paralysis. T h e results s u g g e s t e d that by supporting injured spinal cord tissue with o x y g e n under p r e s s u r e , improvement might occur.  A  large s e r i e s of patient, in this type of study, will be n e c e s s a r y before definite c o n c l u s i o n s c a n be drawn on whether H B O therapy i m p r o v e s recovery in the p a r a l y z e d patient with a bruised spinal c o r d . m o d e l s for compartment compartments  causing  syndrome reduced  S t u d i e s c o n d u c t e d on animal  (increased p r e s s u r e in skeletal m u s c l e  capillary  perfusion,  leading  to  ischemia,  nonfunctional a n d n e c r o s i s of tissue) have all s h o w e d promising results of r e d u c e d m u s c u l a r n e c r o s i s a n d e d e m a in H B O treated v e r s u s control groups [81, 84].  Z a m b o n i et al [93,94] have a l s o c o n d u c t e d s e v e r a l experiments  e x a m i n i n g the effects of hyperbaric o x y g e n on i s c h e m i c m u s c l e on animal m o d e l s , again with positive results s h o w i n g a reduction in e d e m a a n d improved m i c r o v a s c u l a r perfusion.  Further r e s e a r c h , however, n e e d s to supplement the  existing body of literature in terms of h u m a n , controlled, double-blinded studies.  Hyperbaric o x y g e n therapy c a n provide various benefits in the treatment of w o u n d s . T o s u m m a r i z e : it e n c o u r a g e s the growth of n e w tissue by providing extra o x y g e n , it counteracts the c h a n c e of infection by indirectly providing cells (i.e. white blood cells) the extra o x y g e n that they n e e d a n d directly by killing a n a e r o b i c o r g a n i s m s , stopping their.multiplication a n d neutralizing the toxins that s o m e produce [12], it e n c o u r a g e s bone repair by supplying o s t e o c l a s t s a n d  24  o s t e o b l a s t s the rich supply of o x y g e n that they require a n d lastly it provides a c l e a r line of d e m a r c a t i o n between tissue which is b e y o n d repair a n d that which c a n be s a v e d [223]. Therefore, the effectiveness of hyperbaric o x y g e n therapy has  b e e n beneficial in minimizing tissue death, reduction  in swelling  and  promoting healing.  2.4 The Role of Hyperbaric Oxygen in Sports and Exercise Medicine T h e r e is a c o n s i d e r a b l e amount of significant r e s e a r c h s u g g e s t i n g the efficacy of H B O therapy a s both a primary a n d adjunct in the treatment of a variety of illnesses a n d injuries. H o w e v e r , only a paucity of k n o w l e d g e exists in terms of its u s e , benefits a n d m e c h a n i s m of action in sport a n d e x e r c i s e - r e l a t e d injuries [Table 7,8]. Although the few studies c o n d u c t e d thus far looking at a c u t e injuries in sports a n d e x e r c i s e h a v e proven to be promising in terms of using H B O a s a treatment modality, t h e s e studies h a v e b e e n limited by their s a m p l e s i z e a n d study d e s i g n [1, 2]. L e a c h et al states,  "the gap between the knowledge gleaned  from the laboratory and severely traumatized patients and the athlete in the locker room remains vast [40]". T h i s statement clearly s u m m a r i z e s how' m u c h r e s e a r c h n e e d s to be c o n d u c t e d to establish or refute the role of hyperbaric o x y g e n therapy in sports a n d e x e r c i s e medicine. Hyperbaric Oxygen - "Applications of HBO arise as a result of medical adventurism; A therapy in search of a disease"  Gabb G & Robin ED  70  Studies  to  date  Oriani et al [88] first s u g g e s t e d the u s e of H B O to a c c e l e r a t e the rate of recovery from injuries suffered in sports. H o w e v e r , it wasn't until recently that a study on H B O w a s first p u b l i s h e d . T h i s study looked at the n u m b e r of d a y s lost to injury in professional s o c c e r players in S c o t l a n d [1].  T h e results s u g g e s t e d a 5 5 %  reduction in days-lost-to-injury b a s e d on a physiotherapists estimation of the time-course for the injury v e r s u s the actual n u m b e r of d a y s lost with routine therapy a n d H B O treatment  sessions.  Although promising, this study w a s  subjective in nature, n e e d e d a control group, input from a n objective third party  25  a n d required a greater h o m o g e n e i t y of injuries. R a n d o m i z e d , controlled, doubleblinded studies with quantifiable injuries are required for significant validity of results.  A n ankle inversion study c o n d u c t e d at T e m p l e University s u g g e s t e d that patients e x p o s e d to H B O treatments returned to activity 3 0 % faster than control g r o u p s [2].  Unfortunately, the results were inconclusive in this study s i n c e a large  amount of variability existed in the study d e s i g n . T h e authors attributed m u c h of this o b s e r v e d variability to the difficulty in quantifying the severity of ankle s p r a i n s [33]. Furthermore, this study h a s b e e n countered by a recent r a n d o m i z e d doubleblind d e s i g n in which H B O treatment did not improve time to recovery after a n a c u t e ankle sprain injury [95].  S t a p l e s et al [96, 97] c o n d u c t e d both a n animal a n d h u m a n study on a m u s c l e injury m o d e l .  T h e animal m o d e l , which m e a s u r e d m y e l o p e r o x i d a s e levels in  treated v e r s u s untreated rats, w a s s u g g e s t i v e of an inhibitory effect of hyperbaric o x y g e n on the inflammatory p r o c e s s or the ability of H B O to actually modulate the injury to the tissue.  T h e h u m a n study e m p l o y e d a r a n d o m i z e d , double-  blinded d e s i g n with controlled start a n d e n d points.  T h e promising results  revealed that the treatment group with H B O h a d a greater recovery of eccentric strength from d e l a y e d - o n s e t m u s c l e s o r e n e s s ( D O M S ) . H o w e v e r , this treatment modality h a d no effect on pain levels [96, 97].  O n e of the best quantitative studies to date c o m e s from a rat m o d e l of surgically lacerated medial collateral ligaments.  T h i s study c o m p a r e d ligament strength  a n d stiffness in injured a n d uninjured ligaments over a n 8-week period a n d c o n c l u d e d that the H B O a p p e a r s to have promoted the return to normal stiffness of the ligaments at 4 w e e k s a s well a s e n h a n c e d recovery of ligament strength [98].  26  A n o t h e r clinical study c o m p l e t e d by S o o l s m a et al [32] at the University of British C o l u m b i a e x a m i n e d the short-term recovery of grade II medial collateral ligament injuries of the k n e e . Positive results s u g g e s t that recovery w a s more rapid in the H B O e x p o s e d individuals a s c o m p a r e d to the control group [32].  A recent study c o m p l e t e d by B e s t et al [99], demonstrated in a rabbit m o d e l , that a  5-day  treatment  regimen  with  H B O a p p e a r s to  improve  functional  and  morphologic recovery at 7 d a y s after a controlled reproducible m u s c l e stretch injury.  C o u n t e r i n g the a b o v e studies is the work c o n d u c t e d by Harrison B C et a l , at the University of C o l o r a d o , which e x a m i n e d the effects of treating e x e r c i s e - i n d u c e d m u s c l e injury v i a hyperbaric o x y g e n . T h e s e r e s e a r c h e r s c o n c l u d e d that H B O w a s not effective  in the treatment  of m u s c l e injury a s e v i d e n c e d by  MRI, C K  r e s p o n s e , isometric strength testing a n d p e r c e i v e d s o r e n e s s [246].  Mekjavic et a l , h a v e also recently reported that H B O is not effective therapy for the treatment of D O M S b a s e d on m a x i m a l isometric m u s c l e strength of the elbow flexor m u s c l e s , right upper arm circumference a n d ratings of p e r c e i v e d m u s c l e s o r e n e s s [225].  1  Although few in number, all of the a b o v e studies provide s o m e insight into the application of H B O a s an adjunctive treatment in sports a n d e x e r c i s e m e d i c i n e , a n d warrants the n e e d for additional r e s e a r c h to better define the therapeutic indications of hyperbaric o x y g e n .  T h e d e m a n d for hyperbaric o x y g e n therapy is increasing throughout the m e d i c a l field. M e d i c a l professionals, clinicians a n d r e s e a r c h e r s have b e e n a n d  are  currently venturing on n e w a r e a s that haven't previously b e e n investigated. A growing interest in sports a n d e x e r c i s e medicine is appearing throughout literature.  the  However, with this new spark of e n t h u s i a s m c o m e s a high d e g r e e of  27  s k e p t i c i s m that h a s b e e n d e v e l o p e d regarding its u s e . T o date only a handful of studies exist in this a r e a a n d of t h e s e studies, only a select few support the benefits of utilizing this intervention.  Table 7: Benefits of HBO in Sports Injuries* • • • •  Reduction of pain and swelling in the acute stage Speeds up recovery and return to activity training Improves fracture healing Aids in the recovery from exhaustion and collapse  *JainKK • 9  Table 8: Proposed Mechanism of Action in Sports Injuries* • • • •  Vasoconstriction Reduction in neutrophil-adhesion Free radical quenching ability Enhancement of leukocyte killing and hydroxyproline formation  *JainKK  9  2.5 Acute Soft Tissue Injury: Delayed Onset Muscle Soreness  Characteristics  of  DOMS  At s o m e point in time, nearly all of us have e x p e r i e n c e d the s e n s a t i o n of d e l a y e d onset m u s c l e s o r e n e s s ( D O M S ) ; m a n y of which have h a d n u m e r o u s encounters with this c o m m o n self-limiting ailment.  E v e n trained individuals will e x p e r i e n c e  s o m e s o r e n e s s following a novel bout of u n a c c u s t o m e d e x e r c i s e [100]. T h i s condition is usually characterized by a s e n s a t i o n of pain a n d discomfort that o c c u r s in skeletal m u s c l e s following  a bout of u n a c c u s t o m e d e x e r c i s e a n d  exertion, a n d is often a c c o m p a n i e d by t e n d e r n e s s a n d stiffness, with a reduction in mobility or flexibility of the m u s c l e s involved.  T h i s r e d u c e d mobility  and  flexibility is e x a c e r b a t e d during palpation, p a s s i v e stretching a n d contraction of the involved m u s c l e s  [101-103].  T h e pain m a y be slight a n d m a y d i s a p p e a r  upon repeated activity or m a y be s e v e r e e n o u g h to interfere with a n d limit 28  movement.  T h i s pain a n d t e n d e r n e s s is usually localized to the distal third  portion of the m u s c l e , in the region of the m u s c u l a r - t e n d i n o u s junction  where  m u s c l e pain receptors are most c o n c e n t r a t e d , with eventual s p r e a d i n g to the center of the m u s c l e belly by 4 8 hours [104]. N e w h a m et al [104] have reported that the pain a s s o c i a t e d with D O M S a p p e a r s medially, laterally a n d then distally, becoming  more  diffuse  throughout  the  muscle 24-48  hours  post-exercise.  G e n e r a l l y , however, the pain is evident throughout most of the affected m u s c l e belly.  T h e s o r e n e s s that is prominent with D O M S initially a p p e a r s 8-24 hours  after e x e r c i s e , i n c r e a s e s in intensity within the first 2 4 hours, p e a k s from 2 4 - 7 2 hours a n d finally s u b s i d e s 5-7 d a y s p o s t - e x e r c i s e [102, 105, 106].  Type of Activity Inducing Muscle Injury High intensity, short duration e x e r c i s e results in the largest increase in m u s c l e d a m a g e a n d D O M S [107].  Eccentric e x e r c i s e (e.g. downhill running - involving  forced lengthening of a m u s c l e a s it d e v e l o p s tension [108]) h a s b e e n s h o w n to produce  greater  damage  to  the  m u s c l e fibres  as  o p p o s e d to  concentric  contractions (e.g. uphill running), s i n c e the former requires lower energy c o s t s a s fewer motor units are activated for a given load submaximal  has  been  demonstrated to be lower during eccentric (negative) work than concentric  [111,  112].  force  or  power  output,  EMG  [104, 109, 110]. At a given  activity  in  muscle  T h i s leads to higher tensions per c r o s s - s e c t i o n a l a r e a of active skeletal  m u s c l e fibres [104,  110].  The  i n c r e a s e d tension could c a u s e m e c h a n i c a l  disruption of the structural e l e m e n t s in the m u s c l e fibres t h e m s e l v e s  [102, 104,  110, 113] or in the connective tissue that is in s e r i e s with the contractile e l e m e n t s [114].  E v a n s [115, 116] s u g g e s t e d that the r e a s o n why eccentric e x e r c i s e  c a u s e s far greater a m o u n t s of m u s c l e d a m a g e than concentric e x e r c i s e might be d u e to different fiber recruitment patterns. In other words, eccentric contraction conditions provide a situation where relatively few fibres are recruited a n d are producing relatively large forces  [110]. C l e a k [105] reported that lengthening  m u s c l e fibres i m p o s e s additional tension a n d stretch on connective tissue a n d a p p e a r s to increase localized s o r e n e s s in the region of the m u s c l e involved.  29  C a n n o n et al  [108] cite that repeated force lengthening of a m u s c l e a s it  d e v e l o p s tension c a u s e s immediate ultrastructural d a m a g e to the s a r c o m e r e s , followed  by d e l a y e d - o n s e t m u s c l e s o r e n e s s a n d a  r e l e a s e of  myocellular  e n z y m e s . C o n s e q u e n t l y , it h a s b e e n reported that eccentric work requires l e s s o x y g e n , lower a m o u n t s of A T P , p r o d u c e s l e s s lactate a n d s h o w s lower motor unit activity than concentric work [114, 117-120]. Furthermore, eccentric contractions require l e s s time to reach p e a k tension [113].  M o r e recent e v i d e n c e s u g g e s t s that m u s c l e d a m a g e is not a function of absolute force or tension g e n e r a t e d but the magnitude of strain during active lengthening [121].  T h e s e authors determined that active strain during eccentric work w a s  related to the s p e e d at which a m u s c l e w a s lengthened while it w a s contracting. It has  b e e n s u g g e s t e d that the  greater d a m a g e elicited  by faster  eccentric  contractions w a s d u e to the fact that cross-bridge cycling c o u l d not k e e p p a c e with the c h a n g e in length of the m u s c l e [122].  M u s c l e fiber type h a s also b e e n reported to be a c a u s e for m u s c l e d a m a g e .  It  h a s b e e n s u g g e s t e d that specific d a m a g e c a n be noticed in the type 2 B fast glycolytic fibers, thus hypothesizing that m u s c l e injury m a y be a result of the m u s c l e fiber, oxidative capacity [227]. It is b e l i e v e d that type 2 fibers might be more  susceptible to  stretch  induced  injury  b e c a u s e of  a  less developed  e n d o m y s i u m that type 1 fibers [228].  F r o m a review of the literature, it s e e m s apparent that m u s c l e d a m a g e m a y not only be a result of high force generation but the magnitude of strain during active lengthening m a y a l s o play a role in the injury p r o c e s s .  Eccentric  Strength  Loss  Accompanying  DOMS  Throughout the literature, it is evident that D O M S is a c o n s e q u e n c e of m u s c l e "over-use" [100-107].  A n y activity in which the m u s c l e p r o d u c e s high forces or  forces over a longer period of time is c a p a b l e of producing the s e n s a t i o n of  30  DOMS  [107]. In addition, although the d e g r e e of s o r e n e s s is related to both the  intensity of the m u s c u l a r contractions a n d the duration of e x e r c i s e , intensity s e e m s to be the more important determinant of the two [101, 107, 123].  Hough  [124] w a s the first to d e s c r i b e in detail the p h e n o m e n o n of D O M S , hypothesizing the etiology a n d m e c h a n i s m s involved.  H e d e m o n s t r a t e d that D O M S w a s a  result of structural d a m a g e to the m u s c l e and/or c o n n e c t i v e tissue a n d to a reduction in m u s c u l a r performance  [124].  T h i s reduction in performance m a y  result from a reduction in voluntary effort due to the s e n s a t i o n of s o r e n e s s and/or a lowered inherent capacity of the m u s c l e to produce force [101]. Therefore, DOMS  is a l s o a c c o m p a n i e d by a significant  d e c r e a s e in  force-generating  c a p a c i t y of the m u s c l e resulting in eccentric strength d e c r e m e n t s [125].  The  time c o u r s e for the d e v e l o p m e n t of s o r e n e s s a n d l o s s of strength s u g g e s t s little or no relationship b e t w e e n the two parameters  [112].  Newham and colleagues  [110] found that m a x i m a l voluntary force of the knee e x t e n s o r s h a d returned to normal by 2 4 hours after e x e r c i s e while s o r e n e s s at this time w a s most intense. Furthermore, H o u g h s h o w e d that the o c c u r r e n c e of d e l a y e d pain w a s directly related to the p e a k f o r c e s d e v e l o p e d a n d to the rate of force d e v e l o p m e n t in rhythmic contractions, but not to the rate of fatigue  [124, 126].  A reduction in  m a x i m u m force production h a s b e e n o b s e r v e d after eccentric e x e r c i s e , a s early a s 1-hour post-exercise  [113, 127]. Although N e w h a m a n d c o l l e a g u e s h a v e  s u g g e s t e d that strength returns to pre-exercise levels within 2 4 hours  [110],  others h a v e reported a return to b a s e l i n e levels a s long a s 1 w e e k  [113].  G l e e s o n a n d c o l l e a g u e s [128]  have reported that d e c r e m e n t s in  maximum  isometric force ( 5 0 % of normal) are greatest immediately following eccentric e x e r c i s e , with recovery taking p l a c e by d a y s 4-7.  In addition, they report that a  d e c r e a s e in m a x i m u m d y n a m i c power output ( 8 0 % of normal) persists up to 4 days.  *  M a c l n t y r e a n d colleague's [129] have reported a bimodal pattern of eccentric torque that o c c u r s 0 a n d 2 0 - 2 4 hours p o s t - e x e r c i s e . T h i s bimodal pattern w a s the first report demonstrating this pattern in h u m a n s .  F a u l k n e r et al [130] reported  31  two d e c l i n e s in the m u s c l e force in a n animal m o d e l . T h e y s u g g e s t that the initial decline in force m a y b e a function of m e c h a n i c a l injury a n d fatigue myofibrillar disruption at the level of the Z-line), leading to a n acute response  (including  inflammatory  [129]. F a u l k n e r a n d c o l l e a g u e s [130] further s u g g e s t that the s e c o n d  decline in force o c c u r s in r e s p o n s e to phagocytic activity at the site of the initial d a m a g e . T h i s deficit in force d o e s not a p p e a r to b e related to the level of s o r e n e s s s i n c e it o c c u r s prior to the s o r e n e s s a n d c a n remain for a greater period [131]. Maclntyre et al [132] h a v e a l s o reported that no relationship exists between the development of s o r e n e s s a n d l o s s of m u s c l e strength s i n c e the latter a p p e a r s immediately after e x e r c i s e .  Therefore, this bimodal pattern of eccentric torque  further grounds support for the theory that more than o n e m e c h a n i s m is involved in e x e r c i s e - i n d u c e d m u s c l e s o r e n e s s  Theories  Associated  with  [129,130].  DOMS  T h e pain a n d discomfort a s s o c i a t e d with D O M S h a s b e e n studied extensively s i n c e 1 9 0 2 a n d a s a c o n s e q u e n c e , several theories h a v e b e e n s u g g e s t e d to explain this condition. accumulation  T h e o r i e s s u c h a s the lactic  in the muscle), muscle  spasm  theory  acid  theory  (originally  D e V r i e s [133],in 1961 that e x e r c i s e c a u s e s i s c h e m i a which production of a pain s u b s t a n c e ) , connective  tissue damage  (lactic  acid  d e s c r i b e d by results  in the  theory (rupture a n d  d a m a g e of the m u s c l e , predominantly the connective tissue), muscle  damage  theory (skeletal m u s c l e d a m a g e the primary m e c h a n i s m contributing to m u s c l e s o r e n e s s ) a n d inflammation  (tissue d a m a g e triggering a n inflammatory response)  a s c a u s e s of D O M S h a v e b e e n postulated [105, 134].  B a s e d o n a review of the available published literature, it a p p e a r s that D O M S is a result of two p r o c e s s e s , a m e c h a n i c a l a n d b i o c h e m i c a l p r o c e s s  [102, 132]. A  s e q u e n c e of events representing t h e s e two p r o c e s s e s during fatiguing e x e r c i s e is outlined in Figure 2 [ 1 0 1 , 1 2 6 , 135].  \  3 2  Figure 2: S e q u e n c e of events a s s o c i a t e d with d e l a y e d onset m u s c l e s o r e n e s s , including m e c h a n i c a l a n d b i o c h e m i c a l p r o c e s s e s .  S e q u e n c e Of E v e n t s High MechanicalForce  disruption of —) structural — structural proteins damage to in muscle fibre and sarcolemma connective tissue  influx Ca2+ frominterstium to muscle fibre  mitochondrial accumulation of ions  i macrophage  attraction of monocytes  degradationsstructural components of contractile apparatus  accumulation of histamine and " kinins  increased pressure from edema  activation of nociceptors  activation of proteolytic enzymes  inhibition of cellular respiration  DOMS  'adapted from Appell et al , Armstrongi101, et 102 al 126  33  S t a u b e r a n d c o l l e a g u e s [136] h a v e s u g g e s t e d that the pain a n d a s s o c i a t e d with D O M S  inflammation  may be d u e to the swelling a n d disruption  of  the  extracellular matrix. In contrast, J o n e s et al [137] h a v e reported the stiffness to be a result of connective tissue d a m a g e . P y n e [138,  139] & E b b e l i n g a n d  C l a r k s o n [112] h a v e s u g g e s t e d that m e c h a n i c a l stress ( m e c h a n i c a l s h e a r force production during exercise) a n d metabolic s t r e s s (disturbances in normal cellular metabolism  provoked  by exhaustive exercise) account for  exercise-initiated  d a m a g e to skeletal m u s c l e fibres. Within the connective tissue of the m u s c l e s are myelinated group III (A-delta) a n d unmyelinated group'IV (C) afferent receptors. T h e large myelinated group III fibers are believed to transmit "sharp" l o c a l i z e d pain, w h e r e a s the group IV fibers carry "dull", diffuse pain. T h e r e f o r e , it s e e m s likely that the group IV receptors carry the s e n s a t i o n of D O M S s i n c e the pain is usually dull a n d diffuse, with the free nerve e n d i n g s responding to m e c h a n i c a l a n d c h e m i c a l (metaboceptors) a s well a s noxious stimuli (nociceptors)  [112].  Bradykinin, serotonin a n d histamine m a y activate the free nerve e n d i n g s of nociceptors to produce s o r e n e s s a n d pain e x p e r i e n c e d after e x e r c i s e  [138].  W a r h o l et al [140] reported c o n s i d e r a b l e d i s t u r b a n c e s in the contractile apparatus of the g a s t r o c n e m i u s m u s c l e during competitive  marathon  running.  Z-band  streaming, myofibrillar lysis a n d contracture b a n d s were noted upon histological examination.  Furthermore, pathological c h a n g e s o c c u r r e d in the mitochondria,  s h o w i n g focal swelling a n d crystalline  inclusions a n d the s a r c o l e m m a  and  sarcotubular s y s t e m s h o w e d dilatation a n d disruption [126, 141]. C r e n s h a w a n d c o l l e a g u e s [103, 142] investigated whether D O M S of the v a s t u s lateralis m u s c l e w a s a s s o c i a t e d with elevated intramuscular p r e s s u r e . Intramuscular  pressure  (IMP) is defined a s the fluid p r e s s u r e c r e a t e d by a m u s c l e during contraction [142] a n d is correlated linearly with the force of contraction during isometric a n d isokinetic e x e r c i s e [142]. B a s e d on their findings, they first c o n c l u d e d that D O M S of the v a s t u s lateralis m u s c l e is a s s o c i a t e d with extensive intracellular swelling a n d with elevated IMP  [103], h o w e v e r in a follow-up study, they reported that  IMP w a s not an etiologic indicator of D O M S [142]. N e w h a m [143] s u g g e s t e d that  34  intramuscular p r e s s u r e s are raised in s o m e , but not all, painful  compartments  a n d e v e n w h e n raised, follow a different time c o u r s e to the pain that a p p e a r s .  B i o c h e m i c a l l y , a variety of clinical m e a s u r e s h a v e b e e n a s s o c i a t e d with D O M S . M u s c l e fibres contain proteolytic e n z y m e s that are r e l e a s e d following injury a n d initiate degradation of lipid a n d protein structures in the injured cell [144]. T h e p r e s e n c e of intramuscular e n z y m e s in the blood h a s b e e n c o n s i d e r e d to be indicative of d a m a g e to m u s c l e fibres, particularly to the s a r c o l e m m a [101]. N u m e r o u s reports s u g g e s t the time c o u r s e s of i n c r e a s e d levels of  plasma  e n z y m e s are similar to the time c o u r s e of D O M S following e x e r c i s e a n d the intensity of s o r e n e s s a n d level of p l a s m a e n z y m e s are a l s o correlated [107, 119, 145].  H o w e v e r , Donnelly et al [146] h a s s u g g e s t e d that m u s c l e e n z y m e r e l e a s e  a n d m u s c l e s o r e n e s s are unrelated. T h i s s u g g e s t i o n w a s b a s e d on the findings that decline in m u s c l e strength a n d in 5 0 % e n d u r a n c e time did not differ between the first a n d s e c o n d period (10 w e e k gap) of the study, indicating 1) that the repeat bout effect for m u s c l e e n z y m e r e l e a s e w a s not demonstrated a n d 2) m u s c l e s o r e n e s s r e a c h e d the s a m e level after both e x e r c i s e bouts.  In contrast,  Armstrong et al [101] demonstrated in an animal m o d e l that elevations in p l a s m a e n z y m e s might o c c u r simultaneously with e x e r c i s e - i n d u c e d n e c r o s i s of skeletal m u s c l e fibers. P l a s m a e n z y m e s s u c h a s myoglobin ( 1 8 , 0 0 0 - D a heme-containing o x y g e n carrier protein of skeletal m u s c l e cells [147]), hydroxyproline,  creatine  kinase ( 8 0 , 0 0 0 - D a e n z y m e found in large concentrations in m u s c l e tissue [147]), a n d h y p e r k a l e m i a have b e e n indicative of m u s c l e injury [101]. Increases in s e r u m activities  of  enzymes  glutamic-oxaloacetic  transaminase  (GOT),  lactic  d e h y d r o g e n a s e (LDH) a n d aspartate a m i n o t r a n s a m i n a s e ( A S T ) h a s also b e e n reported  to  reflect  muscle  fibre  permeability [138, 139, 148-151].  damage  involving  increased  membrane  Mair a n d c o l l e a g u e s [152] h a v e reported that  a n increase in myosin heavy chain ( M H C ) fragment p l a s m a concentrations is demonstrated after eccentric e x e r c i s e .  35  C r e a t i n e k i n a s e ( C K ) is found almost exclusively in m u s c l e tissue a n d is therefore c o n s i d e r e d the most c o m m o n indicator of skeletal m u s c l e d a m a g e [153]. T h e i n c r e a s e in s e r u m or p l a s m a C K activity after e x e r c i s e is d e l a y e d a n d the extent of delay d e p e n d s upon the type of e x e r c i s e [112]. T o explain this delay in C K release, Clarkson and Tremblay  [154] h a v e theorized that e x e r c i s e - i n d u c e d  d a m a g e m a y c a u s e a n accumulation of C a  2 +  resulting in; a) production of noxious  stimuli s u c h a s bradykinin a n d histamine c a u s i n g m u s c l e s o r e n e s s , b) m u s c l e contractures  leading  sarcoplasmic  reticulum  to  decreased  range  a n d mitochondrial  of  motion,  functioning  c)  impairment  a n d d) activation  of of  s a r c o p l a s m i c p r o t e a s e s resulting in l o s s of s a r c o l e m m a l integrity a n d d e l a y e d release of C K . Although C K h a s b e e n widely u s e d a s a clinical marker for d a m a g e to the m u s c l e , the relationship between the magnitude of C K r e l e a s e and  histological  e v i d e n c e of the extent  of m u s c l e  damage  h a s not b e e n  established [155].  Smith [156] p r o p o s e d that the most likely c h e m i c a l stimulant  for the induction  of D O M S  m a y be prostaglandin  E  2  (PGE ) 2  that c a u s e s  i n c r e a s e d sensitivity of pain receptors a n d that invading m a c r o p h a g e h a v e the capability of synthesizing P G E PGE  2  2  [156]. Similarities in time c o u r s e for i n c r e a s e s in  a n d D O M S (p<0.05), 2 4 hours after a bout of eccentric e x e r c i s e w a s  reported by Smith a n d c o l l e a g u e s [157]. Furthermore, S a l m i n e n a n d c o l l e a g u e s [158] have demonstrated in a n animal study that Indomethacin, a non-steroidal anti-inflammatory drug that inhibits prostaglandin synthesis, h a d reduced d a m a g e to the m u s c l e in e x e r c i s e d m i c e . O n the other h a n d , K u i p e r s a n d c o l l e a g u e s [159] found that prostaglandins are not involved in a n e x e r c i s e - i n d u c e d r e s p o n s e s i n c e flurbiprofen effect on m u s c l e s o r e n e s s .  inflammatory  ( c y c l o - o x y g e n a s e inhibiting drug) did not have a n y Smith a n d c o l l e a g u e s [160] a l s o studied  whether  there would be a reduction in total cholesterol levels in r e s p o n s e to microtrauma induced by eccentric activity.  T h i s w a s founded on the notion that cholesterol is  a c o m p o n e n t of the cell m e m b r a n e a n d that cholesterol levels a r e temporarily reduced in r e s p o n s e to trauma a n d post-surgery. Interestingly, the findings from this study s h o w e d a significant d e c r e a s e in total cholesterol for both groups e x a m i n e d [160].  It h a s a l s o b e e n s u g g e s t e d that thiol proteases, s u c h a s  36  c a l p a i n , d e g r a d e structural proteins s u c h a s alpha-actinin, resulting in Z-line streaming a n d disorganization of the normal alignment of the myofilaments in conditions of altered metabolic and/or, functional d e m a n d s [132].  In a n a n i m a l  m o d e l involving the rat hindlimb m u s c l e , B e l c a s t r o a n d c o l l e a g u e s [161] h a v e reported i n c r e a s e d c a l p a i n activity following level treadmill running, p r o p o s i n g that this  i n c r e a s e d activation  of skeletal  muscle calpain  m a y result  from  i n c r e a s e d intracellular c a l c i u m . A p p e l l et al [126] s h o w e d that nifedipine,  a  c a l c i u m c h a n n e l blocker, diminished e x e r c i s e - i n d u c e d m u s c l e d a m a g e in a m o u s e m o d e l , supporting  the notion  that C a  2 +  ions a r e implicated  in the  mediation of tissue d a m a g e .  Inflammation  and Delayed-Onset  Muscle  Soreness  It h a s b e e n s u g g e s t e d that m u s c l e s o r e n e s s is related to a n response.  inflammatory  Inflammation is the body's normal r e s p o n s e to a n insult, s u c h a s a n  injury, infection or antigen. T h e p u r p o s e of this r e s p o n s e is to c l e a r a n d eliminate d a m a g e d tissue a n d microbial invaders, thus leading to tissue reparation. T h e m a i n pathological feature of inflammation c o n s i s t s of leukocyte infiltration a n d exudation of p l a s m a into the lesion in the early stage followed by proliferation of connective  tissue  including  fibroblasts,  which  leads  to  the formation  of  granulation tissue.  T w o sub-classifications of inflammation exist. First a n d foremost is the acute inflammatory p r o c e s s with local a n d systemic c h a n g e s c h a r a c t e r i z e d by a rapid change  in blood flow  a n d a c c o m p a n i e d by a n immigration  of  neutrophils  (histological hallmark of acute inflammation) a n d m o n o c y t e s . Neutrophils a r e k e y nonspecific host d e f e n s e cells responsible for p h a g o c y t o s i s of microbial, bacterial a n d viral p a t h o g e n s  [139, 162]. T h e y play both efferent  (phagocytosis a n d  degranulation) a n d afferent (release of immunomodulatory m o l e c u l e s ) roles in the immune r e s p o n s e [139]. T h e cardinal s i g n s of r e d n e s s , swelling, heat a n d  pain; "rubor et tumor cum calore et dalore" (Cornelius Celsus) are noted in the acute-local inflammatory p h a s e [163, 164]. In addition, the induction of fever  37  (due to the production of IL-1, T N F , IFN-alpha) a n d the production of p l a s m a proteins constitute the s y s t e m i c reaction.  Chronic  inflammation  is c h a r a c t e r i z e d by the p r e s e n c e of l y m p h o c y t e s a n d  m o n o c y t e s [132]. M o n o c y t e s a n d m a c r o p h a g e s are primarily r e s p o n s i b l e for the removal of neutrophils a n d necrotic tissue [116, 132, 165]. After a n insult o c c u r s to the tissue, the body r e s p o n d s both at a v a s c u l a r a n d cellular level. T h e former involves vasoconstriction (5-10 minutes), followed by vasodilation a n d i n c r e a s e d v a s c u l a r permeability  [156, 166]. T h e latter r e s p o n s e involves the interaction of  v a r i o u s inflammatory mediators, mainly neutrophils a n d m o n o c y t e s . A short time after the t r a u m a o c c u r s , circulating neutrophils dramatically i n c r e a s e in number, aggregating at the site of injury, reaching p e a k concentrations approximately 1-4 hours post-injury. T h i s initial i n c r e a s e in neutrophils in the capillary b e d of the injured tissue is due to the slowing d o w n of blood flow within the capillary a n d the leak of p l a s m a proteins from the capillary [132]. Smith et al [167] demonstrated a m a r k e d i n c r e a s e , a b o v e b a s e l i n e , in neutrophil levels b e t w e e n 1 a n d 2 hours post-exercise.  It h a s b e e n d e m o n s t r a t e d that during t h i s . p e r i o d , neutrophils  b e c o m e intimately a s s o c i a t e d with the endothelial cells, upregulating a d h e s i o n proteins ( C D 1 1 / C D 1 8 ) that allow the neutrophils to role along the endothelial surface, b e c o m e adherent a n d eventually migrating out into the tissue [132].  It  is important to realize that only the p r e s e n c e of neutrophils in the interstitum or muscle  is indicative  of  s e v e r e inflammation  [132].  After this  peak,  the  concentration d e c l i n e s rapidly a n d is followed by the migration of m o n o c y t e s . T h i s immigration rises in concentration a n d is maintained for 4 8 hours, after which the m o n o c y t e s mature into m a c r o p h a g e . M a c r o p h a g e s play a pivotal role in the recovery p r o c e s s following e x e r c i s e - i n d u c e d m u s c l e injury adult  macrophage  remove  necrotic  tissue a n d foreign  M o n o c y t e s / m a c r o p h a g e are responsible for the  [132]. T h e s e  b o d i e s [156,  166].  resorption of neutrophils  necrotic tissue a n d the sequestration of foreign material or antigens  in  [132].  Furthermore, they are c a p a b l e of producing a wide variety of cytokines in large a m o u n t s , thus contributing to the cytokine network. In addition, they play a pivotal  38  role in the specific r e s p o n s e of T a n d B-lymphocytes to antigen [132]. T h e r e is a great deal of e v i d e n c e s u g g e s t i n g the role of m o n o c y t e s / m a c r o p h a g e  during  eccentric e x e r c i s e , both in animal a n d h u m a n m o d e l s .  B e n d s t r u p [168] w a s the first to h y p o t h e s i z e that tissue d a m a g e resulting from intense e x e r c i s e m a y trigger a n inflammatory r e s p o n s e a n d the time required for the  r e s p o n s e to  o c c u r e x p l a i n s the  s o r e n e s s delay  [169].  It  has  been  demonstrated that e x e r c i s e - i n d u c e d m u s c l e injury triggers mobilization of s o m e a s p e c t s of the  inflammatory  response  [132], h o w e v e r the  specific  events  initiating this s e e m s s o m e w h a t unclear. It m a y be p o s s i b l e that the inflammatory r e s p o n s e m a y be responsible for initiating, amplifying and/or resolving skeletal m u s c l e injury [132].  It h a s b e e n demonstrated that cytokines play a role in the i m m u n e r e s p o n s e following s t r e n u o u s e x e r c i s e . H o w e v e r , Northoff et al [234] h a s reported that c h a n g e s in cytokine levels o b s e r v e d in s e r u m or p l a s m a are a l w a y s subtle, thus explaining significance in s o m e studies while being borderline significant  or  undetectable in others. C y t o k i n e s are essential c o m p o n e n t s of our d e f e n s e a n d repair  systems  but  immunoinflammatory  also  potentially  harmful  mediators  of  infectious  and  reactions [245]. C y t o k i n e s are r e l e a s e d at the site  of  inflammation w h e n there is a local r e s p o n s e to a n infection or tissue injury. T h e y facilitate an influx of lymphocytes, neutrophils, m o n o c y t e s a n d other cells into the tissue, a n d t h e s e cells participate in the c l e a r a n c e of the antigen a n d the healing of the tissue  [187, 188]. A c c o m p a n y i n g this local inflammatory r e s p o n s e is a  s y s t e m i c r e s p o n s e known a s the acute p h a s e r e s p o n s e . T h e  "inflammatory"  cytokines p r o d u c e d a s a result of this r e s p o n s e include tumor n e c r o s i s factor-a ( T N F - a ) , interleukin 1 (IL-1) a n d interleukin 6 (IL-6). IL-1 a n d T N F i n c r e a s e after e x e r c i s e a n d induce the release of a third cytokine, IL-6. Both IL-1 a n d T N F h a v e proinflammatory effects while IL-6 h a s b e e n cited to be restorative in nature, with anti-inflammatory a n d i m m u n o s u p p r e s s i v e effects [139, 231].  39  First  introduced  in  1980  by  Weissenbach  [235],  IL-6  is  a  pleiotropic,  "multifunctional" cytokine involved in the regulation of i m m u n e r e s p o n s e s , the acute p h a s e r e s p o n s e ( A P R ) a n d h e m a t o p o i e s i s [238].  M a n y different cells  p r o d u c e it after stimulation during infection, t r a u m a or immunological c h a l l e n g e [231]. Its receptor s y s t e m c o n s i s t s of two m o l e c u l e s : a ligand-binding 8 0 - k D a m o l e c u l e a n d a non-ligand binding signal transducer, gp 130, both of which w e r e found to belong to the cytokine receptor family [232]. A theoretical m o d e l of the immunological a n d inflammatory  r e s p o n s e s to e x e r c i s e a n d m u s c l e d a m a g e  s h o w i n g the central role of cytokines a n d neutrophils in the repair of d a m a g e d t i s s u e w a s p r o p o s e d by P y n e [138, 139] [Figure 3].  S e v e r a l authors  have  reported that eccentric e x e r c i s e c a u s i n g m u s c l e d a m a g e is a s s o c i a t e d with a n i n c r e a s e in s e r u m IL-6 concentrations a n d this increase is significantly correlated with the concentration of creatine k i n a s e in the d a y s following e x e r c i s e [150, 187, 188, 189]. T h e time c o u r s e of cytokine production, the c l o s e a s s o c i a t i o n with m u s c l e d a m a g e a n d the finding of i n c r e a s e d IL-6 after intense e x e r c i s e support the idea that during eccentric e x e r c i s e , myofibers are m e c h a n i c a l l y d a m a g e d , thus stimulating the local production of inflammatory cytokines [187]. R o h d e et al [188]  h a v e reported that eccentric e x e r c i s e induced an i n c r e a s e in p l a s m a  concentrations of IL-6 by 5 7 0 % , 2 hours post-exercise a n d return to p r e - e x e r c i s e levels by day 2. B a u e r et al [233] h a v e cited that elevated levels of IL-6 c a n be found a s early a s a few hours after the onset of a pathogenic event a n d m a y persist for only a few hours or up to a few d a y s . Furthermore, B r u u n s g a a r d et al [189] demonstrated that IL-6 levels i n c r e a s e d five-fold a n d C K levels i n c r e a s e d almost 4 0 fold, 4 d a y s post-eccentric e x e r c i s e . Furthermore,  IL-6 s e e m s to  be  the o n e cytokine that provides the most reliable results, being e l e v a t e d shortly after strenuous e x e r c i s e [234].  40  Figure 3: Theoretical model showing role of cytokines and neutrophils during exercise inducing damage to skeletal muscle . , 1 3 9  Theoretical Model Showing Role Cytokines & Neutrophils Exercise  Mechanical Stress  Oxidative Damage  —•  Cytokines (IL-1, 2, 6, TNF)  _^  1  pain  Metabolic Stress  hormonal activation  acute phase response —  macrophage  I  chemoattractants  i  neutrophils  I  —  repair tissue damage  1  growth/repair/restorative  .'"adapted from Pyne DB [138, 139)  41  Free  Radicals  and  Exercise-Induced  Muscle  Damage  During intense e x e r c i s e , whole body o x y g e n uptake c a n i n c r e a s e 20-fold a b o v e resting levels a n d in active m u s c l e fibers, o x y g e n c o n s u m p t i o n m a y rise 200-fold [237, 238].  It h a s b e e n estimated that 4 - 5 % of the o x y g e n c o n s u m e d during  respiration is not completely r e d u c e d to water, instead forming free radicals [240]. A growing a m o u n t of e v i d e n c e indicates that free radicals play a n important role a s mediators of skeletal m u s c l e d a m a g e a n d inflammation  [171].  F r e e radicals  are c h e m i c a l s p e c i e s with one or more unpaired electrons in their outer orbit m a k i n g them highly reactive [112] s i n c e they strive to b a l a n c e their unpaired electrons by c o m b i n i n g with electrons with opposite s p i n s in other s u b s t a n c e s [239].  O x y g e n free radicals are i n c r e a s e d during e x e r c i s e a s a result of  i n c r e a s e s in mitochondrial o x y g e n c o n s u m p t i o n a n d electron transport  flux,  inducing lipid peroxidation [239] [Table 9]. T h e c a s c a d e of lipid peroxidation is characteristic of inflammation  in D O M S .  Lipid peroxidation initiated by free  radicals d e c r e a s e s the barrier function of cell m e m b r a n e s a n d m a y b e a s s o c i a t e d with m u s c l e fibre n e c r o s i s a n d e n z y m e r e l e a s e following d a m a g i n g e x e r c i s e [112].  During e x e r c i s e , two potentially harmful free radical generating s o u r c e s  are s e m i q u i n o n e (in the mitochondria) a n d xanthine o x i d a s e (in the capillary endothelial cells).  \  W h e n the microcirculation is d a m a g e d , free radical formation  may  activate  proteolytic e n z y m e s [126, 2 2 9 , 230]. R e a c t i v e o x y g e n s p e c i e s are a l s o activated during i s c h e m i a / h y p o x i a a n d s u b s e q u e n t reperfusion/oxygenation m u s c l e [229]. Degradation of a d e n o s i n e triphosphate  in skeletal  ( A T P ) forms xanthine,  which l e a d s to a c a s c a d e effect on xanthine d e h y d r o g e n a s e , xanthine o x i d a s e and  uric  acid,  causing  malfunctioning  of  the  ion  pumps  and  increasing  intracellular levels of c a l c i u m [111, 229]. T h e resulting effect is the generation of o x y g e n free radicals, which induce disruption of phospholipid layers a n d lipid peroxidation [229].  42  Sjodin et al [171] h a v e p r o p o s e d that high intensity e x e r c i s e i n c r e a s e s the flow of  oxygen  through  the  skeletal  muscles,  causing  metabolic  b i o c h e m i c a l c h a n g e s leading the skeletal m u s c l e d a m a g e a n d  stress  and  inflammation  [Figure 4].  C o n t r o v e r s y a l s o exists on the role of hyperbaric o x y g e n therapy in free radicalmediated tissue injury. Hyperbaric o x y g e n h a s b e e n s h o w n to e n h a n c e the antioxidative d e f e n s e m e c h a n i s m s in s o m e a n i m a l m o d e l s , but it h a s a l s o b e e n reported to i n c r e a s e the production of o x y g e n free radicals  [193]. S e v e r a l  authors have reported that o x y g e n under certain conditions c a n generate highly reactive free radicals that mediate tissue injury a n d impair the p r o c e s s of w o u n d healing while others h a v e reported that hyperoxia i n c r e a s e s the b i o c h e m i c a l d e f e n s e m e c h a n i s m s against free radicals [193].  T o a s s e s s the level of lipid peroxidation that o c c u r s during oxidative s t r e s s on tissue, s e v e r a l reliable, analytical methods are available. T h e a s s e s s m e n t of m a l o n d i a l d e h y d e ( M D A ) , a product of lipid peroxidation, h a s b e c o m e the most c o m m o n technique to m e a s u r e the d e g r e e of oxidative d a m a g e in biological s y s t e m s [190]. M a l o n d i a l d e h y d e is o n e of s e v e r a l low-molecular weight  end  products formed v i a the d e c o m p o s i t i o n of certain primary a n d s e c o n d a r y lipid peroxidation products [191, 192]. complex  with  various  tissue  M D A , which m a y exist in a free form or a s a  constituents,  is formed  during  the  oxidative  degradation of s o m e m a c r o m o l e c u l e s , a s a product of free radical generation by ionizing radiation in vivo, a n d a s a by-product of prostaglandin biosynthesis [190]. M D A is formed during the last s t a g e s of the b r e a k d o w n of e n d o p e r o x i d e s formed during intramolecular rearrangements in the structure of. polyunsaturated fatty a c i d s [190]. A m o n g the various methods to evaluate m a l o n d i a l d e h y d e , which include direct spectrophotometry  or high-pressure liquid chromatography,  the  reaction with thiobarbituric acid (TBA) to form a colored adduct a p p e a r s a s a more rapid, inexpensive a n d sensitive technique  [190, 191]. T h e s a m p l e under  investigation is heated with T B A at low p H , a n d a pink c h r o m o g e n (a T B A 2  43  m a l o n d i a l d e h y d e adduct) is m e a s u r e d by its a b s o r b a n c e at or c l o s e to 5 3 2 nm or by f l u o r e s c e n c e at 5 5 3 nm [236]. T h e f l u o r e s c e n c e technique will be applied for our p u r p o s e s in examining lipid peroxidation resulting from e x e r c i s e - i n d u c e d muscle damage.  >  T a b l e 9: M e c h a n i s m by w h i c h e x e r c i s e g e n e r a t e s free radicals * • • •  Increases in epinephrine a n d other c a t e c h o l a m i n e s that c a n produce o x y g e n radicals w h e n they are metabolically inactivated Production of lactic acid that c a n convert a w e a k l y d a m a g i n g free radical (superoxide) into a strongly d a m a g i n g o n e (hydroxyl) Inflammatory r e s p o n s e s to s e c o n d a r y m u s c l e d a m a g e incurred with overexertion *Clarkson PM, et al 2000  44  Figure 4: B i o c h e m i c a l m e c h a n i s m for oxygen-free radical formation resulting in skeletal m u s c l e d a m a g e a n d inflammation during e x e r c i s e  Biochemical Mechanism For Oxygen-Free Radical Formation During Exercise High intensity exercise —  Increased flow of ~ " oxygen thru skeletal muscle cells  Skeletal muscle Cellular damage and necrosis inflammation (lipid peroxidation of cell membrane)  ATP utilization — greater than ATP generation  Loss of cell viability 4*  Metabolic Stress ™  Biochemical changes —  Attack of free radicals on <cell membrane  Increase in oxygen free radical  Decrease in cellular defense system  *adapted from Sjodin  etal[171]  45  <-  Training  and Adaptation  Effect  A training a n d adaptation r e s p o n s e h a s b e e n reported in the literature. S c h w a n e et al [145] s u g g e s t e d that training c o u l d reduce the magnitude of pathological alterations that o c c u r after eccentric e x e r c i s e . N e w h a m et al [131]  further  s u g g e s t e d a few p o s s i b l e explanation for this training effect: 1) there is a c h a n g e in the pattern of motor unit recruitment (i.e. either s u s c e p t i b l e fibres are s p a r e d o n s e c o n d a n d s u b s e q u e n t o c c a s i o n or more fibres are recruited a n d the forcefibre ratio is reduced), 2) there m a y be m u s c l e fibre adaptation s o that they b e c o m e more  resistant to the fatiguing  a n d d a m a g i n g effects of eccentric  e x e r c i s e a n d 3) the first bout of e x e r c i s e h a d c a u s e d d a m a g e a n d destruction to a population of susceptible fibres, possibly t h o s e near the e n d of their life c y c l e . T h e time between e x e r c i s e bouts would then be sufficient for regeneration thus m a k i n g available a fibre population of high m e c h a n i c a l resistance [126].  Byrnes  et al [172] a n d N o s a k a et al [173] found that the effect of training on reducing indicators of eccentric e x e r c i s e - i n d u c e d m u s c l e d a m a g e might be o b s e r v e d after only a single e x e r c i s e bout.  Similar results were a l s o evident s h o w i n g that  training a l s o r e d u c e s the i n c r e a s e in s e r u m C K a n d m y o g l o b i n , upon repeat bouts of activity [112, 172, 173]. B y r n e s et al [172] s u g g e s t that the m e c h a n i s m s responsible for s o r e n e s s a n d e n z y m e r e l e a s e are the s a m e but the magnitude of the c h a n g e between repeat e x e r c i s e s e s s i o n s c a n be altered by the performance on the first bout of work.  In addition, C l a r k s o n a n d c o l l e a g u e s [127] s h o w e d a  d e c r e a s e in r e s p o n s e to a s e c o n d bout of e x e r c i s e after a 9 w e e k separation b e t w e e n e x e r c i s e while B y r n e s et al [172] a n d N o s a k a et al [173] reported that this prophylactic effect on the generation of m u s c l e s o r e n e s s a n d s e r u m protein r e s p o n s e m a y last up to 10 w e e k s .  2.6 The Role of Magnetic Resonance Imaging in the Detection of DOMS D i s c o v e r e d in the  1940's, the p h e n o m e n o n of nuclear magnetic r e s o n a n c e  ( N M R ) h a s b e c o m e a valuable instrument for the non-invasive study of m u s c l e bioenergetics during e x e r c i s e [176]. N M R c a n be u s e d to p r o d u c e high quality images  using magnetic  r e s o n a n c e imaging (MRI)  a n d h a s b e e n useful  46  in  e x a m i n i n g healthy a n d d i s e a s e d skeletal m u s c l e  [174].  M R I , u s e d to study  protons, u s e s large v o l u m e radiofrequency coils a n d gradient magnetic fields that allow 2 D a n d 3 D i m a g e s to be constructed [174]. T h e intensity of the g e n e r a t e d i m a g e s d e p e n d s on the rate of relaxation of the protons in the s a m p l e a r e a . O b s e r v i n g different types of n u c l e u s relaxation, n a m e l y T 1 a n d T 2 relaxation, c a n generate i m a g e s of varying contrast [174].  T 2 - w e i g h t e d proton i m a g e s have b e e n particularly useful in detecting differences in the c h e m i c a l environment of cellular water, although T1 relaxation h a s a l s o b e e n investigated. E x e r c i s e c h a n g e s the c h e m i c a l properties of water m o l e c u l e s in m u s c l e (e.g. i n c r e a s e in total water content [extracellular fluid]). T h e resulting effect is longer T1 a n d T 2 relaxation times a n d brighter s i g n a l s within m u s c l e immediately following exercise [177-179]. Furthermore, the magnitude of t h e s e immediate p o s t - e x e r c i s e c h a n g e s h a v e b e e n s h o w n to be linearly related to e x e r c i s e intensity [177-179].  F l e c k e n s t e i n et al [179] found that after sports-  related m u s c l e injuries, d e l a y e d i n c r e a s e s in m u s c l e T 2 relaxation times a n d signal intensities were demonstrated a n d that this outlasted all other indicators of m u s c l e injury.  T h i s h a s also b e e n reported in n u m e r o u s other studies of  eccentric e x e r c i s e [180-183].  Nurenburg et al [182] has s u g g e s t e d that the sensitivity of M R I - g u i d e d biopsy for the detection of e x e r c i s e induced m u s c l e d a m a g e is by far more accurate in determining the extent a n d location of m u s c l e injury than b i o p s i e s guided by DOMS.  T h e s e investigators cite that there is a poor correlation between D O M S  a n d C K a n d the extent of ultrastructural m u s c l e injury but that there is a g o o d correlation  between  signal  intensity  grades  by  MRI  and  the  degree  of  ultrastructural d a m a g e . S e v e r a l studies have u s e d MRI to investigate c h a n g e s in c r o s s sectional a r e a of m u s c l e following a protocol of lengthening contractions [181, 183]. T h e s e studies demonstrate that the c r o s s sectional a r e a of m u s c l e i n c r e a s e s in a d e l a y e d m a n n e r with a time c o u r s e similar to the c h a n g e s in signal intensity a n d T 2 relaxation in m u s c l e .  47  Functional MRI refers to imaging not only the a n a t o m y of a tissue but a l s o the extent to which the t i s s u e is involved in performing s o m e task functional  magnetic  resonance  imaging  is  used  to  compare  [226]. M u s c l e the  relative  involvement of different m u s c l e s recruited during e x e r c i s e . T h i s method relies on the activity-induced i n c r e a s e in the nuclear magnetic transverse relaxation time (T2) of the m u s c l e water, which is c a u s e d by osmotically driven shifts of fluid into the myofibrillar s p a c e [226]. In addition to imaging of whole m u s c l e recruitment, m u s c l e MRI m a y reveal c h a n g e s in motor unit organization during d i s e a s e [226].  T h e availability a n d sophistication of magnetic r e s o n a n c e imaging s c a n n e r s h a v e i n c r e a s e d e n o r m o u s l y during the last d e c a d e a n d h a s now b e c o m e the method of c h o i c e for clinical imaging of most soft tissue pathologies, including sportsrelated injuries of m u s c l e a n d joints [226]. B e c a u s e of its noninvasive t e c h n i q u e a n d its i n d e p e n d e n c e from ionizing radiation, r e s e a r c h e r s now u s e this imaging technique for basic a n d applied morphometric studies of h u m a n subjects. F o r the p u r p o s e of our study, MRI c a n easily m e a s u r e the effects of e x e r c i s e training on m u s c l e a n d fat v o l u m e a n d the inflammation a s s o c i a t e d with d e l a y e d - o n s e t muscle soreness.  2.7 Perceived Muscle Soreness and the Visual Analog Scale P a i n is very subjective in nature, thereby m a k i n g it very difficult to objectively m e a s u r e a n d quantify. pain  T h e visual a n a l o g s c a l e is o n e m e a s u r e for quantifying  [155, 1 9 5 - 1 9 7 , 199, 201].  It c o n s i s t s of a 10 c m long line with m a r k e d  e n d i n g s indicating no pain at one e n d a n d extreme pain at the other e n d [ A P P E N D I X B].  T h i s line m a y be horizontal or vertical in nature with both  s h o w i n g a high correlation  (r=0.99)  [200].  In order to quantify a n d objectively  m e a s u r e p a i n , subjects are a s k e d to p l a c e a mark along this line with respect to their level of pain at the time of m e a s u r e m e n t .  T h i s distance is then m e a s u r e d  from o n e extreme of no pain to the a r e a m a r k e d .  Although a great d e a l of  criticism exists in the literature with respect to the high variability inherent in this pain rating s y s t e m , the visual a n a l o g s c a l e h a s b e e n found to bring greater  48  sensitivity a n d statistical power to data collection a n d a n a l y s i s by allowing a broader range of r e s p o n s e s than traditional  categorical r e s p o n s e s .  It a l s o  r e m o v e s the bias brought on by e x a m i n e r questioning a n d allows for graphical temporal c o m p a r i s o n s , thus minimizing bias a n d boosting statistical p o w e r  [155,  198].  49  C H A P T E R 3:  METHODOLOGY  3.1 Experimental design T h e experiment w a s divided into four s t a g e s :  •  STAGE  1  (baseline/pre-exercise)  Evaluation period - Height - Weight - Perceived muscle soreness (VAS) - Eccentric strength (torque) assessment - Quadricep circumference - Blood analysis - Creatine Kinase - Interleukin 6 - Malondialdehyde - Magnetic resonance imaging •  STAGE 2 (eccentric  exercise  protocol/DOMS)  Familiarization and warm-up period Exercise period (inducing muscle injury (DOMS)) •  STAGE  3 (treatment)  Treatment period (HBO therapy) •  STAGE  4 (post-exercise  evaluation  - days 2-5)  Evaluation period (post-exercise) - Height - Weight - Perceived muscle soreness (VAS) - Eccentric strength (torque) assessment - Quadricep circumference - Blood analysis - Creatine Kinase - Interleukin 6 - Malondialdehyde - Magnetic resonance imaging (days 1, 3 and 5)  50  1ft X  X  X  X  X  X  X  X  X  X  X  X  X  X  X  X  X  o _. °- 3  •? E Q oo —  X  </) Q  X  X  N "  >- a 3 < x: £  X  a4  UJ  <  ro a  X  X  CQ  CD  o 0) <  a5 Q p  z  a.*  UJ_j  £ 8 9.  Lu< CL UJ  CC <  Q >  CD  o -  w  cn'i= C  TD  CU CD O — 3 O 11 0- CO O 00  l  7 T' r  co  -2 CO c LU -I CL  <  a) c CL  2  Q.  g  UJ CD  O CD  3.2 Subjects S i x t e e n f e m a l e volunteers b e t w e e n the a g e s 18-40 participated in this study. S u b j e c t s w e r e recreationally active with no weight training, running, or t e a m sports a s part of their p h y s i c a l regimen. A n y previous e x p e r i e n c e with eccentric e x e r c i s e would h a v e c a u s e d adaptation of the m u s c l e s u c h that the m u s c l e w o u l d h a v e b e e n more resistant to the effects of s u b s e q u e n t bouts of intense e x e r c i s e . S u b j e c t s did not e n g a g e in physical activity for more than 3 hours a w e e k a n d h a d to have met both inclusion a n d e x c l u s i o n criteria to participate. E x c l u s i o n from participation in the study included athletes w h o actively weight train, run, jog, play in t e a m sports, and/or s k i . T h e s e activities involve repetitive eccentric loading of the q u a d r i c e p s a n d therefore the eccentric e x e r c i s e would not h a v e p r o d u c e d the desired effect of D O M S .  In addition, individuals who h a d  e x p e r i e n c e d d e l a y e d - o n s e t m u s c l e s o r e n e s s to their q u a d r i c e p s in the previous three months prior to participating or who h a d a past history of s e v e r e joint injury, arthritis or other chronic i l l n e s s e s were e x c l u d e d . S u b j e c t s taking a n a l g e s i c s or prescription drugs were a l s o e x c l u d e d .  Contraindications  to  hyperbaric  contraindications:  diabetes,  lung  oxygen cysts,  were  epilepsy,  also upper  assessed  (HBO  respiratory  tract  infections, pregnancy, fever). After recruitment to the study, the subjects who met the a b o v e criteria were then required to carefully read a n d sign a c o n s e n t form a n d fill out a questionnaire that w a s a p p r o v e d by the University of British C o l u m b i a C l i n i c a l Ethics C o m m i t t e e for r e s e a r c h involving h u m a n  subjects  ( A P P E N D I X A).  3.3 Procedure T h e r e s e a r c h protocol w a s a r a n d o m i z e d double-blind d e s i g n . Subjects were brought into the A l l a n M c G a v i n Sport M e d i c i n e Clinic where they w e r e s c r e e n e d a n d e x a m i n e d thoroughly to e n s u r e that all the inclusion a n d e x c l u s i o n criteria h a d b e e n met a n d that they were not at any risk by undergoing H B O treatment. T h e subjects were then randomly a s s i g n e d to one of two groups: a control (N=8)  52  a n d experimental group (N=8) a n d blinded to their specific treatment a n d group assignment.  E c c e n t r i c torque  (strength),  perceived  muscle  soreness  and  quadricep c i r c u m f e r e n c e m e a s u r e m e n t s w e r e taken at b a s e l i n e (Day 1) a n d after e a c h of the four treatment s e s s i o n s (i.e. D a y s 2-5). B l o o d s a m p l e s w e r e collected by antecubital venipuncture for the p u r p o s e of s e r u m creatine k i n a s e , interleukin6 a n d m a l o n d i a l d e h y d e a s s e s s m e n t at b a s e l i n e (Day 1), 4-hours p o s t - e x e r c i s e (Day 2) a n d e a c h day following treatment s e s s i o n s (Day 3, 4, 5).  Magnetic  r e s o n a n c e i m a g e s were collected at b a s e l i n e (Day 1), 2 4 - h o u r s p o s t - e x e r c i s e (Day 3) a n d 7 2 - h o u r s (Day 5) p o s t - e x e r c i s e .  STAGE  1  Initially subjects were given a brief explanation of the experimental p r o c e s s . T h e subjects were then evaluated p r e - e x e r c i s e for p e r c e i v e d m u s c l e s o r e n e s s , eccentric m u s c l e strength (torque) a n d quadricep c i r c u m f e r e n c e . B l o o d s a m p l e s were collected to a s s e s s levels for C K , IL-6 a n d M D A a n d a n MRI w a s taken of their quadricep m u s c l e s (both dominant a n d non-dominant).  T h e height a n d  weight of e a c h subject w a s a l s o recorded daily.  Perceived  muscle  soreness  Subjects w e r e requested to give a subjective rating of m u s c l e s o r e n e s s in their quadricep of the  non-dominant  leg. T h e testing o c c u r r e d p r e - e x e r c i s e a n d  immediately after e a c h hyperbaric e x p o s u r e on d a y s 2-5. T h e y w e r e instructed to complete four d e e p knee b e n d s a n d then rate the s o r e n e s s they e x p e r i e n c e d during t h e s e s q u a t s . T h e ratings w e r e c o m p l e t e d on the visual a n a l o g s c a l e ( V A S ) . T h e V A S is a 10cm line with "no pain or discomfort" the line a n d "worst pain or discomfort"  (e.g.  (e.g. 0) at one e n d of  10) at the other e n d . T h e subjects  were given a form a n d a s k e d to p l a c e a mark on the line a s to where they felt the level of p e r c e i v e d m u s c l e s o r e n e s s ( A P P E N D I X B).  T h e V A S w a s u s e d to  record the subjects' perception of s o r e n e s s of the quadricep on the e x e r c i s e d leg d u e to e x e r c i s e d - i n d u c e d injury a n d not s o r e n e s s e x p e r i e n c e d upon recollection of past injuries.  53  Eccentric  strength  Isokinetic strength w a s m e a s u r e d using the K i n C o m D y n a m o m e t e r . T h e K i n C o m D y n a m o m e t e r (Chattecx C o r p . , C h a t t a n o o g a , T e n n e s s e e ) is a  hydraulically  p o w e r e d , c o m p u t e r controlled exercise-testing d e v i c e . T h i s a p p a r a t u s w a s u s e d to m e a s u r e a n d record the eccentric torque (Newton -  metres, Nm) of the  q u a d r i c e p m u s c l e a s well a s create the e x e r c i s e - i n d u c e d m u s c l e s o r e n e s s ( D O M S ) in the subjects.  S u b j e c t s were first instructed to ride a stationary bike for 5 minutes followed by a stretching e x e r c i s e (lunges) before being s e a t e d on the e x e r c i s e equipment. T h e subjects were then instructed to sit on the isokinetic d y n a m o m e t e r , with their hips at 80°, their b a c k supported a n d their pelvis stabilized on the b e n c h with strapping  (fastening all three seatbelts (waist  belt, left s h o u l d e r a n d right  s h o u l d e r belt) to limit m o v e m e n t during the testing protocol). T h e r e s e a r c h e r set the lever a r m length of the K i n C o m d y n a m o m e t e r for e a c h subject to 7 5 % of the length from the h e a d of the fibula to the lateral malleolus with the lateral joint line of the k n e e in alignment with the center of the rotational axis point of the m a c h i n e . S e c u r i n g the test leg on the upper third of the quadricep a n d the lower leg by a tight V e l c r o strapped shin p a d e n s u r e d stabilization a n d limited the m o v e m e n t of the subjects' leg during the e x e r c i s e protocol. T h e y were further instructed to hold the s i d e s of the seat for additional stability. T h e angular velocity w a s set at 30° through a range of 60° at a long m u s c l e length (110 - 35° of k n e e flexion).  T h e subjects were then given a practice trial consisting of three s u b m a x i m a l a n d o n e m a x i m a l contraction (both concentric a n d eccentric contractions), followed by four m a x i m a l test contractions.  A 2-minute rest period o c c u r r e d b e t w e e n the  practice a n d test contractions, with the practice serving a s the w a r m - u p for e a c h test s e s s i o n . F o r the p u r p o s e of this study, only eccentric torque of the k n e e extensors w a s c o l l e c t e d . T h e baseline m e a n torque value w a s collected from the a v e r a g e of three m a x i m a l efforts of the four repetitions.  54  Quadricep  circumference  T h e m e a s u r e m e n t s of q u a d r i c e p s circumference w e r e taken before the eccentric exercise  protocol  and  after  each  treatment  sessions  anthropometric G u l i c k m e a s u r i n g tape (JUZO®).  using  a  standard  The s a m e Gulick measuring  tape w a s u s e d throughout the entire testing protocol a n d the s a m e s i d e of tape w a s u s e d consistently (i.e. the centimetre side of the tape would be on the superior side of the quadricep). E s t a b l i s h e d l a n d m a r k s were identified at the 10 a n d 2 0 c m point a b o v e the superior border of the patella, a s the subject lay s u p i n e on a n e x a m i n i n g table. A permanent, waterproof pen mark w a s p l a c e d at t h e s e two  points s o that the  investigator  c o u l d m e a s u r e the s a m e  points  throughout the testing period a n d e n s u r e s a c c u r a c y over the five d a y s . T h i s mark w a s reinforced on a daily b a s i s . A m e a n of two m e a s u r e m e n t s w a s obtained during every testing period. T h i s m e a s u r e m e n t w a s u s e d to evaluate c h a n g e s in the circumference of the quadricep m u s c l e , indicating the p r e s e n c e of e d e m a .  Blood  analysis  Thirty minutes after e a c h treatment s e s s i o n , subjects were driven to St. P a u l ' s Hospital ( S P H ) where twelve millilitres (mi's) of blood w a s withdrawn e a c h day, during  the  five  day testing  period. T r a i n e d  hospital  laboratory  technicians  collected blood s a m p l e s by standard antecubital venipuncture. S i x milliliters of blood w a s collected in an S S T (serum separator) tube for C K a n d M D A a n a l y s i s a n d the remaining six milliliters w a s collected in a n E D T A (ethylene diamine tetra-acetic acid) tube for IL-6 analysis. T h e blood s a m p l e s were then s p u n to isolate the p l a s m a , s e p a r a t e d in 1.5 ml cryotubes a n d frozen at - 7 0 ° C .  A n a l y s e s of all blood m e a s u r e m e n t s were c o n d u c t e d  at the  phlebotomory  laboratory at St. P a u l ' s Hospital a n d V a n c o u v e r Hospital & Health S c i e n c e s Centre.  The  levels for  each  parameter  (i.e.  CK/MDA/IL-6)  were  carefully  quantified a n d m e a s u r e d by highly qualified t e c h n i c i a n s in the laboratory a s well a s using state-of-the-art  measurement  d e v i c e s (e.g. instruments  measuring  reflection densities, f l u o r e s c e n c e , etc.).  55  Quality control w a s maintained for all blood a n a l y s e s in the study. T h i s allowed the  investigator to be certain that the a s s a y w a s reproducible a n d v a l u e s  obtained were consistent on a day-to-day b a s i s . Calibration of instruments u s e d in the laboratory is c o n d u c t e d every six m o n t h s or w h e n c h a n g e s o c c u r to the slide generator (reagent s l i d e s of the instrument). Adjustments to the instrument are m a d e internally by the c o m p u t e r operating s y s t e m . Internal Quality  Control  (IQC) a n d c h e c k s are carried out three times a d a y a n d are. s u b s e q u e n t l y monitored on a monthly b a s i s . Evaluations are sent out for c o m p a r i s o n with other laboratories worldwide to further e n s u r e reliability a n d validity of results. Quality  External  Control ( E Q C ) is monitored by CEQAL®' w h i c h is m a n d a t e d by the B C  Medical  Association,  Department  of  Diagnostic  Accreditation  Program.  S u b s c r i b e r s from C a n a d a a n d the United S t a t e s provide s a m p l e s every 2 months a n d evaluations are p r o c e s s e d with the results, including statistical a n a l y s e s , being  reported.  accreditation  Finally, the  laboratory  is fully a c c r e d i t e d , with a  period. T o e n s u r e reliability  a n d validity  of  five-year  M D A samples, a  standard curve w a s constructed prior to a s s a y a n a l y s e s . U s i n g a s t a n d a r d i z e d curve e n s u r e d g o o d reproducibility a n d precision of results on a daily b a s i s . All s a m p l e s were a n a l y z e d in a batch to further e n s u r e reliability of the results. In addition, every subject s e r v e d a s their own control to e x a m i n e c h a n g e s of malondialdehyde levels post-injury, therefore s h o w i n g lipid peroxidation c h a n g e s over the five-day treatment period.  Creatine  Kinase  A n a l y s i s of C K activity  in s e r u m  plasma was conducted  C h e m i s t r y Calibrator Kit 3™ a n d the Vitros C K Slide.  using the  Vitros  T h e Vitros C K Slide is a  dry, multilayered, analytical element c o a t e d on a polyester support. A n 11 uL drop of s a m p l e is deposited on the slide a n d evenly distributed by s p r e a d i n g the layer to the underlying layers. T h i s layer contains N-acetylcysteine ( N A C ) to activate C K without pretreating the s a m p l e . W h e n the s a m p l e is deposited on the slide, creatine kinase c a t a l y z e s the c o n v e r s i o n of creatine p h o s p h a t e a n d A D P to creatine  and  A T P . In  the  presence  of  glycerol  kinase  ( G K ) , glycerol  56  is  phosphorylated  to  L-a-glycerophosphate  by  ATP.  Oxidation  of  L-oc-  g l y c e r o p h o s p h a t e to d i h y d r o x y a c e t o n e p h o s p h a t e a n d h y d r o g e n peroxide o c c u r s in the p r e s e n c e of L - a - g l y c e r o p h o s p h a t e o x i d a s e (oc-GPO). Finally leuco d y e is o x i d i z e d by hydrogen peroxide in the p r e s e n c e of p e r o x i d a s e to form a d y e . Reflection densities are monitored during incubation a n d the rate of c h a n g e in reflection density is then c o n v e r t e d to C K e n z y m e activity.  Interleukin-6 S e r u m IL-6 w a s a n a l y z e d using a standard E L I S A kit obtained by  ChemiKine™.  T h i s kit is a s a n d w i c h e n z y m e i m m u n o a s s a y (EIA), which m e a s u r e s the "free" forms  of  the  cytokine  IL-6.  With  this  assay  system,  pre-coated  mouse  m o n o c l o n a l antibodies g e n e r a t e d against h u m a n IL-6 are u s e d to capture h u m a n IL-6 in a s a m p l e . S i m u l t a n e o u s l y , IL-6 specific rabbit polyclonal antibodies detect IL-6 in the s a m p l e . With the addition of goat anti-rabbit  conjugated-alkaline •  p h o s p h a t a s e (which binds to the rabbit anti-human polyclonal cytokine antibody), followed by the addition of the supplied color generating solution, the amount of IL-6 is detected.  Malondialdehyde  Finally, the a n a l y s i s of s e r u m p l a s m a for M D A levels w a s c o m p l e t e d using the s t a n d a r d T B A a s s a y a n d fluorimetric  analysis.  Tetramethoxy  propane  was  diluted with ethanol. A n aliquot (250 ul) with distilled water w a s treated with 1.5 ml of 2 0 % trichloroacetic a c i d ( T C A ) a n d then mixed with 1.5 ml of 0 . 6 7 % thiobarbituric acid solution. T h e mixture w a s h e a t e d for 3 0 minutes a n d then s p u n at 2 5 0 0 rpm for 10 minutes. Relative f l u o r e s c e n c e intensity of the  reaction  product w a s m e a s u r e d at 5 1 5 nm excitation a n d 5 5 3 nm e m i s s i o n .  Magnetic  resonance  imaging  All s c a n s were performed on a 1.5 t e s l a S i e m e n s S y m p h o n y MRI s y s t e m . T h e patients were positioned s u p i n e , a n d centered on the MR I s c a n n i n g table, with the legs a d d u c t e d .  C o r o n a l a n d axial scout i m a g e s were obtained for initial  57  localization. A x i a l i m a g e s were obtained from the level of the l e s s e r trochanter to the superior pole of the patella. T h e field of view included both thighs.  The  positioning a n d slice locations w e r e identical for all subjects a n d s c a n s . T h e body coil w a s u s e d for signal reception. T 2 relaxation time a n d Short Tip R e c o v e r y (STIR) i m a g e s were a s s e s s e d . S u r f a c e coils of the  Inversion  radiofrequehcy  s y s t e m (i.e. body array coil) for magnetic r e s o n a n c e imaging w e r e tested a n d rotated on a daily b a s i s to maintain quality control for this m e a s u r e m e n t .  T h e pulse s e q u e n c e s were axial T 2 , axial S T I R a n d c o r o n a l S T I R . T h e axial T 2 i m a g e s u s e d a T R 5 5 0 0 m s e c , T E 110 m s e c , e c h o train 1 1 , matrix 2 5 6 by 2 5 6 , 10 m m slice thickness, 5 m m slice g a p , o n e signal a v e r a g e d .  T h e axial S T I R  i m a g e s u s e d a T R 4 3 0 0 m s e c , T E 3 0 m s e c , T l 160 m s e c , e c h o train 18, matrix 2 5 6 by 2 5 6 , 10 m m slice t h i c k n e s s , 5 m m slice g a p , o n e signal a v e r a g e d .  The  coronal S T I R i m a g e s u s e d the s a m e parameters a s the axial S T I R i m a g e s , except that the slice t h i c k n e s s w a s 5 m m , with no interslice g a p .  T h e i m a g e s were a n a l y z e d on a P C - b a s e d computer workstation using e F i l m (eFilm 1.5.0 software version).  T h i s workstation software allows s i m u l t a n e o u s  viewing, windowing, a n d m e a s u r e m e n t s to be performed on multiple s l i c e s from multiple imaging s e r i e s . F o r m e a s u r e m e n t of m u s c l e signal intensity, the i m a g e s from the 24-hour post e x e r c i s e s c a n were visually inspected to identify a n y a r e a s of i n c r e a s e d m u s c l e signal (indicative of e d e m a ) .  T h e a r e a of m a x i m a l e d e m a  w a s identified. A n oval region of interest w a s manually traced a r o u n d the a r e a of m a x i m a l m u s c l e e d e m a . T h e region of interest w a s greater that 1 c m in a r e a , thus it represented at least 1 c c v o l u m e of m u s c l e tissue.  C a r e w a s taken to not  include n o n - m u s c l e t i s s u e s s u c h a s ligament, blood v e s s e l s , or fat in the region of interest. T h e corresponding a r e a w a s identified in the opposite (dominant) leg, a n d in the identical anatomic location of the left leg on the first a n d third MRI scans.  T h e signal intensity of the m u s c l e w a s r e c o r d e d .  T h e ratio of signal  intensity of the e x e r c i s e d (non-dominant)/signal intensity of the identical anatomic location in the dominant leg w a s calculated. T h e s a m e p r o c e s s w a s repeated for  58  all three m u s c l e s : rectus femoris m u s c l e , v a s t u s intermedius a n d v a s t u s lateralis m u s c l e . A subjective s c o r e b a s e d on visual a s s e s s m e n t w a s a l s o r e c o r d e d : 0 = no visible e d e m a , 1 = minimal m u s c l e e d e m a , 2 = moderate m u s c l e e d e m a , 3 = marked muscle edema.  STAGE  2  D e l a y e d - o n s e t m u s c l e s o r e n e s s w a s i n d u c e d on the K i n C o m D y n a m o m e t e r in the quadricep m u s c l e of the non-dominant leg. Subjects w e r e s e a t e d on the e x e r c i s e equipment a s outlined in s t a g e 1. T h e subjects w e r e given a w a r m - u p s e s s i o n to acquaint them with the eccentric e x e r c i s e .  This warm-up session  c o n s i s t e d of o n e set of 10 repetitions at a s u b m a x i m a l effort. After this w a r m - u p period, subjects were instructed to perform repeated eccentric contractions of their non-dominant leg (110° - 35° of k n e e flexion) at a s l o w s p e e d (30° per s e c o n d ) on the K i n C o m D y n a m o m e t e r . T h e subject were instructed not to resist the concentric m o v e m e n t on the w a y up but to resist the m a c h i n e ' s eccentric force on the w a y d o w n . voluntary  T h e e x e r c i s e required the completion of 3 0 0 m a x i m a l  eccentric contractions.  The  subjects  completed  30  s e t s of  10  repetitions with e a c h set beginning every minute for 30 minutes, allowing for a 15 second  rest between e a c h set.  T h e y received verbal f e e d b a c k from  the  r e s e a r c h e r (verbally e n c o u r a g e d to e n s u r e that they are giving m a x i m a l effort) a s well a s b i o f e e d b a c k from the resistive force or force v e r s u s velocity curve displayed on the K i n C o m monitoring unit.  STAGE  3  Immediately after the e x e r c i s e protocol a n d for the following 3 d a y s , subjects w e r e e x p o s e d to a hyperbaric environment (a total of 4 hyperbaric/normoxic e x p o s u r e s ) . Subjects were required to read a two-page information sheet on the hyperbaric unit a n d p r o c e d u r e s for c o m p r e s s i o n a n d d e c o m p r e s s i o n , prior to the first treatment s e s s i o n .  Subjects w e r e a l s o required to try the available aviator-  style g a s m a s k to e n s u r e a comfortable fit a n d to e n s u r e that a tight s e a l w a s maintained between the mouth a n d n o s e . T h i s m a s k w a s sterilized after e a c h  59  treatment s e s s i o n to m a k e sure that proper hygiene w a s maintained throughout the study period.  E m e r g e n c y equipment a n d clearing t e c h n i q u e s w e r e a l s o  e x p l a i n e d (i.e. swallowing, v a l s a l v a m a n o e u v r e , etc). Subjects w e r e  verbally  instructed a s to how they c o u l d d e c o m p r e s s the c h a m b e r on their own a n d let t h e m s e l v e s out if n e c e s s a r y . A m i c r o p h o n e in the c h a m b e r w a s continually o p e n a n d pointed out to the subjects to allow t h e m to c o m m u n i c a t e a n y p r o b l e m s that they m a y e x p e r i e n c e during c o m p r e s s i o n .  Control For  group  e a c h treatment  session,  the  subjects  were  seated  inside the  HYOX  m o n o p l a c e c h a m b e r ( A b e r d e e n , Scotland). T h e c h a m b e r w a s then c o m p r e s s e d to a p r e s s u r e of 1.2 A T A (an i n c r e a s e of 140 m m Hg). During c o m p r e s s i o n , the subjects breathed the ambient air in the c h a m b e r .  O n c e at a p r e s s u r e of 1.2  A T A , the subjects were instructed to w e a r the g a s m a s k , inspiring normoxic air ( 2 1 % oxygen).  T h e c h a m b e r w a s then r e d u c e d to barometric p r e s s u r e for the  remainder of the 60-minute treatment s e s s i o n . After 60 minutes, the subjects were instructed to remove the m a s k a n d again b e g a n breathing the ambient air of the c h a m b e r while d e c o m p r e s s i o n w a s initiated.  T h i s i n c r e a s e in p r e s s u r e to 1.2 A T A w a s sufficient to c a u s e the subjects to experience  the . c o m m o n  tympanic  membrane  sensations  associated  with  increasing ambient p r e s s u r e . Therefore, it w a s unlikely for subjects to determine their group designation. T h e air w a s delivered through a s e r i e s of regulators to the m a s k from high-pressure cylinders external to the c h a m b e r .  Experimental  group  T h e c o m p r e s s i o n a n d d e c o m p r e s s i o n procedure for the hyperbaric o x y g e n group w a s identical with the p r o c e d u r e s for the control group, with the exception that this group w a s c o m p r e s s e d to 2.0 A T A . O n c e at 2.0 A T A , the subjects in the experimental group received 1 0 0 % o x y g e n , which w a s delivered to the aviatorstyle m a s k from high-pressure cylinders external to the c h a m b e r .  60  STAGE  4  T h e s a m e evaluation w a s performed a s stage 1 after e a c h treatment s e s s i o n . A l l d e p e n d e n t v a r i a b l e s were m e a s u r e d immediately p o s t - H B O / n o r m o x i c treatment s e s s i o n s (days 2-5).  3.4 Statistical Analysis T h e study d e s i g n involved 2 groups (experimental a n d control group) with a n u m b e r of d e p e n d e n t variables (i.e. p e r c e i v e d s o r e n e s s , eccentric  strength,  quadricep c i r c u m f e r e n c e , blood e n z y m e s a n d M R I ) that w a s m e a s u r e d o n a n u m b e r of repeated o c c a s i o n s . A two w a y (group x time) A N O V A w a s performed with repeated m e a s u r e s on the d e p e n d e n t variables, with the exception of p e r c e i v e d m u s c l e pain ( V A S s c o r e s ) . V A S s c o r e s were nonparametric test  a n a l y z e d using a  (Friedman test). F o r all statistical a n a l y s e s , the significant  level w a s set at p<0.05.  Statistical a n a l y s e s w e r e performed using a n I B M -  compatible c o m p u t e r a n d S P S S 9.0 statistical software.  3.5 Statistical Power P o w e r for the study (0.76) w a s b a s e d o n calculations of C o h e n ' s D c (deita/s(i-r) ). 1/z  C a l c u l a t i o n s w e r e b a s e d on a n a l p h a level set at 0.05, a 2 0 % e x p e c t e d c h a n g e , correlations of 0.6 a n d a standard deviation of 3 3 % for the individual variables. T h e s e v a l u e s were determined from previous literature a n d a s s u m p t i o n s were b a s e d o n clinical significance.  61  CHAPTER 4: RESULTS Anthropometric  Data  T h e m e a n physical characteristics of the 16 subjects w h o c o m p l e t e d the study are listed in T a b l e 10. N o significant differences w e r e detected between subjects in their height, weight a n d a g e (p<0.05).  T a b l e 10: P h y s i c a l characteristics for both g r o u p s (age, height a n d weight). V a l u e s reported a s M e a n ± S D .  AGE (yr*)  HEIGHT (cm)  WEIGHT (kg)  Control (n=8)  25.25 ± 4 . 1 0  162.31 ± 6 . 0 3  57.0 ± 12.8  Experimental (n=8)  25.49 ± 4.24  160.90 ± 3 . 2 7  58.5 ± 10.1  Note: no significant differences between groups on any of the variables  Muscle  Pain  Results for the rating of p e r c e i v e d s o r e n e s s of the non-dominant  leg a r e  illustrated in Figure 5. All groups s h o w e d a significant i n c r e a s e in s o r e n e s s after the e x e r c i s e protocol (p<0.05, p=0.0001) but there w a s n o statistical difference between groups for treatment effects (p=0.571).  A significant interaction effect  between treatment a n d time w a s evident (p<0.05, p=0.010).  T h e control group s h o w e d a n i n c r e a s e in pain from b a s e l i n e , at 4 hours poste x e r c i s e , p e a k i n g at 2 4 hours a n d 4 8 hours p o s t - e x e r c i s e a n d d e c r e a s i n g 7 2 hours post-insult ( A P P E N D I X C - Figure A ) . T h e experimental group h a d pain levels i n c r e a s e from b a s e l i n e , 4 hours p o s t - e x e r c i s e , p e a k i n g at 2 4 hours a n d d e c r e a s i n g 4 8 hours a n d further d e c r e a s i n g 7 2 hours post-exercise ( A P P E N D I X C - Figure B) [Table 11]. Both g r o u p s e x p e r i e n c e d l e s s pain over time e v e n though the p e a k time period for pain differed b e t w e e n groups.  62  T a b l e 11: A v e r a g e ratings of p e r c e i v e d s o r e n e s s for the quadricep m u s c l e of the non-dominant leg before (baseline) and after (days 2, 3, 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s visual a n a l o g s c o r e (1-10) ± S E M .  Baseline (Day 1)  4-Hours PostExercise (Day 2)  24 Hours PostExercise (Day 3)  48 Hours PostExercise (Day 4)  72 Hours PostExercise (Day 5)  Control (n=8)  0.00 ± 0.00  2.38 ± 0.57  3.25 ± 0.45  3.38 ± 0.71  1.88 ± 0 . 5 2  Experimental (n=8)  0.00 ± 0.00  2.75 ± 0.65  4.38 ± 0.57  2.12 ± 0 . 4 4  0.25 ± 0 . 1 6  U « A w l  V> 1 U v./  Figure 5: Average rating of perceived soreness for the quadricep muscle of the non-dominant leg, according to the visual analog scale (range before (baseline) and after hyperbaric/normoxic exposure.  1-10),  •  Control Group  Hi—Experimental Group  5.0  4.0  3.0  2.0  LO H Baseline  4 Hours Post-Ex  24 Hours Post-Ex  48 Hours Post-Ex  72 Hours Post-Ex  Day(s)  63  Eccentric  Strength  A n a l y s i s of the m e a n eccentric torque indicated that there w e r e differences within the g r o u p s  (p<0.05, p=0.0001);  significant  however, there  statistical difference between g r o u p s a n d there w a s n o significant  w a s no interaction  effect (p<0.05, p=0.102; p=0.100) (Figure 6).  T h e control group s h o w e d eccentric torque to b e significantly lower 2 4 hours post-exercise,  with  gradual  recovery  by d a y 5 (72 hours  post-exercise)  ( A P P E N D I X C - Figure C ) . T h e experimental group d e m o n s t r a t e d a d e c r e m e n t in eccentric torque at 4 hours p o s t - e x e r c i s e , followed by gradual recovery 2 4 , 4 8 a n d 7 2 hours p o s t - e x e r c i s e ( A P P E N D I X C - Figure D) [Table 12]. Both groups d e m o n s t r a t e d immediate eccentric strength d e c r e m e n t s , followed by a gradual pattern of recovery.  Table 12: A v e r a g e  m a x i m a l eccentric torque for the q u a d r i c e p m u s c l e before (baseline) a n d after (days 2 , 3 , 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . Strength v a l u e s reported a s m a x i m a l torque (Nm) ±SEM.  Baseline (Day 11  4-Hours Post- 24 Hours Exercise :;Rpst-Exercise (Dry 2) (Day 3)  48 Hours Post-Exercise (Day 4)  72. Hours PostExercise f Day 5)  Control (n=8)  136.0 + 13.75  91.0 + 9.02  87.25 + 9.92  95.38 ± 10.13  98.38 ± 1 1 . 1 0  Experimental (n=8)  142.38+7.44  109.50 ± 7 . 9 5  113.0 ± 7 . 8 2  125.38 ± 9 . 5 6  125.63 ± 8 . 4 9  64  Figure 6: Average maximal eccentric torque for the quadricep muscle, before the exercise protocol (baseline) and after hyperbaric/normoxic exposure.  Quadricep  Circumference  A n a l y s i s of the quadricep c i r c u m f e r e n c e at the 10cm reference point a b o v e the superior portion  of the  patella indicated that quadricep circumference w a s  significantly different within the groups (p<0.05, p=0.005); however, there w a s no significant  difference  between  (p<0.05, p=0.815; p=0.939)  groups  (Figure 7).  a n d there  was  no  interaction  effect  At the 2 0 c m point a b o v e the knee,  quadricep circumference w a s not significantly different within the groups (p<0.05, p=0.253) a n d again there w a s no significant difference between groups.  Also,  there w a s no interaction effect (p<0.05, p=0.677, p=0.676) (Figure 8).  Both groups (control  a n d experimental), for the  10cm m e a s u r e m e n t  point,  demonstrated a slight i n c r e a s e in e d e m a at 4 hours p o s t - e x e r c i s e , p e a k i n g at day  65  3 (24 hours post-exercise), d e c r e a s i n g o n d a y 4 (48 hours) a n d finally s h o w i n g a further d e c r e a s e o n d a y 5 (72 hours) post-exercise ( A P P E N D I X C - Figure E , F ) .  T h e 2 0 c m m e a s u r e m e n t for both groups w a s slightly more variable. T h e control group h a d a slight d e c r e a s e 4 hours post-exercise, increasing 2 4 a n d 4 8 hours post-insult a n d further d e c r e a s i n g 7 2 hours post-exercise ( A P P E N D I X C - Figure G ) . T h e experimental group s h o w e d a slight i n c r e a s e in c i r c u m f e r e n c e 4 hours post-exercise, d e c r e a s i n g below b a s e l i n e 2 4 hours post-insult, increasing 4 8 hours a n d finally d e c r e a s i n g 7 2 hours post-exercise ( A P P E N D I X C - Figure H) [Table 13]. i  T a b l e 1 3 : A v e r a g e quadricep c i r c u m f e r e n c e m e a s u r e d at both the 10 a n d 2 0 c m point a b o v e the superior portion of the patella. M e a s u r e m e n t s were taken before (baseline) a n d after (days 2, 3 , 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . Q u a d r i c e p circumference reported a s centimetres (cm)) ± S E M .  Baseline (Day 1)  4-Hours PostExorciso (Day 2)  24 Hours PostExercise (Day 3)  48 Hours PostExercise (Day 4)  72 Hours PostExe-cise (Day 5)  Control (n=8)  44.99 ± 1.93  45.10 ± 1.78  45.31 ± 1 . 7 1  44.82 ± 1.91  44:13 ± 1.86  Experimental (n=8)  45.42 ± 2.04  45.73 ± 1.99  45.96 ± 2 . 1 8  45.71 ± 2.05  44.79 ± 1 . 9 9  48 Hours ,_ " Exorcifn (Day 4)  72 Hours PostExercise (Day 5)  10 cm  Baseline (Day 1) on ^ 20 cm  . 4-Hours ,_ Exercise (Day 2i P o s t  24 Hours P'wt,_ Exorcise (Day 3)  :  P o s t  Control (n=8)  53.49 ± 1 . 9 6  52.97 ± 1.99  53.19 ± 1.99  53.21 ± 2.01  52.89 ± 2.01  Experimental (n=8)  54.49 ± 2 . 1 7  54.64 ± 2 . 2 8  54.38 ± 2.38  54.68 ± 2.28  53.91 ± 2 . 1 5  66  Figure 7: Average quadricep circumference (10 cm location), before eccentric exercise (baseline) and after hyperbaric/normoxic exposure.  42.0 -I  -•—Control Group - • — Experimental Group  41.0 40.0 39.0 Baseline  4 Hours Post-Ex  24 Hours Post-Ex Day(s)  48 Hours Post-Ex  72 Hours Post-Ex  F i g u r e 8: A v e r a g e q u a d r i c e p c i r c u m f e r e n c e (20 c m l o c a t i o n ) , b e f o r e e c c e n t r i c e x e r c i s e ( b a s e l i n e ) a n d after h y p e r b a r i c / n o r m o x i c exposure.  i 1|  a  » - — - - - — _ _ _ _ _ _  —i I  •  "  1  •Control Group -Experimental Group  Baseline  4 Hours Post-Ex  24 Hours Post-Ex  48 Hours Post-Ex  72 Hours Post-Ex  Day(s)  67  Blood Enzymes Creatine  Kinase  (CK)  A n a l y s i s of the creatine k i n a s e ( C K ) m e a s u r e m e n t s indicated that C K w a s not significantly  different  within  the  two  study  groups  (p<0.05,  p=0.538).  No  significant difference w a s d e m o n s t r a t e d b e t w e e n g r o u p s a n d there w a s  no  significant interaction effect (p<0.05, p=0.647; p=0.570) (Figure 9).  T h e control group d e m o n s t r a t e d an i n c r e a s e in C K levels 4 hours p o s t - e x e r c i s e , followed by a d e c r e a s e at 2 4 hours a n d 4 8 hours p o s t - e x e r c i s e , a n d a n i n c r e a s e at 72 hours post-insult ( A P P E N D I X C -  Figure I). T h e experimental  group  s h o w e d a m a r k e d d e c r e a s e from b a s e l i n e at 4, 2 4 , 4 8 a n d 7 2 hours post e x e r c i s e ( A P P E N D I X C - Figure J) [Table 14]. O n e subject h a d an unusually high C K r e s p o n s e b e f o r ^ t h e eccentric e x e r c i s e protocol (>2500 uL). C K a n a l y s i s w a s performed with a n d without this subject to e x a m i n e whether the  high  variability introduced into the data by this subject w a s m a s k i n g any significant statistical findings. R e m o v i n g this subject from the statistical a n a l y s i s did not alter the overall findings. Furthermore, removing this outlier from the data revealed a more stable curve, s h o w i n g an i n c r e a s e from b a s e l i n e at 4 a n d 2 4 hours poste x e r c i s e , followed by a gradual d e c r e a s e at 4 8 a n d 7 2 hours post-injury [Table 14].  T a b l e 14: M e a n s e r u m creatine kinase v a l u e s before (baseline) a n d after (days 2, 3, 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . C K v a l u e s reported a s microlitres (u./L) ± S E M . Baseline (Day1)  4-Hours PostExercise (Day 2)  24 Hours PostExercise (Day 3)  48 Hours PostExercise (Day 4)  72 Hours PostExercise (Day 5)  Control (n=8)  150.50 ± 69.20  208.13 ±76.76  178.58 ±39.85  139.13 ±32.22  168.13 ±56.93  Experimental (n=8)  396.50 ±316.42  271.63 ± 147.45  192.63 ±52.75  143.88 ±25.89  124.50 ± 2 2 . 3 8  125.43 ±22.11  145.30 ± 2 6 . 8 7  120.57 ± 13.02  110.43 ± 2 0 . 1 0  Experimental (minus outlier) 80.14 ± 7 . 2 7  68  Figure 9: Average creatine kinase (CK) levels, before (baseline) and after hyperbaric/normoxic exposure. 800.0  •Control Group  700.0  - Experimental Group 600.0  500.0  400.0  300.0  200.0 i  100.0  Baseline  4 Hours Post-Ex  24 Hours Post-Ex  48 Hours Post-Ex  72 Hours Post-Ex  Day(s)  Malondialdehyde (MDA) M a l o n d i a l d e h y d e ( M D A ) results indicate no significant difference within the two study groups (p<0.05,  p=0.08).  N o significant difference w a s demonstrated  between groups for treatment effects a n d there were no significant interaction effects (p<0.05, p=0.58; p=0.56) (Figure 10).  T h e control group d e m o n s t r a t e d very little variability between baseline v a l u e s a n d post-exercise levels, increasing slightly d a y s 3 a n d 4 a n d d e c r e a s i n g c l o s e to b a s e l i n e by day 5 ( A P P E N D I X C - Figure K). T h e experimental group, however, i n c r e a s e d d a y 2, d e c r e a s e d slightly day 3, i n c r e a s e d again on day 4 a n d slightly d e c r e a s e d by d a y 5. T h e v a l u e s for the experimental group remained slightly higher than what w a s o b s e r v e d at baseline ( A P P E N D I X C - Figure L) [Table 15].  69  T a b l e 1 5 : M e a n m a l o n d i a l d e h y d e v a l u e s before (baseline) a n d after (days 2 , 3 , 4, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . M D A v a l u e s reported a s n m o l e s M D A / m L ± S E M .  Control (n=8) Experimental (n=8)  Baseline (Day1)  4-Hours PostExercise (Day 2)  24 Hours PostExercise (Day 3)  48 Hours PostExercise (Day 4)  72 Hours PostExercise (Day 5)  3.98 ± 0.48  4.0 ± 0.34  4.28 ± 0.35  4.34 ± 0.58  4.03 ± 0.36  4.26 ± 0.38  4.55 ± 0.47  4.46 ± 0.52  4.58 ± 0.43  4.46 ± 0.44  Figure 10: Average malondialdehyde (MDA) levels, before (baseline) and after hyperbaric/normoxic exposure .  Normal levels 3.9-4.2 nmol MDA/mL •  Control  - » — Experimental  Baseline  4 Hous Post-Ex  24 Hours Post-Ex  48 Hours Post-Ex  72 Hours Post-Ex  Day(s)  70  Interleukin  6 (IL-6)  Interleukin 6 (IL-6) results indicate no significant difference within the two groups (p<0.05, p=0.400). N o significant difference w a s demonstrated b e t w e e n groups for treatment  effects a n d there w a s n o significant interaction effect  (p<0.05,  p=0.111; p=0.451) (Figure 11).  Both groups demonstrated m a r k e d differences in the pattern of recovery from e x e r c i s e - i n d u c e d m u s c l e injury a n d there w a s a great deal of variability. T h e control group fluctuated  markedly; d e c r e a s i n g from  b a s e l i n e , 4 hours  post-  e x e r c i s e , increasing a b o v e b a s e l i n e 2 4 hours post-exercise, further d e c r e a s i n g at 4 8 hours to levels similarly to those s e e n at 4 hours post-exercise a n d i n c r e a s i n g , although below b a s e l i n e , by 7 2 hours post-insult ( A P P E N D I X C - Figure M). T h e experimental group w a s more stable with l e s s variability a n d fluctuations in IL-6 levels. F r o m b a s e l i n e , the IL-6 levels slightly d e c r e a s e d 4 hours p o s t - e x e r c i s e , remained at the s a m e level 2 4 hours post-exercise, d e c r e a s e d further by 4 8 hours  a n d increased  above  baseline  levels  by 7 2 hours  post-exercise  ( A P P E N D I X C - Figure N) [Table 16].  Table 16: M e a n interleukin-6 v a l u e s before (baseline) a n d after (days 2, 3, 4 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . IL-6 v a l u e s reported a s p g / m L ± S E M .  Baseline .Day 1)  4-Hours PostExercise (Day 2)  24 Hours PostExercise (Day 3)  48 Hours PostExercise (Day 4)  Control (n=4)  618.13 ±517.87  Experimental (n=4)  59.73 ±21.98  72 Hours PostExercise (Day 5)  67.11 ± 5 7 . 8 3  873.19 ± 519.41  24.54 ± 11.71  459.26 ± 405.81  44.80 ± 14.33  46.98 ± 20.94  26.08 ± 5.75  86.23 ±21.03  71  Figure 11: Average interleukin-6 (IL-6) levels, before eccentric exercise (baseline) and after hyperbaric/normoxic exposure.  1600.0  i  1400.0  -•—Control Group HH— Experimental Group  1200.0  Normal levels <50 pg/ml 1000.0  800.0  600.0  i  400.0  200.0  .0  i  Baseline  4 Hours Post-Ex  2 4 Hours Post-Ex  4 8 Hours Post-Ex  7 2 Hours Post-Ex  Day(s)  Magnetic Resonance  Imagine (MRI)  T2-weighted images Rectus  Femoris  A n a l y s i s of the rectus femoris m u s c l e using T 2 weighted imaging revealed that this  m u s c l e w a s not significantly different  p=0.108).  N o significant  difference  within  the two groups  w a s demonstrated between  (p<0.05,  groups for  treatment effects a n d there w a s no significant interaction effect (p<0.05, p=0.800; p=0.799) (Figure 12).  T h e control group demonstrated very little variability between b a s e l i n e ratios a n d post-exercise ratios, revealing a slight i n c r e a s e by d a y 5 ( A P P E N D I X C - Figure O). T h e experimental group also s h o w e d very little variability b e t w e e n d a y s 1  72  through 5, h o w e v e r a slight i n c r e a s e w a s noted on day 3, followed by another i n c r e a s e on d a y 5 ( A P P E N D I X C - Figure P) [Table 17].  Table 17: A v e r a g e T 2 relaxation times of the rectus femoris m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s T 2 Relaxation T i m e (msec) ± SEM.  Day 1 (Baseline)  Day 3 (24 hours post-exercise)  Day 5 (72 hours post exercise) 1.03 ± 0 . 1 0  Control Group (n=8)  0.91 ± 7 . 0 2 x ^0~'  ^  0.92 ± 4.34 x 10"  Experimental Group (n=8)  0.89 ± 6.13 x 10"  2  0.97 ± 0 . 1 0  2  1.09 ± 0 . 1 9  Figure 12: Mean T2 relaxation times (msec) before (baseline) and after hyperbaric/normoxic exposure (days 3, 5) for the rectus femoris muscle 1.200  1.000  o a> to  .800  E <D  E  c o ra x ro  - • — Experimental Group .600  HI—Control Group  a  DC  .400  .200  .000 Baseline  24 Hours Post-Ex Day(s)  72 Hours Post-Ex  73  Vastus  Intermedius  A n a l y s i s of the v a s t u s intermedius m u s c l e using T 2 weighted imaging revealed that this m u s c l e w a s significantly different within the two study g r o u p s (p<0.05, p=0.007) but no significant difference w a s evident b e t w e e n g r o u p s for treatment effects a n d group interaction effects (p<0.05, p=0.361; p=0.259) (Figure13).  T h e control group d e m o n s t r a t e d a n i n c r e a s e in signal intensity 2 4 hours poste x e r c i s e , followed by a further i n c r e a s e at 7 2 hours post-insult ( A P P E N D I X C Figure Q ) . T h e experimental group a l s o s h o w e d a n i n c r e a s e on d a y 3, but on d a y 5, a d e c r e a s e w a s noted ( A P P E N D I X C - Figure R) [Table 18].  T a b l e 18: A v e r a g e T 2 relaxation times of the v a s t u s intermedius m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s T 2 Relaxation T i m e (msec) ± SEM. Day 1 (Baseline)  Day 3 (24 hours post-exercise)  Day 5 (72 hours post exercise)  Control Group (n=8)  0.95 ± 3.66 x 1 0 "  1.26±9.16x10  1.36 ± 0 . 1 5  Experimental Group (n=8)  0.96 ± 4.85 x 1 0 "  1.25 ± 0 . 1 3  1.104 ± 5 . 9 1 x 10"  2  74  2  Figure 13: Mean T2 relaxation times (msec) before (baseline) and after hyperbaric/normoxic exposure (days 3, 5) for the vastus intermedius muscle. 1.600  1.400 1  1.200  fi  1  1.000  c o  .800  fi E  - • — Experimental Group -•—Control Group  a a  .600  CM  .400  .200  .000 4Baseline  24 Hours Post-Ex  72 Hours Post-Ex  Day(s)  Vastus Lateralis A n a l y s i s of the v a s t u s lateralis m u s c l e using T 2 weighted imaging revealed that there w a s a significant difference within the groups for this m u s c l e  (p<0.05,  p=0.038) but no significant difference w a s evident between groups for treatment effects a n d group interaction effects (p<0.05, p=0.806; p=0.258) (Figure 14).  T h e control group d e m o n s t r a t e d an i n c r e a s e in signal intensity 2 4 hours postexercise, followed by a slight d e c r e a s e at 7 2 hours post-insult ( A P P E N D I X C Figure S ) . T h e experimental group also s h o w e d an i n c r e a s e 2 4 hours post-injury, followed by a further slight i n c r e a s e 72 hours post-insult ( A P P E N D I X C - Figure T) [Table 19].  75  T a b l e 1 9 : A v e r a g e T 2 relaxation times of the v a s t u s lateralis m u s c l e taken before (baseline) a n d after (days 3 , 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s T 2 Relaxation T i m e (msec) ± SEM  Day 1 (Baseline)  Control Group (n=8)  0.88 ± 8.65 x 10~  Experimental Group (n=8)  0.98 ± 7.75 x 10'  2  2  Day 3 (24 hours post-exercise)  Day 5 (72 hours post exercise)  1 . 1 0 ± 7 . 1 2 x 10"  2  1.05 ± 7.54 x 10"  2  1.02 ± 7.55 x 10"  2  1.09 ± 8.59 x 10"  2  Figure 14: Mean T2 relaxation times (msec) before (baseline) and after hyperbaric/normoxic exposure (days 3, 5) for the vastus lateralis muscle.  76  STIR IMAGES Rectus  Femoris  A n a l y s i s of the rectus femoris m u s c l e using S T I R imaging demonstrated that there w a s a significant difference within the groups (p<0.05, p=0.018) but no significant difference w a s evident between g r o u p s for treatment  effects  and  interaction effects (p<0.05, p=0.796; p=0.733) (Figure 15).  Both control a n d experimental g r o u p s demonstrated a similar pattern over the 3 m e a s u r e m e n t s taken over 5 d a y s . T h e control group d e m o n s t r a t e d an i n c r e a s e 2 4 a n d 7 2 hours p o s t - e x e r c i s e from baseline m e a s u r e m e n t s ( A P P E N D I X C Figure U). T h e experimental group a l s o s h o w e d an i n c r e a s e 24 hours post-injury, followed by a further slight i n c r e a s e 7 2 hours post-insult ( A P P E N D I X C - Figure V) [Table 20].  Table 20: A v e r a g e signal intensity ratio for S T I R image of the rectus femoris m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s signal intensity ratio - S T I R (msec) ± S E M .  ' Day 1 (Baseline) Control Group (n=8)  0.88 ± 5.95 x 10"  Day 3 (24 hours posti exercise) 1.00 ± 6.26 x 10"  Experimental Group (n=8)  0.91 ±2.01 x 10"  0.98 ±3.81 x 10"  2  2  2  2  Day 5 (72 hours post exercise) 1.07±6.94x 10"  J  1.02 ± 9.06 x 10"  2  /  77  Figure 15: Mean Signal Intensity Ratio for STIR image before (baseline) and after hyperbaric/normoxic exposure (days 3, 5) for the rectus femoris muscle.  Vastus  Intermedius  A n a l y s i s of the v a s t u s intermedius m u s c l e using S T I R imaging demonstrated that there w a s a significant difference within the groups (p<0.05, p=0.014) but no significant difference w a s evident for b e t w e e n groups for treatment effects a n d group interaction effects (p<0.05, p=0.580; p=0.451) (Figure 16).  T h e control group demonstrated a n i n c r e a s e in signal intensity 2 4 hours poste x e r c i s e , followed by a slight i n c r e a s e at 7 2 hours post-insult ( A P P E N D I X C Figure W ) . T h e experimental group a l s o s h o w e d a n i n c r e a s e 2 4 hours poste x e r c i s e , followed by a slight d e c r e a s e 7 2 hours after injury ( A P P E N D I X C Figure X) [Table 21].  78  Table 21: Average signal intensity ratio for STIR image of the vastus intermedius muscle taken before (baseline) and after (days 3, 5) the eccentric exercise protocol, following hyperbaric/normoxic exposure. Values reported as signal intensity ratio - STIR (msec) ± S E M .  Day 1 (Baseline)  Day 3 (24 hours post-exercise)  Day 5 (72 hours post exercise)  Control Group (n=8)  0.97 ± 2.96 x 10"  2  1.07 ± 4.99 x 10"  2  1.07 ± 5.87 x 10"  2  Experimental Group (n=8)  0.97 ± 1.98 x 1 0 "  2  1.06 ± 5.73 x 1 0 "  2  1.00 ± 2.78 x 10"  2  Figure 16: Mean Signal Intensity Ratio for STIR image before (baseline) and after hyperbaric/normoxic exposure (days 3, 5) for the vastus intermedius muscle. 1.150  .800  1  , Baseline  24 Hours Post-Ex  72 Hours Post-Ex  Day(s)  79  Vastus  Lateralis  A n a l y s i s of the v a s t u s lateralis m u s c l e using S T I R imaging  demonstrated  statistical significance within the groups (p<0.05, p=0.004) but no significant difference  was  evident  between  groups  for  treatment  effects  and  group  interaction effects (p<0.05, p=0.265; p=0.560) (Figure 17).  T h e control group demonstrated an i n c r e a s e in signal intensity 2 4 hours poste x e r c i s e , followed by a very slight d e c r e a s e at 72 hours post-insult ( A P P E N D I X C - Figure Y ) . T h e experimental group a l s o s h o w e d a slight i n c r e a s e 24 hours post-exercise, followed by a further minimal i n c r e a s e 72 hours after  injury  ( A P P E N D I X C - Figure Z) [Table 22].  T a b l e 22: A v e r a g e signal intensity ratio for S T I R image of the v a s t u s lateralis m u s c l e taken before (baseline) a n d after (days 3, 5) the eccentric e x e r c i s e protocol, following hyperbaric/normoxic e x p o s u r e . V a l u e s reported a s signal intensity ratio - S T I R (msec) ± S E M .  Day 1 (Baseline)  Control Group (n=8)  0.88 ± 5.99 x 10"  Experimental Group (n=8)  i . 0 0 ± 3 . 6 3 x 10"  2  2  Day 3 (24 hours postexercise)  Day 5 (72 hours post exercise)  1.07 ± 6.54 x 10 -  1.06 ± 5.36 x 1 0 "  2  1.09 ± 4 . 7 1 x 1 0 "  1.11 ± 5 . 5 6 x 10"  2  2  I  80  F i g u r e 1 7 : M e a n S h o r t T i p I n v e r s i o n R e c o v e r y (STIR) R a t i o s b e f o r e ( b a s e l i n e ) a n d after h y p e r b a r i c / n o r m o x i c e x p o s u r e ( d a y s 3, 5) f o r the v a s t u s l a t e r a l i s m u s c l e . 1.200 -r  1.000 |  .800 I  -•—Experimental Group  .600  -•—Control Group  .400  .200  .000 Baseline  24 Hour Post-Ex  72 Hours Post-Ex  Day(s)  81  CHAPTER 5: DISCUSSION T h e u s e of hyperbaric o x y g e n a s a therapeutic modality is increasing a m o n g athletes, trainers, physiotherapists a n d other m e d i c a l professionals. H o w e v e r , there is a paucity of scientific e v i d e n c e to prove the effectiveness of using hyperbaric o x y g e n a s a m e a n s of treatment. P r e v i o u s studies examining the u s e of hyperbaric o x y g e n to ameliorate e x e r c i s e - i n d u c e d D O M S did not convincingly prove or refute the efficacy of hyperbaric o x y g e n in treating m u s c l e a n d softtissue injuries.  T h e aim of the present study w a s to investigate the influence of  hyperbaric o x y g e n therapy on p e r c e i v e d m u s c l e s o r e n e s s , m u s c l e strength, muscle edema and plasma enzymes.  T h e injury m o d e l that w a s u s e d in this study, d e l a y e d - o n s e t m u s c l e s o r e n e s s , allowed us to induce a quantifiable m u s c l e injury a n d then monitor s e v e r a l variables over the c o u r s e of recovery. T h i s "gold-standard" injury model h a s b e e n investigated in great detail for over 15 y e a r s a n d provides a g o o d representation of m u s c l e injury, d a m a g e a n d c o m p r o m i s e d force that m a y o c c u r during sport and  recreational activities  [101-10,  105-106,  1 1 0 - 1 1 3 , 127, 131].  However,  caution should be applied w h e n using this type of soft tissue injury m o d e l . It h a s b e e n s u g g e s t e d by previous investigators [96, 2 4 6 , 247] that although  DOMS  w a s created in the m u s c l e of the subjects, the d e g r e e of m u s c l e s o r e n e s s m a y be at question, thus negating any beneficial effect the treatment modality m a y h a v e in the rehabilitation p r o c e s s . T h i s study elongated the m u s c l e length of the quadricep to 110°-35° to further e n s u r e high tension to the m u s c l e fibres. In addition, visual estimation for the p r e s e n c e of e d e m a w a s m a d e 2 4 hours postinjury through magnetic r e s o n a n c e imaging. H o w e v e r , although every effort w a s m a d e to e n s u r e that m u s c l e d a m a g e o c c u r r e d to the q u a d r i c e p , the d e g r e e of d a m a g e for this study is a l s o questionable.  1  F e m a l e s were u s e d in the study to m a k e the group a s h o m o g e n e o u s a s p o s s i b l e a n d to control for extraneous variables. Furthermore, s e x differences in m u s c l e fatigue have b e e n reported in the literature, with f e m a l e s generally exhibiting a  82  greater relative fatigue resistance than m a l e s [252-253].  This phenomenon has  b e e n o b s e r v e d in a variety of m u s c l e s with the u s e of various fatigue protocols. H i c k s et al [254] h y p o t h e s i z e d that m u s c l e m a s s , substrate utilization a n d m u s c l e morphology m a y be m e c h a n i s m s involved attributing to differences in fatigue resistance. E s t r o g e n m a y a l s o be implicated in modulating m u s c l e fatigue. E s t r o g e n receptors are found on v a s c u l a r endothelial a n d s m o o t h m u s c l e cells [255].  E s t r o g e n s affect  v a s c u l a r tone  indirectly  by modulating  release of  endothelium-derived v a s o a c t i v e factors a n d directly by modulating intracellular calcium  in v a s c u l a r s m o o t h  thrombotic  events  and  m u s c l e cells [255]. E s t r o g e n s indirectly  inflammation  (  by  altering  platelet  affect  aggregation  and  leukocyte a d h e r e n c e a n d migration, respectively [255]. E s t r o g e n s a l s o influence production of mitogens w h i c h , w h e n r e l e a s e d at sites of v a s c u l a r injury, affect v a s c u l a r remodeling [255]. Therefore, o n e g e n d e r w a s s e l e c t e d to control for g e n d e r differences a s s o c i a t e d with fatigue resistance.  T h e study, in its entirety, w a s c o m p l e t e d with 16 female participants. Although it w a s anticipated that the study would recruit 2 0 subjects, time constraints a n d the exclusion of a few individuals d u e to interruptions in treatment hindered this effort. B e c a u s e of this d e c r e a s e in s a m p l e s i z e , power of the study w a s r e d u c e d from the pre-calculated 0.76, thereby limiting the ability to detect a treatment effect, given that an effect actually e x i s t e d . T h i s limitation of a relatively small s a m p l e s i z e c o u l d therefore  have h a d an effect on not attaining  statistical  significance in the study for the variables e x a m i n e d .  This  double-blind  study  examined  numerous  variables. Pain  and  strength  parameters were e x a m i n e d to demonstrate p e r c e i v e d m u s c l e s o r e n e s s a n d eccentric  strength  decrements,  MRI  and  quadricep  circumference  were  m e a s u r e d to indicate m u s c l e e d e m a that is d e m o n s t r a b l e in m u s c l e injury, a n d blood levels for CK, damage,  the  IL-6,  cytokine  MDA  w e r e e x a m i n e d to illustrate skeletal m u s c l e  response  and  lipid  peroxidation.  Through  these  m e a s u r e m e n t s , the patterns of recovery could be s e e n between both groups a n d  83  therefore would allow us to determine whether the experimental group s h o w e d a faster c o u r s e of recovery to b a s e l i n e levels over the five d a y testing period in contrast to the control group.  Perceived muscle soreness It w a s h y p o t h e s i z e d that the m u s c l e s o r e n e s s e x p e r i e n c e d by the D O M S protocol would be reduced by hyperbaric o x y g e n therapy over the testing period.  The  results of this study do not support this hypothesis. Hyperbaric o x y g e n therapy did not significantly d e c r e a s e m u s c l e pain or s o r e n e s s levels over the c o u r s e of four treatments.  T h e s e results contradict the findings of other reports indicating  that H B O therapy s u c c e s s f u l l y alleviated m u s c l e s o r e n e s s in athletes [1, 95].  B o r r o m e o et al [95] u s e d the visual a n a l o g s c a l e to a s s e s s pain levels for acute ankle sprains over a c o u r s e of three treatments. H e demonstrated pain levels peaking at 3.25 a n d 2.6 for the H B O a n d control  group,  respectively,  d e c r e a s i n g to near baseline levels by completion of the third treatment.  and  Harrison  et al [246] a n d Mekjavic et al [225] u s e d the visual a n a l o g s c a l e to a s s e s s pain levels in their s a m p l e population.  Their studies demonstrated peak ratings a s  high a s 7 for perceived m u s c l e s o r e n e s s for both groups. B a l n a v e et al [147] s h o w e d a rating of 6 for p e a k m u s c l e s o r e n e s s . A n o t h e r study by Z h a n g et al [229] s h o w e d peak pain levels of 2.5 a n d 4.5 for the intervention a n d p l a c e b o , respectively.  T h e s e studies all s h o w  ratings  of  muscle soreness  ranging  a n y w h e r e between 2.5 a n d 7. T h i s is similar to the results of this study, which demonstrate that m u s c l e s o r e n e s s p e a k e d in range between 3.5 a n d 4.5 for the control a n d H B O group, respectively.  T h e majority of previous studies cite perceived s o r e n e s s to peak  anywhere  between twenty-four and forty-eight hours post-exercise [129, 147, 2 2 5 , 246]. Similarly, this study supported t h e s e finding for both groups.  T h e perception of  m u s c l e s o r e n e s s p e a k e d 24 hours after the high-force eccentric e x e r c i s e protocol  84  for the experimental group while the control group p e a k e d 4 8 hours  post-  eccentric e x e r c i s e .  S e v e r a l r e a s o n s m a y be postulated for the failure of the H B O treatments  to  alleyiate m u s c l e s o r e n e s s . T h e findings of this study m a y be d u e to intersubject variability.  P a i n or p e r c e i v e d s o r e n e s s remains a subjective variable in which the  subject is a s k e d to graphically display their perception of p a i n .  P a i n thresholds  a n d tolerance levels are different a m o n g individuals [241]. T h e perception of s o r e n e s s is very difficult to interpret a s p e r c e i v e d m u s c l e s o r e n e s s is subjective in nature a n d its perception varies from individual to individual. Stewart [241] h a s cited that the p e r c e i v e d pain is a product of interpretation  by the mind; the  intensity felt c a n be i n c r e a s e d or d e c r e a s e d by c o n s c i o u s a n d u n c o n s c i o u s thought  or emotions.  The sensory component  subject's past e x p e r i e n c e where  attitudes  may be m o d u l a t e d  by  and psychological variables  the may  influence description of the s e n s a t i o n [129]. Error is easily introduced a s a n individual m a y report more or l e s s pain that what is actually felt. L e s s pain is usually the direction in which individuals follow in reporting pain [241].  T h e visual a n a l o g s c a l e h a s b e e n cited in previous literature  a s a reliable  instrument in m e a s u r i n g pain or p e r c e i v e d s o r e n e s s in subjects [15, 195-197, 199, 201].  However, by marking on a 10 c m line the level of pain felt c a n  introduce error in a c c u r a c y of what is actually felt v e r s u s what is begin reported. Furthermore, frequency in s a m p l i n g m a y a l s o bias results. S u b j e c t s m a y have b e e n a w a r e of their previous r e s p o n s e a n d p e r h a p s s u b c o n s c i o u s l y bias their following r e s p o n s e s .  N e w h a m et al [104] reported that pain a n d t e n d e r n e s s is usually localized to the distal third portion of the m u s c l e , in the region of the m u s c u l o t e n d i n o u s junction w h e r e m u s c l e pain receptors are most concentrated. In addition, pain a s s o c i a t e d with D O M S a p p e a r s medially, laterally a n d then distally, eventually s p r e a d i n g to the center of the m u s c l e belly by 4 8 hours [104]. G e n e r a l l y , however, pain is  85  evident throughout most of the affected m u s c l e belly while no pain is e x p e r i e n c e d during rest [132]. All subjects in the study d e m o n s t r a t e d a similar pattern of pain, with the m u s c u l o t e n d i n o u s junction to be the primary location of  soreness.  Furthermore, a s e x p e c t e d , all subjects (both control a n d experimental groups) d e m o n s t r a t e d no pain at rest but e x p e r i e n c e d discomfort during m o v e m e n t .  A n o t h e r p o s s i b l e explanation for the lack of significant differences in pain data w a s that the stimulus in this study did not c a u s e e n o u g h m u s c l e d a m a g e a n d injury. E v e n though w e i n c r e a s e d the angle of m u s c l e elongation to a n 110° - 3 5 ° m u s c l e range, p e r h a p s there wasn't e n o u g h stimulus to induce m u s c l e injury a n d d a m a g e to warrant sufficient D O M S to s h o w significant results in treatment. T h e e x e r c i s e protocol that w a s u s e d in this study w a s one that h a s b e e n tested by Mclntyre et al [129].  T h e s e investigators h a v e u s e d this protocol on the n o n -  dominant q u a d r i c e p m u s c l e to elicit a n d i n d u c e sufficient m u s c l e s o r e n e s s . L a u r e n c e [247], h a d s u g g e s t e d that in his study, at 95° -  35° flexion, the  quadricep m u s c u l a t u r e m a y not be elongated e n o u g h to induce sufficient m u s c l e s o r e n e s s . Therefore, this study e m p l o y e d his suggestion of using 110° -  35°  flexion to further elongate the quadricep m u s c l e , thus ensuring that the stimulus would in fact induce sufficient m u s c l e s o r e n e s s .  All subjects w e r e carefully  monitored during the e x e r c i s e protocol a n d were e n c o u r a g e d to exert m a x i m a l effort on e a c h contraction. T h e r e efforts were a l s o o b s e r v e d on the visual display to s e e that they w e r e not only m a x i m i z i n g their efforts but also maintaining this level consistently throughout the protocol.  H o w e v e r , e v e n though w e e n s u r e d  t h e s e efforts to induce m u s c l e injury, p e r h a p s the level of m u s c l e d a m a g e w a s not sufficient e n o u g h to warrant D O M S .  Eccentric Strength It w a s h y p o t h e s i z e d that treatment with hyperbaric o x y g e n would return strength faster than the normal c o u r s e of recovery. T h i s h o w e v e r w a s not the c a s e a s there were non-significant findings between g r o u p s in terms of treatment placebo sessions.  Hyperbaric o x y g e n did not accelerate the  injury  and  healing  86  involved in restoring strength a s neither the rate or magnitude of recovery of eccentric strength differed between groups. T h i s contradicts the findings  of  S t a p l e s et al [96] w h o demonstrated significant results w h e n a n a l y z i n g m e a n torque data, thus s h o w i n g a reduction in H B O v s . control group.  A n initial decline in strength w a s apparent b e t w e e n both g r o u p s in the study immediately after the eccentric e x e r c i s e protocol. T h i s w a s followed by a g r a d u a l return of torque o v e r four day, although t h e s e levels did not return to b a s e l i n e (pre-exercise) by d a y 5. M a c l n t y r e et al [129] h a v e reported similar findings. In their study, torque m e a s u r e m e n t s did not return to b a s e l i n e for 6 a n d 7 d a y s , respectively. T h e initial decline in strength corroborates other findings of strength d e c r e m e n t s immediately following a high intensity e x e r c i s e protocol [129, 130]. T h e y suggest that the initial decline in force m a y be a function of m e c h a n i c a l injury a n d fatigue (including myofibrillar disruption at the level of the Z-line), leading to an acute inflammatory r e s p o n s e  [129, 130]. A s e c o n d decline in  strength ( b i m o d a l pattern) w a s s e e n with six of the sixteen subjects in this study. T h i s corroborates findings in previous r e s e a r c h s u g g e s t i n g that a bimodal pattern of eccentric strength is s e e n in both an animal m o d e l a n d h u m a n s [129, 130]. F a u l k n e r a n d c o l l e a g u e s [130] further s u g g e s t that the s e c o n d decline in force o c c u r s in r e s p o n s e to phagocytic activity at the site of the initial d a m a g e . T h i s deficit in force d o e s not a p p e a r to be related to the level of s o r e n e s s s i n c e it o c c u r s prior to the s o r e n e s s a n d c a n remain for a greater period [131]. P r e v i o u s studies h a v e reported that there is no relationship between the level of s o r e n e s s a n d the decline in m u s c l e strength [129, 130, 1 3 1 , 132].  Strength d e c r e m e n t s  are s e e n immediately p o s t - e x e r c i s e while s o r e n e s s d e v e l o p s 2 4 - 4 8 hours poste x e r c i s e . O b s e r v i n g a bimodal pattern of eccentric torque, lends more e v i d e n c e a n d support to the theory that more than one m e c h a n i s m (i.e. m e c h a n i c a l , biochemical) is involved in e x e r c i s e - i n d u c e d m u s c l e d a m a g e .  Both groups d e m o n s t r a t e d  gradual  recovery o v e r time,  although  individual  variability in eccentric torque w a s evident a m o n g all subjects [ A P P E N D I X C  87  -  Figures D, E]. T o explain this variability, s e v e r a l explanations m a y be worth introducing. First, it would h a v e b e e n worthwhile to o b s e r v e the recovery pattern of all subjects for more than 4 treatment s e s s i o n s . B y only taking eccentric torque m e a s u r e s at p o s t - e x e r c i s e , 2 4 , 4 8 a n d 72 hours, w e weren't a b l e to follow the c o u r s e of recovery over the previous cited 5-7 d a y s recovery period for D O M S [132]. Furthermore, a learning effect c o u l d h a v e played a role in the recovery pattern s e e n a m o n g subjects. N u m e r o u s other investigators h a v e reported a training a n d adaptation effect during eccentric e x e r c i s e [131, 145, 172, 173]. N o s a k a et al [173] h a s s u g g e s t e d that a s u b s e q u e n t single bout of eccentric e x e r c i s e m a y reduce indicators of m u s c l e d a m a g e .  N e w h a m and colleagues  [104, 110, 131, 143] h a v e s u g g e s t e d that this m a y b e - c a u s e d by a c h a n g e in motor unit recruitment pattern, m u s c l e fibre adaptation and/or a regeneration of n e w m e c h a n i c a l l y resistant fibres resulting from d a m a g e a n d destruction to the original recruited fibres. T h e subjects could h a v e b e c o m e more comfortable a n d more e x p e r i e n c e d day by day a s they performed the eccentric strength ,test. Although subjects were e n c o u r a g e d verbally by the investigator to  produce  m a x i m a l effort throughout the e x e r c i s e protocol a n d isokinetic strength tests, a n d their efforts were monitored on the visual display, their energy level m a y h a v e fluctuated daily. T h i s would result in variability in strength levels throughout the c o u r s e of the testing period.  Quadricep Circumference Q u a d r i c e p circumference w a s m e a s u r e d at the 10 a n d 2 0 c m point a b o v e the superior portion of the patella, over the c o u r s e of the 5 d a y s . T h e premise for this m e a s u r e m e n t w a s that during the strenuous eccentric e x e r c i s e , m u s c l e fatigue a n d injury would give rise to m u s c u l a r e d e m a in the quadricep m u s c l e of the n o n dominant leg. E d e m a is a result of the inflammatory p r o c e s s a n d could lead to i n c r e a s e d pain a n d d e c r e a s e d range of motion.  T h e eccentric e x e r c i s e - i n d u c e d e d e m a , a s reflected in quadricep circumference, did  not  demonstrate  significance  between  groups.  Slight  increases  88  in  c i r c u m f e r e n c e were d e m o n s t r a t e d between groups at both the 10 a n d 20 c m mark, h o w e v e r not e n o u g h to attain statistical significance.  !  E v a n s et al [150, 165] s u g g e s t that e v i d e n c e of swelling h a s ranged from i n c r e a s e d circumference of the e x e r c i s e d m u s c l e 2 4 - 4 8 hour post-exercise to ultrastrucfural e v i d e n c e of p o s t - e x e r c i s e e d e m a . Similarly, the results of this study demonstrated that p e a k swelling (i.e. i n c r e a s e d quadricep circumference) at the 10 c m point o c c u r r e d on d a y 3 (24 hours post-exercise) for both groups. P e a k swelling at the 2 0 c m point w a s evident on d a y 4 (48 hours post-exercise) for both groups, h o w e v e r the m e a s u r e m e n t s for d a y s 2-5 for the control group remained lower than b a s e l i n e . Mekjavic et al [225] a l s o d e m o n s t r a t e d i n c r e a s e s in arm circumference on d a y s 3 a n d 5, p o s t - e x e r c i s e .  E d e m a is a result of either a n i n c r e a s e in v a s c u l a r permeability of small blood v e s s e l s , or l e a k a g e of intracellular fluid into the  extracellular s p a c e [131],  resulting in tissue hypoxia, d u e to the i n c r e a s e d o x y g e n diffusion distance from the capillaries to s o m e cells, a n d the i n c r e a s e d interstitial p r e s s u r e around the capillaries due to the fluid a c c u m u l a t i o n [225]. It h a s b e e n reported that m u s c l e e d e m a is c a u s e d by i n c r e a s e d a m o u n t s of d e g r a d e d protein c o m p o n e n t s of the m u s c l e a n d the release of protein-bound ions in d a m a g e d m u s c l e cells [42, 165, 154]. A s a result, an i n c r e a s e in intracellular osmotic fluid is present. Taylor et al [242] h a s s u g g e s t e d that an i n c r e a s e in p l a s m a proteins p r o d u c e a n i m b a l a n c e a c r o s s the v e s s e l wall a s t h e s e proteins m o v e into the interstitial s p a c e a n d fluid is drawn out. proteolytic  F r e e radicals m a y a l s o play a role in e d e m a a s they give rise to  enzymes  when  the  microcirculation  is  compromised.  Lipid  peroxidation initiated by free radicals d e c r e a s e s the barrier function of cell m e m b r a n e s a n d m a y be a s s o c i a t e d with m u s c l e fibre n e c r o s i s a n d e n z y m e release following d a m a g i n g e x e r c i s e [112].  T h e o x y g e n diffusion distance from  the capillaries c a n be up to four times greater than normal with hyperbaric o x y g e n . T h i s is mainly d u e to the larger p r e s s u r e gradient between capillary a n d tissue PO2, which i n c r e a s e s the n u m b e r of cells that c a n be o x y g e n a t e d w h e n  89  cellular oxygenation is limited by e d e m a [61]. Therefore, hyperbaric o x y g e n m a y be beneficial in reducing e d e m a by ensuring a d e q u a t e cellular oxygenation to maintain cellular energy production, allowing the cell to fuel its A T P driven p u m p s a n d c h a n n e l s , thus e n h a n c i n g the reabsorption of fluid from the extracellular s p a c e a n d d e c r e a s e e d e m a [83, 225]. Furthermore, s e c o n d a r y vasoconstriction by H B O would reduce blood inflow by 2 0 % without d e c r e a s i n g o x y g e n delivery. T h i s would a l s o reduce e d e m a by d e c r e a s i n g the, intravascular  hydrostatic  p r e s s u r e , a n d establishing a more favorable p r e s s u r e gradient for fluid m o v e m e n t out of the interstitial s p a c e b a c k into the capillaries [225].  In contrast to studies that have demonstrated a reduction in e d e m a by hyperbaric o x y g e n therapy [3, 5, 7 2 - 7 4 , 91], there is inconclusive e v i d e n c e that H B O therapy r e d u c e s e d e m a with soft tissue athletic injuries. [95]. O u r study indicates that this modality of treatment  w a s not effective  in minimizing the e d e m a  formation  a s s o c i a t e d with e x e r c i s e - i n d u c e d m u s c l e injury. T h i s finding is similar to that of Mekjavic et al [225] w h o s u g g e s t e d that e d e m a in their study w a s not of sufficient magnitude to establish the i n c r e a s e d diffusion d i s t a n c e s a n d i n c r e a s e d interstitial p r e s s u r e , which might promote tissue hypoxia.  Blood Analysis: Creatine Kinase, Malondialdehyde and Interleukin-6  Creatine  Kinase  (CK)  C r e a t i n e kinase h a s b e e n studied extensively in relation to m u s c l e d a m a g e d u e to strenuous e x e r c i s e s i n c e this p l a s m a e n z y m e is found exclusively in the skeletal a n d c a r d i a c m u s c l e [41, 7 7 , 127, 1 5 3 , 160, 162]. It h a s b e e n reported that the extent of release of this p l a s m a e n z y m e is d e l a y e d a n d is a direct 1  c o n s e q u e n c e of m u s c l e d a m a g e a n d e x e r c i s e [41, 1 1 2 , 1 6 0 , 162]. Furthermore, the magnitude a n d duration of the i n c r e a s e in C K activity is affected by the type a n d intensity of activity a s well a s previous level of activity [115]. It h a s a l s o b e e n cited that C K h a s a high d e g r e e of variability a n d its r e s p o n s e varies between 90  individuals [41, 7 7 , 162]. F o r e x a m p l e , one study in which subjects performed a n eccentric e x e r c i s e s h o w e d i n c r e a s e s in C K activity up to 3 0 0 0 0 m-units/ml, while other subjects s h o w e d i n c r e a s e s of less than 5 0 0 m-units/ml [248].  It w a s h y p o t h e s i z e d that hyperbaric o x y g e n would attenuate the r e s p o n s e of creatine k i n a s e over the c o u r s e of treatment. e x e r c i s e levels would  i n c r e a s e from  dampened  to the  in contrast  control  More specifically, 4-hours post-  b a s e l i n e , but this  r e s p o n s e would  group. A n a l y s i s of the  data did  be not  demonstrate significant findings b e t w e e n groups for treatment with hyperbaric o x y g e n o v e r the c o u r s e of one w e e k .  O u r findings are consistent with t h o s e of  previous studies indicating that the eccentric e x e r c i s e protocol, w h i c h w a s u s e d in our investigation, resulted in a quantifiable m u s c l e injury [100, 153, 177, 179]. H o w e v e r the lack of significance a c r o s s both groups s u g g e s t that H B O w a s not effective in treating m u s c l e injury or D O M S .  In f e m a l e s , the level of C K r a n g e s a n y w h e r e between 4 5 - 2 3 0 u/L, with e x e r c i s e , trauma, surgery a n d other ailments increasing levels 10-15 times normal range v a l u e s [249]. Furthermore, s o m e individuals may have i n c r e a s e s up to 50 times the upper limit of normal range but this r e s p o n s e is s e e n in conditions s u c h a s m u s c u l a r dystrophy [249]. Elevation in levels may be s e e n a s early a s 2-4 hours post e x e r c i s e , with v a l u e s returning to normal by 4 8 a n d 72 hours post-injury. F o r the p u r p o s e of the present study, blood w a s withdrawn from subjects 4 hours post-exercise, followed by s u b s e q u e n t testing for 3 d a y s (24, 4 8 a n d 72 hours) to e x a m i n e i n c r e a s e s in s e r u m C K activity levels.  A statistical a n a l y s i s on the creatine k i n a s e data w a s performed on two s e p a r a t e o c c a s i o n s , one with the entire subject pool a n d a s e c o n d with the removal of one r e s p o n d e r in the experimental group w h o a p p e a r e d to h a v e a C K value well b e y o n d the normal limits (>2500 uL). T h i s w a s d o n e to e n s u r e that this high variability  introduced  into the  data by this subject  did not  mask  statistical  91  significant findings in the s a m p l e . Both a n a l y s e s demonstrated no significance b e t w e e n g r o u p s for treatment effects.  T h e s e r u m creatine kinase r e s p o n s e to the eccentric e x e r c i s e protocol in the current investigation s h o w e d c o n s i d e r a b l e individual variability, although most subjects d i s p l a y e d an immediate i n c r e a s e in s e r u m C K 4 hours p o s t - e x e r c i s e . E b b e l i n g a n d C l a r k s o n [127] s u g g e s t e d that subjects w h o a p p e a r to be quite similar w h e n performing the s a m e amount  of eccentric e x e r c i s e c a n have  c h a n g e s in circulating C K activity that are different by orders of magnitude. M a n y factors influence intersubject variability, including a g e , gender, body composition a n d race [147]. In contrast, E b b e l i n g a n d C l a r k s o n [127] point out that the i n c r e a s e in circulating C K is "unrelated to either the development of s o r e n e s s , the amount of strength l o s s after e x e r c i s e , fitness level of the subject, or lean body weight". Other investigators h a v e looked at genetic variation a n d s e r u m C K activity to understand the variability [112]. It is likely that the p o s t - e x e r c i s e rise in circulating C K activity is a manifestation of skeletal m u s c l e d a m a g e but not a direct indicator [127].  Malondialdehyde  (MDA)  A growing amount of e v i d e n c e indicates that free radicals play a n important role a s mediators of skeletal m u s c l e d a m a g e a n d inflammation  after  strenuous  e x e r c i s e [239, 2 4 0 , 244]. It h a s b e e n postulated that the generation of o x y g e n free radicals is i n c r e a s e d during e x e r c i s e a s a result of mitochondrial o x y g e n c o n s u m p t i o n a n d electron transport flux, inducing lipid peroxidation [239].  Lipid  peroxidation is potentially a very d a m a g i n g p r o c e s s to the o r g a n i z e d structure a n d function of m e m b r a n e s . R e c e n t studies indicate that a) o x y g e n free-radicals mediate, at least in part, the i n c r e a s e d m i c r o v a s c u l a r permeability p r o d u c e d by reoxygenation, a n d b) free radical s c a v e n g e r s c a n reduce skeletal m u s c l e n e c r o s i s occurring after prolonged i s c h e m i a [244]. T h e literature supports the notion of the interrelationship b e t w e e n i s c h e m i c tissue a n d inflammatory cells a n d therefore, c o n c l u d e s that capillary plugging by granulocytes a n d o x y g e n free  92  radical formation m a y contribute to the i s c h e m i c injury [244]. T h e role of reactive o x y g e n s p e c i e s in the mediation of e x e r c i s e - i n d u c e d oxidative d a m a g e to m u s c l e a n d the protection offered by anti-oxidant d e f e n s e s y s t e m s have b e e n h a v e a l s o b e e n well studied [138, 139]. M a l o n d i a l d e h y d e , a product of lipid peroxidation is a w a y of estimating free radical generation a s a result of skeletal m u s c l e damage.  It w a s h y p o t h e s i z e d that the eccentric e x e r c i s e protocol would induce skeletal m u s c l e d a m a g e , thus ultimately giving rise to free radicals a n d lipid peroxidation. Hyperbaric o x y g e n therapy  would  h a v e a n effect  on  lipid peroxidation  by  e n h a n c i n g the antioxidative d e f e n s e m e c h a n i s m s a n d increasing the b i o c h e m i c a l d e f e n s e m e c h a n i s m s against free radicals [193]. A s a result, H B O therapy would attenuate M D A levels.  A n a l y s i s of the data did not demonstrate significant findings for treatment with hyperbaric o x y g e n . Hyperbaric o x y g e n h a d no effect on reducing free radical d a m a g e a n d limiting lipid peroxidation levels a s e v i d e n c e d by m a l o n d i a l d e h y d e . T h e results of both groups demonstrated very little fluctuation in m a l o n d i a l d e h y d e levels, increasing slightly after the eccentric e x e r c i s e protocol a n d d e c r e a s i n g c l o s e to b a s e l i n e levels by day 5. T h e m e a n v a l u e s for M D A in this study r a n g e d between 4.0 a n d 4.6 nmol/ml. Other studies have reported M D A v a l u e s in the range of 1.0 - 3.0 nmol/ml [229, 2 5 0 , 251]. T h i s difference may be due to the intensity a n d duration  of the prescribed e x e r c i s e protocol, the variability  s a m p l i n g times for blood a n a l y s i s ^and nature of injury. C h i l d et al [251]  in  only  s a m p l e d M D A levels pre a n d immediately p o s t - e x e r c i s e . T h i s , therefore would not allow us to determine what the levels of M D A would be at 24, 4 8 a n d 72 hours post-exercise. Furthermore, Novelli et al [250] e x a m i n e d b i o p s i e s taken from the right femoral quadricep m u s c l e at three time points during  aortic  surgery.  93 v  O n e plausible explanation for not demonstrating significance in the present study c o u l d be that the eccentric e x e r c i s e protocol didn't elicit e n o u g h skeletal m u s c l e d a m a g e to o b s e r v e c h a n g e s p o s t - e x e r c i s e .  In other w o r d s , the effect w a s  minimal which did not allow us to o b s e r v e a noticeable c h a n g e with H B O treatments,  a s e x p e c t e d . If the e x e r c i s e protocol  induced sufficient  muscle  d a m a g e or injury, the levels of M D A would have i n c r e a s e d higher than what w a s observed.  Furthermore,  if  H B O therapy  was  beneficial  in  reducing  lipid  peroxidation, the present study might then h a v e b e e n able to s e e a noticeable effect a n d obtain statistical significance.  A n o t h e r factor that m a y h a v e played a role in lipid peroxidation detection m a y be the m o d e of a s s a y s a m p l i n g (i.e. direct spectrophotometry).  M D A a s s a y is the  most generally u s e d test in the appreciation of the role of oxidative s t r e s s in injury a n d d i s e a s e . M D A is o n e of several products formed during the radical i n d u c e d d e c o m p o s i t i o n or b r e a k d o w n of e n d o p e r o x i d e s during the last s t a g e s of the oxidation of polyunsaturated fatty a c i d s . Most often, at high temperature a n d low p H , M D A readily participates in nucleophilic addition reaction with 2-thiobarbituric acid ( T B A ) , generating a red, fluorescent 1:2 M D A : T B A adduct [191]. B e c a u s e of this fact, a n d facile a n d sensitive m e t h o d s to quantify M D A (as the free a l d e h y d e or its T B A derivative), the " T B A test" is u s e d a s a routine test to detect a n d quantify lipid peroxidation in a wide array of s a m p l e types [191]. T h i s reaction that o c c u r s is very sensitive but its specificity, e v e n with improvement of preanalytical (sampling, preservatives), a n d analytical s t a g e s (fluorescence, H P L C ) is still a matter of debate [243].  M D A itself participates in reactions with  m o l e c u l e s other than T B A a n d is a catabolic substrate. O n l y certain  lipid  peroxidation products generate M D A (invariably with low yields) a n d M D A is neither the s o l e e n d product of fatty peroxide formation a n d d e c o m p o s i t i o n nor a s u b s t a n c e generated exclusively through lipid peroxidation [191]. A n extensive review of the literature h a s c o n c l u d e d that M D A determination a n d the T B A test c a n offer, at best, a narrow a n d s o m e w h a t empirical window on the c o m p l e x p r o c e s s of lipid peroxidation [191, 234]. It is subject to interferences, which if not  94  c o n s i d e r e d , m a y lead to e r r o n e o u s results. Future studies s h o u l d p e r h a p s f o c u s o n more sensitive a n d specific (chemical or p h y s i c a l methods) m o d e s of lipid peroxidation a s s e s s m e n t .  Interleukin  6 (IL-6)  Increasing n u m b e r s of reports h a v e d e s c r i b e d the IL-6 r e s p o n s e to injury [187188, 231-235]. Eccentric e x e r c i s e is a s s o c i a t e d with a n i n c r e a s e in s e r u m IL-6 concentrations a n d is significantly correlated with the concentration of C K in s u b s e q u e n t d a y s following injury. T h e time c o u r s e of cytokine production, the c l o s e association with m u s c l e d a m a g e a n d the finding of IL-6 in skeletal m u s c l e b i o p s i e s after intense e x e r c i s e lend support to the i d e a that during eccentric e x e r c i s e myofibers are m e c h a n i c a l l y d a m a g e d a n d that this p r o c e s s stimulates the local production of inflammatory cytokines [197].  IL-6 is an integral cytokine  mediator of the acute p h a s e r e s p o n s e to injury a n d infection. It plays a n active role in the post-injury i m m u n e r e s p o n s e , making it a n attractive therapeutic target in attempts to control hyperinflammatory-provoked injury [235].  It w a s h y p o t h e s i z e d that the  eccentric e x e r c i s e protocol  would  induce  an  inflammatory r e s p o n s e that would lead to the production of IL-6. T h i s i n c r e a s e in p l a s m a levels of IL-6 would be alleviated by hyperbaric o x y g e n treatments over the four d a y s of therapy a n d the inflammatory r e s p o n s e would be r e d u c e d by treatment.  Statistical a n a l y s i s of the data demonstrated non-significant findings for both groups. T h i s must be interpreted cautiously, a s the s a m p l e size w a s quite small a n d standard errors of the m e a n s large. analysis was demonstrated.  A great d e g r e e of variability in the  T h i s variability w a s unexplainable a n d may largely  be attributed to error in laboratory technique. A n o t h e r limitation w e e n c o u n t e r e d in the study w a s that the laboratory, in conducting the a n a l y s i s , contaminated a n d destroyed 8 of the 16 subjects (50%), thus leaving a total of four subjects per group (n=4). T h i s therefore diminished our s a m p l e s i z e a n d a s a result, w a s not  95  sufficient e n o u g h attain significance to substantiate or refute the involvement of hyperbaric o x y g e n therapy in reducing IL-6 elevations during m u s c l e d a m a g e a n d injury.  E x a m i n a t i o n of the m e a n s of both groups provided perplexing results. T h e experimental group d e m o n s t r a t e d a relatively stable level of IL-6 while the control group s h o w e d an irregular pattern [Figure 11]. O v e r a l l , all subjects in both groups h a d variability in the r e s p o n s e of IL-6 to the eccentric e x e r c i s e protocol a n d s u b s e q u e n t treatment s e s s i o n s . T h e s e results do not typify the normal c o u r s e of IL-6 that h a s b e e n reported in the literature.  Biffl et al [235] h a s s u g g e s t e d that  IL-6 concentration rose within 2-4 hours post-trauma. T h i s w a s not evident in our investigation a s IL-6 levels elevated 2 4 hours post-exercise for both groups. T h e magnitude of elevation of IL-6 is related directly to the d e g r e e of tissue injury [235], ranging <50 pg/ml in healthy individuals a n d rising a s high a s 1000 ng/ml in certain s e v e r e d i s e a s e states [233]. A s s u m i n g that the results w e r e reliable a n d valid, it a p p e a r s that hyperbaric o x y g e n w a s doing s o m e t h i n g to the p l a s m a levels of IL-6 in the experimental group, m a k i n g the r e s p o n s e s stable a n d less s p o r a d i c than the control group. T h i s w a s evident in the graphical depictions of the  means  of  both  control  and  experimental  groups  [Figure  11].  Could  administering 1 0 0 % o x y g e n at high p r e s s u r e h a v e s o m e impact on IL-6? A study by R o h d e et al [188] at the C o p e n h a g e n M u s c l e R e s e a r c h C e n t r e demonstrated that a n i n c r e a s e of 5 7 0 % in IL-6 concentration w a s s e e n in the control trial (pree x e r c i s e to 2 hours post-exercise) a n d returned to p r e - e x e r c i s e levels at d a y 2. B a u e r et al [233] reported that elevated levels of IL-6 c o u l d be found a s early a s a few hours or up to a few d a y s a n d t h e s e levels fluctuate d e p e n d i n g on the acute inflammatory r e s p o n s e . F o r e x a m p l e , patients undergoing elective surgery r e a c h e d IL-6 p l a s m a concentrations of meningitis ranged b e t w e e n 10 -  100 pg/ml, while patients with viral  1000 ng/ml [233].  A review of the  literature  proved u n s u c c e s s f u l , a s w e were unable to support or explain the variable findings found in this study. Future studies n e e d to further e x a m i n e this question more in detail by p e r h a p s f o c u s s i n g on the inflammatory r e s p o n s e a n d e x a m i n i n g  96  in more detail how hyperbaric o x y g e n therapy affects the e n s u i n g  cytokine  response.  Magnetic  Resonance  Imaging  Magnetic  r e s o n a n c e imaging, being a powerful  non-invasive  measurement,  allowed us to determine whether the eccentric protocol did in fact  produce  e x e r c i s e - i n d u c e d m u s c l e d a m a g e a n d a s well a s quantify this level of m u s c l e injury a s e v i d e n c e d by m u s c l e e d e m a .  S k e l e t a l m u s c l e T 2 relaxation time h a s  b e e n a s s o c i a t e d with c h a n g e s in the fluid c o m p o n e n t of injured m u s c l e a n d u s e d a s a marker of e d e m a , inflammation a n d injury [177, 179, 246]. T h i s is consistent with previous findings that s u g g e s t that e d e m a results from  high  intensity  eccentric e x e r c i s e [174, 177-179].  In the current study, it w a s h y p o t h e s i z e d that hyperbaric o x y g e n treatment would d e c r e a s e m u s c l e e d e m a that w a s induced by the eccentric protocol over the time c o u r s e of therapy.  Statistical a n a l y s i s for both T2-weighted i m a g e s a n d S T I R  i m a g e s demonstrated non-significant findings for treatment effect o v e r time. T h i s w a s evident in all three m u s c l e s : rectus femoris, v a s t u s m e d i u s , a n d v a s t u s lateralis.  Although T 2 a n d S T I R i m a g e s indicated a n i n c r e a s e in e d e m a 2 4 -  hours post-exercise for the s a m e three m u s c l e s , respectively, treatment  with  hyperbaric o x y g e n did not induce a n effect sufficient e n o u g h to reduce e d e m a a n d a c h i e v e statistical significance.  T h e nuclear magnetic r e s o n a n c e ( N M R ) signal from which M R i m a g e s are constructed a r i s e s from the magnetic behavior of the hydrogen nuclei in tissue water a n d fat m o l e c u l e s w h e n tissue is p l a c e d in a strong magnetic field. Inside a strong magnet, the hydrogen nuclei c a n be excited by the input of energy, specifically, by a "pulse" of energy at the resonant radiofrequency [226]. T h i s excitation c a u s e s a fraction of the nuclei to oscillate together in the magnetic field in a n orientation  that g e n e r a t e s a detectable magnetic  signal that c a n  be  recorded electronically a n d a s a result, form i m a g e s [226]. H o w e v e r , immediately  97  after excitation, nuclei in different  magnetic environments begin to oscillate  differently, the oscillation b e g i n s to b r e a k d o w n a n d the o b s e r v e d signal d e c a y s a w a y [226]. D e c a y of the N M R signal is referred to a s transverse relaxation time (T2 relaxation time) a n d is the time constant that c h a r a c t e r i z e s the exponential d e c a y of the signal after the initial excitation [226]. T 2 relaxation time, h o w e v e r d o e s not s u p p r e s s fat on the o b s e r v e d i m a g e s . Short Tip Inversion R e c o v e r y (STIR) i m a g e s allows o n e to sort out tissue a s this form of imaging only s h o w s fluid (bright signal) a n d s u p p r e s s e s fat [ A P P E N D I X D].  MRI is the most sensitive n o n - i n v a s i v e imaging method for detection  and  quantification of m u s c l e e d e m a . T 2 weighted i m a g e s a n d S T I R i m a g e s are particularly sensitive for detection of e d e m a a n d i n c r e a s e d water content in body tissue.  T h e MRI data in this study is b a s e d o n signal intensity m e a s u r e m e n t s . MRI signal intensity m e a s u r e m e n t s are e x p r e s s e d in arbitrary units, which w e r e variable due to a n u m b e r of factors. T h e signal intensity obtained from a specific v o l u m e of tissue d e p e n d s on the physical properties of the tissue (such a s water content, tissue composition, a n d tissue density), the spatial location of the tissue relative to the signal reception coil, the performance characteristics of the radiofrequency system,  the  performance  performance  characteristics  characteristics of  the  of  the  amplification  reception system.  coil, Many  and of  the  these  characteristics are variable from m o m e n t to moment a n d from location to location within the body. T o o v e r c o m e this variability, this study u s e d a ratio of signal intensity in the s t r e s s e d m u s c l e divided by the signal intensity simultaneously obtained in the c o r r e s p o n d i n g exact a n a t o m i c location in the opposite quadricep muscle.  T h i s type of internal control o v e r c o m e s variability a s s o c i a t e d with (for  instance) body hydration, body position relative to the coil, a n d the operating characteristics of the MRI s y s t e m .  98  T h i s m e a s u r e m e n t s y s t e m is s u s c e p t i b l e to signal intensity variability resulting from a s y m m e t r i c positioning of the body within the MRI s c a n n e r , h o w e v e r c a r e w a s taken to e n s u r e that all subjects w e r e positioned in a symmetric a n d midline fashion within the s c a n n e r s y s t e m .  A n o t h e r potential s o u r c e of error is injury  (acute or chronic) in the dominant (control) leg, which c o u l d affect signal intensity measurements.  The  images  were  visually  inspected  and  any  area  of  abnormality/injury in the control leg w a s a v o i d e d . D u e to the e x p e n s e involved in conducting M R I ' s on all subjects, this study h a d to limit image s c a n s to 3 per subject. A s a result, b a s e l i n e i m a g e s w e r e t a k e n , followed by a s c a n 2 4 hours post-exercise  and  finally  72  hours  post-exercise.  Ideally,  images  taken  throughout the entire w e e k of the study a n d p e r h a p s e v e n 96 a n d 120 hours post-injury would h a v e b e e n beneficial to s e e a pattern of recovery.  B y only  taking i m a g e s at 0, 24 a n d 72-hour time points, introduces the opportunity for potentially m i s s i n g what is h a p p e n i n g b e t w e e n those periods. Finally, e n o u g h m u s c l e injury m a y not  have b e e n d e m o n s t r a t e d  by the eccentric e x e r c i s e  protocol to attain statistical significance in treating this soft-tissue injury with hyperbaric o x y g e n treatments.  <  99  CHAPTER 6: SUMMARY, CONCLUSIONS AND RECOMMENDATIONS Summary Hyperbaric o x y g e n is a field of m u c h controversy a n d s k e p t i c i s m . T h e handful of studies that have b e e n published, differ in results a s to whether this form of therapy p r o v e s effective purpose  of this  in treating sport a n d exercise-related injuries. T h e  study w a s to determine  whether  intermittent e x p o s u r e to  hyperbaric o x y g e n played a beneficial role in the recovery from a n acute soft tissue injury. A group of sedentary f e m a l e students (n=16) were  randomly  a s s i g n e d to o n e of two groups (control [air] & experimental [hyperbaric oxygen]). T h e subjects performed a high intensity e x e r c i s e protocol, followed by the administration  of four  daily  treatment  sessions.  N o significant  differences  (p<0.05) a m o n g a n y of the d e p e n d e n t v a r i a b l e s were o b s e r v e d between those treated with hyperbaric o x y g e n a n d those that received normoxic conditions.  Conclusions F r o m a review a n d a n a l y s i s of pain, strength, quadricep c i r c u m f e r e n c e , C K , IL-6, M D A a n d MRI data, it a p p e a r s that hyperbaric o x y g e n treatment did not h a v e a n y effect o n the following:  1. reducing p e r c e i v e d m u s c l e s o r e n e s s over the five-day testing period. 2. improving eccentric m u s c l e strength after the e x e r c i s e protocol over the four-day c o u r s e of treatment. 3. reducing e d e m a a s e v i d e n c e d by 10 a n d 2 0 c m q u a d r i c e p s c i r c u m f e r e n c e measurements. 4. reducing the C K r e s p o n s e , w h i c h , is e x a c e r b a t e d by skeletal m u s c l e damage. 5. reducing IL-6 levels which h a v e recently b e e n demonstrated to i n c r e a s e considerably following eccentric e x e r c i s e .  100  6. reduce M D A levels indicative of lipid peroxidation, which o c c u r s during eccentric e x e r c i s e a n d s u b s e q u e n t skeletal m u s c l e d a m a g e . 7. reducing e d e m a a s e v i d e n c e d by MRI imaging  Recommendations for future studies T h e present study d o e s not lend support to the efficacy of using this treatment modality in soft-tissue injuries. H o w e v e r , a more f o c u s e d study on fewer "key" variables a n d a larger s a m p l e s i z e m a y a d d light to this a r e a of r e s e a r c h .  The  cost of doing the a b o v e study w a s quite e x p e n s i v e a n d therefore, modifications were implemented to reduce c o s t s (e.g. 5 to 3 MRI s c a n s / s u b j e c t ) .  Future studies s h o u l d :  1. focus on certain important variables s u c h a s strength, blood p a r a m e t e r s (i.e. M D A & IL-6) a n d MRI over more time periods. T h e s e variables do not carry the subjective c o m p o n e n t that pain a n d q u a d r i c e p s circumference m e a s u r e m e n t s employ a n d the variability c o m p o n e n t that C K h a s b e e n cited a s having. 2.  increase the s a m p l e size to e n s u r e a higher power a n d reduction of type 2 error.  3.  e x a m i n e p l a s m a P G E , s i n c e prostaglandins are p r o d u c e d by invading 2  m a c r o p h a g e during injury a n d have b e e n cited to sensitize pain receptors. Furthermore, P G E  2  h a s b e e n implicated in the generation of inflammatory  pain [156, 157]. 4.  e x a m i n e whether the D O M S m o d e l is appropriate for inducing injury to the m u s c l e . In other words, d o e s this type of soft-tissue injury create a n oxygen  diffusion  distance  great  enough  for  hyperbaric  oxygen  to  demonstrate its beneficial effects, increasing the o x y g e n diffusion gradient. Investigations s h o u l d p e r h a p s focus on other types of injuries s u c h a s ligamentous injuries, where there is a n i n c r e a s e in the o x y g e n diffusion  101  distance a n d the n e e d for a reduction in the inflammatory p r o c e s s is warranted.  It is worth noting that the H B O treatment protocol that w a s u s e d (60 minute s e s s i o n s at 2.0 A T A , o n c e a d a y for four days) in this study w a s b a s e d on previous r e s e a r c h protocols looking at the therapeutic effects of this modality. However,  p r e s s u r e , duration  of treatment,  frequency  a n d total  number  of  treatments vary for a given indication. C a r e must be taken to e n s u r e that o x y g e n toxicity levels are c l o s e l y monitored. F o u r treatment s e s s i o n s w e r e c h o s e n from a practical perspective, a s a n athlete being treated for a n injury s h o u l d h a v e indication of significant improvement in the rate of recovery within s e v e r a l d a y s .  C a u t i o n s h o u l d be applied w h e n negating the effects of hyperbaric o x y g e n therapy on soft tissue injuries b a s e d on the results of this study. Limitations of the study s h o u l d be carefully c o n s i d e r e d a s m a n y factors m a y affect the therapeutic effects of H B O on m u s c l e injury. Harrison et al [246] s u g g e s t that H B O therapy will be effective  in the treatment  of athletic  injuries that involve a  greater  magnitude of soft tissue d a m a g e or for injuries in which o x y g e n availability m a y be more of a limiting factor d u e to the location of the injury or magnitude of local edema.  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Lupus 1999;8:409-15.  119  APPENDIX  120  Study Procedure: S h o u l d I be c h o s e n to participate in this study, I will be a s k e d to attend 4 s e s s i o n s of treatment (1 per d a y for 5 d a y s ) . In addition, I a m fully aware that 12 ml of blood (5ml is equivalent to 1 tsp.) will be withdrawn prior to a n d 6 hours post-exercise, a s well a s e a c h d a y following treatment a n d magnetic r e s o n a n c e imaging (MRI) will a l s o be taken of my q u a d r i c e p m u s c l e . I will a l s o be a s k e d to refrain from any form of e x e r c i s e 12 hours post-injury a n d k e e p a brief diary of my activities a s a reference. I understand that I will be p l a c e d in either a control or experimental group by randomization of n a m e s using a computer. In order to avoid bias, neither the subject nor the investigator will know what treatment the subject is receiving. H o w e v e r , in c a s e of a n e m e r g e n c y , the c o d e c a n a n d will be broken.  Exclusion: I understand that I m a y be e x c l u d e d from the study if I play on a t e a m sport, run or weight train a s part of my physical regimen, more than 3 hours per week. Individuals w h o s e activities involve jumping and/or squatting will also be e x c l u d e d from the study. In addition, individuals w h o have e x p e r i e n c e d d e l a y e d onset m u s c l e s o r e n e s s to their q u a d r i c e p s in the last three months, who have h a d a past history of s e v e r e joint injury, arthritis or other chronic illnesses a n d w h o are taking prescription drugs or a n a l g e s i c s will be e x c l u d e d . Hyperbaric o x y g e n contraindications (e.g. diabetes, lung c y s t s , epilepsy, upper respiratory tract infections, pregnancy, fever or confinement anxiety) will also be evaluated.  Risks and Benefits: I understand that the risks are minimal in this study, with mild aural barotraumas (ear a c h e ) , n a u s e a , tooth a n d s i n u s pain a n d blurred vision occurring rarely. T h e benefit, if s u c c e s s f u l , will be no pain a n d a return of strength of the quadricep m u s c l e s o o n e r than the normal c o u r s e of recovery for this condition.  Confidentiality: I understand that any information resulting from this study will be kept strictly confidential a n d all d o c u m e n t s will be identified only by a c o d e n u m b e r a n d kept in a locked filing cabinet. I will not be identified by n a m e in any report of the c o m p l e t e study.  122  [  Contact: If I h a v e a n y c o n c e r n s about my treatment or rights a s a subject, I m a y contact Dr. R . D . Spratley, Director of R e s e a r c h S e r v i c e s at the University of British C o l u m b i a , at 8 2 2 - 8 5 9 5 . If I h a v e a n y q u e s t i o n s or desire further information with respect to the study, I s h o u l d contact any of the a b o v e investigators.  New Findings: I will be a d v i s e d of any n e w information that b e c o m e s available that m a y affect my willingness to remain in this study.  Patient Consent: I understand that participation in this study is entirely voluntary a n d that I m a y refuse to participate or withdraw from the study at any time without consequences. I h a v e received my c o p y of the c o n s e n t form for my own records. I c o n s e n t to participate in this study.  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