THE FEASIBILITY OF BLOOD

FLOW RESTRICTION EXERCISE FOR INDIVIDUALS WITH

INCOMPLETE SPINAL CORD INJURIES

A dissertation submitted to the

Kent State University College

of Education, Health, and Human Services

in partial fulfillment of the requirements

for the degree of Doctor of Philosophy

By

Jonathon R. Stavres

August 2017
 A dissertation written by

Jonathon R. Stavres

B.S., Indiana University of Pennsylvania, 2013

M.S., Indiana University of Pennsylvania, 2014

Ph.D., Kent State University, 2017

Approved by

__________________________, Director, Doctoral Dissertation Committee

John McDaniel

__________________________, Member, Doctoral Dissertation Committee

Adam Jajtner

__________________________, Member, Doctoral Dissertation Committee

J. Derek Kingsley

__________________________, Member, Doctoral Dissertation Committee

Kimberly Peer
Accepted by

__________________________, Director, School of Health Sciences

Lynne E. Rowan

__________________________, Dean, College of Education, Health and Human

James C. Hannon Services

ii
STAVRES, JONATHON R., Ph.D., August, 2017 Exercise Physiology

THE FEASIBILITY AND EFFICACY OF BLOOD FLOW RESTRICTION EXERCISE

FOR INDIVIDUALS WITH INCOMPLETE SPINAL CORD INJURIES (151 pp.)

Director of Dissertation: John D. McDaniel, Ph.D.

INTRODUCTION: Individuals with incomplete spinal cord injuries (iSCIs)

suffer from significant functional impairments which impact quality of life and limit

exercise intensity. Blood flow restriction (BFR) exercise has been shown to maximize

muscular adaptations to low-intensity in the general population, but has yet to be

translated to the spinal cord injured population. PURPOSE: The purpose of this project

was to examine the feasibility and safety of BFR exercise in a sample of subjects with

iSCI; and to compare physiological responses between BFR and non-BFR exercise.

METHODS: Nine individuals with iSCIs completed two bouts of knee extension (three

sets of ten repetitions) in counterbalanced order. One set included BFR, and the other did

not. Signs and symptoms of deep vein thrombosis (DVT) formation and autonomic

dysreflexia (AD) were monitored during and after BFR exercise. Muscular activation

(iEMG), hemodynamics, tissue oxygenation characteristics, work output, perceived

difficulty, pain, and lactate formation were also compared across each trial and between

conditions. RESULTS: Our data indicate that subjects were able to perform BFR

exercise without experiencing any signs/symptoms of DVT or AD; and without any

decrements in work output, muscular activation, or lactate accumulation compared to

non-BFR exercise of the same workload. Our data also indicate that BFR elicited venous
pooling and hyperemia between contractions, indicated by increases in oxyhemoglobin,

deoxyhemoglobin, and total hemoglobin (+12.3±96.7 and +105.4±76.7, respectively);

while tissue oxygenation decreased (6.5±3.0%; p<0.05 for all comparisons).

CONCLUSION: These data suggest that BFR exercise can be safely performed in

individuals with iSCIs without elevated pain or difficulty. Future investigations may

consider exploring the effects of sustained BFR training on muscular characteristics and

functional outcomes in this population.

 ACKNOWLEDGEMENTS

I would like to acknowledge the following individuals who were integral to the

successful completion of this dissertation. First, Dr. John D. McDaniel served as the dissertation

advisor for this project, and offered his support and guidance throughout the entirety of this

process. This project could not have been possible without his intellect and abundance of

patience. Second, I would like to thank the dissertation committee members, Dr. J. Derek

Kingsley, Dr. Kimberly S. Peer, and Dr. Adam Jajtner who all provided quality feedback and

thoughtful insight during the proposal and data collection phases. I would also like to thank

Amber Brochetti and Martin Kilbane who offered their services as licenses physical therapists

and aided in recruitment efforts. Lastly, I would like to thank Tyler Singer who dedicated a

substantial portion of his time to data collection for this project. This project could not have

been possible without the help of each individual mentioned above; thank you.

iii
 TABLE OF CONTENTS
Item Page

ACKNOWLEDGEMENTS ........................................................................................ iii

LIST OF FIGURES ....................................................................................................vii

CHAPTER
I. INTRODUCTION..................................................................................................... 1

II. REVIEW OF LITERATURE ................................................................................... 4

Pathology and Treatment of Motor Incomplete Spinal Cord Injuries ......................... 6
Neuropathic pain .................................................................................................... 7
Spasticity ............................................................................................................... 9
Dysautonomia and hemodynamics ....................................................................... 12
Blood pressure regulation and heart rate ............................................................ 12
Blood distribution during exercise ..................................................................... 13
Autonomic dysreflexia ...................................................................................... 14
Motor function ..................................................................................................... 16
Treadmill training ............................................................................................. 17
Over ground walk training................................................................................. 19
Functional electrical stimulation ........................................................................ 20
Blood Flow Restriction Training ............................................................................. 21
Acute responses to blood flow restriction training ................................................ 21
Chronic adaptations to blood flow restriction training .......................................... 25
Potential mechanisms ........................................................................................... 27
Safety issues ........................................................................................................ 29
Need For Research .................................................................................................. 31
Purpose and Hypothesis .......................................................................................... 32

III. METHODS........................................................................................................... 36
Study Design .......................................................................................................... 36
Participants ............................................................................................................. 36
Protocol .................................................................................................................. 37
Experiment 1 ....................................................................................................... 37
Experiment 2 ....................................................................................................... 39
Variables ................................................................................................................ 40
D-dimer ............................................................................................................... 41
Venous blood flow ............................................................................................... 41
Spasticity ............................................................................................................. 42
Neuropathic pain .................................................................................................. 42
Muscle activation ................................................................................................. 42
Perceived effort .................................................................................................... 43
iv
 Blood pressure and heart rate ............................................................................... 43
Whole blood lactate ............................................................................................. 43
Muscle tissue oxygenation ................................................................................... 43
Statistical Analyses ................................................................................................. 44
Experiment 1 ....................................................................................................... 44
Experiment 2 ....................................................................................................... 44

IV. THE FEASIBILITY OF BLOOD FLOW RESTRICTION EXERCISE IN INCOMPLETE

SPINAL CORD INJURED PATIENTS ................................................................. 46
Methods .................................................................................................................. 48
Experimental protocol 1 ....................................................................................... 49
Experimental protocol 2 ....................................................................................... 52
Statistical analysis ................................................................................................ 53
Results .................................................................................................................... 54
Hemodynamics .................................................................................................... 54
Oxyhemoglobin ................................................................................................... 55
Deoxyhemoglobin ................................................................................................ 56
Total hemoglobin ................................................................................................. 57
Tissue oxygenation index ..................................................................................... 57
Protocol 2 ............................................................................................................ 58
Discussion .............................................................................................................. 59
Safety and pain .................................................................................................... 59
Physiological variables ........................................................................................ 61
Limitations........................................................................................................... 63

V. GRANT SUBMISSION: BLOOD FLOW RESTRICTION TRAINING IN INDIVIDUALS

WITH INCOMPLETE SPINAL CORD INJURIES ............................................... 65
Personal Statement .................................................................................................. 65
Positions and Honors .............................................................................................. 66
Contributions to Science ......................................................................................... 66
Additional Information: Research Support .............................................................. 67
Overall Objective .................................................................................................... 68
Specific aims ....................................................................................................... 68
Specific aim 1 ................................................................................................... 68
Specific aim 2 ................................................................................................... 68
Introductory statement and background ................................................................ 69
Protocol 1.......................................................................................................... 69
Protocol 2.......................................................................................................... 70
Preliminary data ................................................................................................... 71
Specific aim 1 ................................................................................................... 71
Specific aim 2 ................................................................................................... 72
Rationale ................................................................................................................ 73
Significance ......................................................................................................... 73
v
 Scientific aim 1 ................................................................................................. 73
Scientific aim 2 ................................................................................................. 74
Relevance to the Neilsen Foundation's mission .................................................... 75
Research Plan ......................................................................................................... 75
Scientific aim 1 ................................................................................................. 75
Scientific aim 2 ................................................................................................. 78
Human subjects .................................................................................................... 79
Subject recruitment and inclusion......................................................................... 80
Data collection ..................................................................................................... 81
Potential risks and protection against risks ........................................................... 83
Potential benefits.................................................................................................. 84
Safety monitoring ................................................................................................ 85
Facilities and Resources .......................................................................................... 86
Major equipment .................................................................................................. 87
Doppler ultrasound ............................................................................................ 87
Near infrared spectroscopy ................................................................................ 87
Beat-by-beat blood pressure monitoring ............................................................ 88
Isokinetic dynamometer .................................................................................... 88
Pneumatic occlusion cuffs ................................................................................. 88
Multi-channel data acquisition system ............................................................... 88
Fitness room and equipment .............................................................................. 89
Other Research Support .......................................................................................... 89

VI. SUMMARY .......................................................................................................... 90

APPENDICES .......................................................................................................... 109

Appendix A: American Spinal Injury Association Impairment Scale ............. 110
Appendix B: Neuropathic Pain Scale ............................................................. 113
Appendix C: Modified Ashworth Scale ......................................................... 116
Appendix D: Craig H. Neilsen Foundation Spinal Cord Injury Research on the
Translational Spectrum Pilot Research Grant Template ................................. 118
Appendix E: Manuscript Abstract ................................................................. 121
Appendix F: Craig H. Neilsen Foundation Grant Submission Budget ............ 124

REFERENCES ......................................................................................................... 126

vi
 LIST OF FIGURES

1. An outline of the study design, including both experimental protocols .............................. 92

2. An outline of the BFR walking protocol ........................................................................... 93

3. Mean arterial pressure (MAP) compared across each experimental trial ........................... 94

4. Mean arterial pressure (MAP) compared across each acute exercise bout ......................... 95

5. Systolic blood pressure (SBP) compared across each experimental trial ........................... 96

6. Systolic Blood pressure (SBP) compared across each acute exercise bout ......................... 97

7. Oxyhemoglobin (Oxy) compared across each experimental trial and between conditions . 98

8. Oxyhemoglobin (Oxy) compared across each acute exercise bout and between conditions

................................................................................................................................. 99

9. Deoxyhemoglobin (Doxy) compared across each experimental trial and between

conditions ................................................................................................................. 100

10. Deoxyhemoglobin (Doxy) compared across each acute exercise bout and between

conditions ................................................................................................................. 101

11. Total hemoglobin compared across each experimental trial and between conditions ....... 102

vii
12. Total hemoglobin compared across each acute exercise bout and between conditions ..... 103

13. Tissue oxygenation index (TOI) compared across each experimental trial and between

conditions ................................................................................................................. 104

14. Tissue oxygenation index (TOI) compared across each acute exercise bout and between

conditions ................................................................................................................. 105

15. A timeline of protocol 1.................................................................................................. 106

16. A timeline of protocol 2.................................................................................................. 107

viii
 CHAPTER I

INTRODUCTION

Spinal cord injuries are common neurological disorders that disrupt the

transmission of a motor signal from the brain to the periphery, or the afferent signal from

the periphery to the brain. This causes motor dysfunction, paralysis, autonomic

disorders, and other symptoms that impact daily activities and quality of life (Kawanishi

& Greguol, 2013). The severity of a spinal cord injury (SCI) depends on the location of

the injury along the vertebral column and the completeness of the injury. In some cases,

individuals can retain near-complete motor and sensory function, while others may lose

total function below the neck. An incomplete spinal cord injury (iSCI) is the former,

defined by residual sensory or motor function at the lower portion of the spinal cord

(Roberts, Leonard, & Cepela, 2016). In many of these cases patients also retain some

degree of ambulation, but with a high degree of variability between patients.

Rehabilitation programs for individuals with iSCI are heavily focused on restoring as

much motor function, specifically locomotor function, as possible (Dietz & Fouad, 2014).

Physical therapy programs for this population also focus on increasing muscular strength,

muscular endurance, work capacity; all with the ultimate goal of improving quality of life

and obtaining (or maintaining) independence (Kozlowski & Heinemann, 2013).

Researchers and practitioners are still developing new training methods that can

potentiate the rehabilitation of an incomplete spinal cord injury.

Blood flow restriction (BFR) training is a form of exercise training that uses

external pressure to occlude venous flow from, and often arterial flow to, the exercising

1
 2

muscle. This type of training has been shown to increase muscular strength, endurance,

elicit hypertrophy, and even increase aerobic exercise capacity at very low (20-30%

maximal exertion) intensities (Abe et al., 2010; Abe, Kearns, & Sato, 2006; Barcelos et

al., 2015). These results have been observed in normal healthy adults, athletes, elderly

individuals, and in only one case individuals with SCI (Gorgey et al., 2016). The

proposed mechanisms for this effect include metabolite accumulation during exercise,

preferential type II fiber recruitment, hypoxemia induced adaptations to mitochondrial

volume and density, elevated growth hormone, and activation of myogenic cells (Goto,

Ishii, Kizuka, & Takamatsu, 2005; Kon, Ikeda, Homma, & Suzuki, 2012; Lang et al.,

1998; Loenneke, Fahs, Wilson, & Bemben, 2011; Meyer, 2006; Takarada, Nakamura, et

al., 2000). While the addition of blood flow restriction training to the rehabilitation

program of an individual with iSCI might present some challenges, the potential benefits

could be substantial. Improving muscular fitness and aerobic capacity in this population

may lead to enhanced motor function and prolonged independence. Therefore, the

purpose of the proposed study is to determine the feasibility and safety of performing

BFR exercise in individuals with iSCI. Furthermore, this study aims to examine the

potential efficacy of BFR in this population by comparing the acute physiological

responses of BFR exercise to exercise without BFR. Results from this study may support

future research examining the outcomes of regular BFR training for individuals with

iSCI.

This study will include 2 separate experiments, each designed to answer one of

the research aims listed above. Data were collected on 9 subjects with chronic (>6
 3

months) iSCI for each experiment. Experiment 1 compared a trial of knee extensor

exercise with BFR to a trial of the same exercise without BFR; and Experiment 2

compared a trial of unassisted walking with BFR to a trial of walking without BFR. We

hypothesize that subjects will be able to perform both walking and knee extensor exercise

with BFR with only moderate reductions in repetitions performed and time walked

compared to control exercise. We also hypothesize that subjects will not exhibit any

signs of Autonomic Dysreflexia (AD) or development of Deep Vein Thrombosis (DVT)

due to BFR exercise; and that the acute physiological responses to BFR exercise (e.g.

increased EMG activity with submaximal exercise and elevated lactate accumulation)

will be similar in this sample to those published in previous studies examining the able-

bodied population.
 CHAPTER II

REVIEW OF LITERATURE

A spinal cord injury (SCI) is a neurological condition caused by damage to the

spinal cord, and can be due to trauma or disease. The effects of a SCI depend on the

location and completeness of the injury, and can range from total loss of sensory and

motor function below the neck to almost no loss in sensory or motor function (Roberts et

al., 2016). It is estimated that 282,000 people in the United States are currently living

with a SCI and there are approximately 17,000 new cases of SCI each year (National

Spinal Cord Injury Research Center [NSCIRC]), 2016). The average cost during the first

year following a SCI can range from $347,896 for someone with low level motor-

incomplete SCI to $1,065,980 for someone with a high level motor-complete SCI; and

recurring annual cost can range between $42,256 and $185,111 for the same injuries.

More importantly, life expectancy is reduced following SCI and is negatively correlated

with degree of injury and age of SCI diagnosis (NSCIRC, 2016). Fortunately, recent

medical advances have started to improved life expectancy during the first 2 years of SCI

(Strauss, Devivo, Paculdo, & Shavelle, 2006).

Classifying the degree of injury and identifying its location along the spinal cord

first requires an understanding of spinal cord anatomy. The spinal cord descends through

the vertebral column beginning with the cervical region and terminating in the lumbar

region (conus medullaris). Spinal nerves, which are comprised of a ventral (efferent) and

dorsal (afferent) root, exit the spinal cord between vertebrae via the intervertebral

foramina. There are 8 spinal nerves that exit the cervical region, 12 in the thoracic

4
 5

region, and 5 in the lumber region. The cauda equina is a junction of spinal nerves that

exits through sacral foramina below the level of the conus medullaris. Each of these

spinal nerves is innervated by specific dermatomes (sites on the skin that trigger sensory

input), and innervate a number of muscles, collectively referred to as a myotome. While

most dorsal roots are innervated by specific dermatomes, a single muscle or muscle group

may be innervated by more than one spinal neuron. The neural level of injury (NLI) can

then be diagnosed by identifying the effected dermatomes and myotomes (Roberts et al.,

2016). The two most basic classifications of NLI are tetraplegia and paraplegia. Damage

to the spinal nerves of the cervical spine is classified as tetraplegia (a.k.a. quadriplegia).

In tetraplegia, function of the legs, trunk and arms are affected. The degree to which these

limbs are affected can vary based on the completeness of the injury. Paraplegia is

characterized by damage to the spinal cord in the thoracic or lumbar regions, manifesting

as loss of function in the legs, and can also include loss of function in the trunk (Roberts

et al., 2016). It is important to note that damage to peripheral nerves (such as the brachial

plexus) is not considered in the diagnoses or classification of a spinal cord injury (i.e.

level and completeness). While tetraplegia and paraplegia are broad categories of the

NLI, the American Spinal Injury Association (ASIA) Impairment Scale is a more

comprehensive tool for assessing NLI and completeness of injury (Appendix A). The

ASIA impairment scale tests voluntary motor function on a 0 (total paralysis) to 5

(normal function) and sensory function on a 0 (no sensory input) to 2 (normal sensory

input) scale to determine level and completeness of injury for the right and left sides.

The most rostral level of injury is selected as the overall NLI (Kirshblum et al., 2011). If
 6

a patient has residual motor or sensory function below the NLI including the sacral region

(specifically S4-S5), their injury is classified as incomplete. An injury without any

sensory or motor function at the most caudal regions (specifically S4-S5) is classified as

complete. This study will primarily focus on those individuals with incomplete SCIs

(iSCIs).

Pathology and Treatment of Motor-Incomplete Spinal Cord Injuries

Incomplete spinal cord injuries are estimated to account for 58.3% (45%

incomplete tetraplegia and 13.3% paraplegia) of all diagnosed SCI's upon hospital

discharge (NSCIRC, 2016). A study by Lee, Leiby, and Marino (2016) examined the

recovery following traumatic SCI and found that 15.5% of patients who were initially

classified as ASIA-A (motor-complete) eventually recovered enough to be reclassified as

motor-incomplete. The same study reported differences in functional scores (Function

Independence Measure [FIM]) and recovery of lower extremity motor function between

NLI’s in a sample of paraplegics, with lower NLI’s showing a greater recovery in lower

extremity motor scores and having a higher functional score after 1 year. Another study

by Kirshblum et al. (2016) examined the efficacy of rehabilitation during the acute phase

of an SCI and found that 20% of ASIA-A (See Appendix A for ASIA scale) diagnoses

were reclassified as motor-incomplete upon discharge, which increased to 27.8% after 1

year; and 33.9% of ASIA-B diagnoses were reclassified as motor incomplete upon

discharge, which increased to 53.6% after 1 year. These data suggest that rehabilitation

following SCI diagnosis can have profound effects on the overall function of an

individual with a SCI. That being said, there are a number of pathologies that result from
 7

a SCI that need to be considered in the rehabilitation program. These pathologies are

discussed below.

Neuropathic pain

Neuropathic pain occurs at or below the NLI, and is caused by damage to the

spinal cord or neural tract, contrary to nocioceptive pain which is caused by agitation of

tissues that provide sensory input to the CNS (D'Angelo et al., 2013). There is no

consensus regarding the estimation of the prevalence of neuropathic pain among

individuals with SCI. These estimates can range from 11% of SCI’s presenting with

below-NLI pain and 24% presenting at-NLI pain (Barrett, McClelland, Rutkowski, &

Siddall, 2003), to combined estimates of 92% of individuals with SCI presenting with

neuropathic pain (Adriaansen et al., 2013). This variance may be due to the subjective

nature of reporting pain, the variation in the populations studied (injury level, time after

injury, etc.), or the variation between the pathologies of each specific injury.

Neuropathic pain treatment is primarily pharmacological. Drugs such as antidepressants

(Bohren et al., 2013), anticonvulsants (Camara et al., 2015), selective serotonin reuptake

inhibitors (Saito, Wakai, Sekiguchi, Kikuchi, & Konno, 2014), capsaicin (Trbovich &

Yang, 2015), non-steroidal anti-inflammatories (Shinozaki, Yamada, Nonaka, &

Yamamoto, 2015), and in severe cases Opioids (Bostick et al., 2015) are used to treat

pain in individuals with iSCI. Opioids are used sparingly due to the well documented

issues with addiction (Juurlink & Dhalla, 2012).

Traditional physical rehabilitation modalities are generally considered ineffective

in treating neuropathic pain (Akyuz & Kenis, 2014; Fattal, Kong, Gilbert, Ventura, &
 8

Albert, 2009). Therefore, researchers and clinicians have developed techniques that

integrate physical rehabilitation with neurological conditioning. Villiger et al. (2013)

examined the effect of virtual-reality augmented training on neuropathic pain

management in a group of individuals with iSCI, 9 of which presented with neuropathic

pain. This virtual reality system used movement sensors attached to the foot and calf of

each leg to control a video game presented on a screen. Each subject played 4 different

games which included ankle dorsiflexion (to help prevent foot-drag), knee extension, and

external rotation of the leg. In each of the games a pair of virtual legs was displayed on

the screen which represented the subjects’ legs and reacted to the movement of the calf

and foot. Each of these training sessions lasted approximately 45 minutes and was

performed 4-5 times per week for 4 weeks. After the training period, 6 out of the 9

patients presenting with neuropathic pain had clinically improved Neuropathic Pain Scale

(NPS, Appendix B) scores (6.6±1.2 at baseline and 3.9±1.8 post-training for intensity,

and 8.0±1.1 at baseline and 4.7±1.8 post-training for unpleasantness). Another study by

Jordan and Richardson (2016) also tested the effects of visual-illusory based

neurorehabilitation on neuropathic pain in SCI with a focus on the differences in location

of pain (above NLI, at NLI, and below NLI). This study also used quantitative sensory

testing (QST) to better determine the effect of virtual-reality augmented

neurorehabilitation on neuronal hypersensitivity, and its relationship to neuropathic pain.

The visual-illusory stimulus required subjects to watch 2 different programs, one of a

person walking along a path and another of a person manually propelling a wheelchair

along a path; and they were to imagine themselves performing these actions. Both of
 9

these videos were presented on a 3 dimensional display for a period of 20 minutes.

Results indicated that pain, independent of location, was significantly reduced following

the virtual walking condition (-1.58±1.62 at-level, and -0.78±1.51 below-level) compared

to the wheelchair condition (p<0.05). Results also indicated that at-NLI sensitivity was

positively correlated to below-level neuropathic pain following the virtual walking

stimulus (p<0.05). These data suggest that sensory enhanced (either through virtual

reality or visual-illusory) neurological rehabilitation can improve neuropathic pain in

iSCI subjects.

Spasticity

Spasticity is an involuntary muscular contraction commonly observed in SCI and

other neurological disorders (i.e. multiple sclerosis, Parkinson’s disease, etc.). Spasticity

can be present during rest (often hypertonia) or during movement (hypertonia and

dyssynergia). This can effect rehabilitation by limiting range of motion, and has been

identified as a significant limitation to activities of daily living (van Cooten, Snoek,

Nene, de Groot, & Post, 2015). The same study also observed the presence of spasticity

to be higher in tetraplegia compared to paraplegia. Mild spasticity may also be beneficial

for rehabilitation if it occurs synergistically with the desired range of motion, which can

benefit transfers or activities of daily living. Spasticity is assessed using the Modified

Ashworth Scale (MAS; appendix C). The MAS is a 9-point scale that evaluates muscle

tone at rest and resistance through the full range of motion, and has been reported to have

good test re-test and interrator reliability in individuals with any form of SCI (Li, Wu, &

Li, 2014). The Tardieu Scale, another method of evaluating spasticity in SCI, was shown
 10

to be less reliable in the same study. Pharmacological treatment of spasticity includes

gamma-aminobutryic acid agonists (GABA) (Heetla, Proost, Molmans, Staal, & van

Laar, 2016) and alpha-2 agonists (Malanga, Reiter, & Garay, 2008). One issue with these

medications is that they impair movement, which itself is contradictory to physical

rehabilitation efforts.

Physical rehabilitation is also used to treat spasticity and would be preferential to

pharmacological interventions. These rehabilitation modalities include passive, active,

and assisted range of motion exercises (Krause, Szecsi, & Straube, 2008; Rayegani et al.,

2011), vibration therapy (Murillo et al., 2011), and transcutaneous nerve stimulation

(Laddha, Ganesh, Pattnaik, Mohanty, & Mishra, 2015). Tashiro et al. (2015) examined

the effects of treadmill training on spasticity in a sample of spinal cord injured rats, and

aimed to explain the mechanisms that mediate the improvement in spasticity. Following

2 weeks of daily body-weight supported treadmill exercise the spasticity of treadmill-

trained rats was reduced, and was far lower that the spasticity in a group of control-rats

who did not undergo treadmill training. Results also indicated that upregulation of Brain

Derived Neurotrophic Factor (BDNF) in the lumbar spine, which was purported to

increase expression of potassium-chloride transporter member 5 (KCC2; controls the

neuronal chloride ion gradient), and KCC2 expression was significantly correlated to

reductions in spasticity. A study by Adams and Hicks (2011) examined the effect of

body weight supported treadmill training and tilt table standing in a group of spinal cord

injured humans (n=7). Subjects in this study ranged from ASIA A to ASIA C, and

spasticity was assessed via the MAS at baseline, after a single session, and after 4 weeks
 11

of training in each condition. Results indicated that flexor spasms (ES=0.57) and passive

resistance to movement (ES=0.69) were lower following a single bout of treadmill

exercise compared to tilt table standing. Results also suggested that treadmill training

was more beneficial than tilt table standing after 4 weeks of training, and that tilt table

standing was more beneficial for extensor spasms than body weight supported treadmill

training (ES=0.68 and 1.32 for single session and 4 weeks respectively). There is limited

research that examines the effect of treadmill exercise independent of other modalities or

therapies on spasticity alone in individuals with SCI. This is likely due to a greater focus

on the functional outcomes (e.g. walking speed, motor scores, etc) of treadmill exercise.

Transcutaneous electrical nerve stimulation (TENS) involves passing an electrical

current through a target nerve via surface stimulation. Ping Ho Chung and Kam Kwan

Cheng (2010) tested the effects of 60 minutes of TENS and 60 minutes of sham TENS

(control) over the common peroneal nerve on spasticity in the ankle dorsiflexors. The

TENS condition elicited a 29.5% reduction in Composite Spasticity Score (a score that

accounts for tendon jerks, resistance to range of motion, and clonus), a 31% reduction in

the resistance to passive ankle dorsiflexion, and a 29.6% reduction in clonus. These

improvements were not observed in the control group who did not receive TENS.

Another study by Laddha et al. (2015) examined the effects of 6 weeks of either 30 or 60

minute sessions of TENS over the common peroneal nerve compared to 6 weeks of task-

oriented exercise training on the spasticity of the ankle plantar flexors. Results indicated

that all three groups improved spasticity, clonus, and Timed Up and Go scores (a measure
 12

of functional ability), and the improvements in spasticity were significantly greater in the

TENS groups (p<0.05).

Dysautonomia and hemodynamics

Dysautonomia is a condition characterized by dysfunction of the autonomic

nervous system. This often leads to abnormal blood pressure regulation, blood flow

distribution, and heart rate during rest and exercise (Bell, Chen, Bahls, & Newcomer,

2013; Krstacic, Krstacic, & Gamberger, 2013). For individuals with SCI, dysautonomia

is highly dependent on the level and completeness of the injury (Zhu, Galea, Livote,

Signor, & Wecht, 2013).

Blood pressure regulation and heart rate. In an individual with a SCI, the

global sympathetic vasoconstriction that occurs in response to exercise, specifically in

response to increases in central command and the activity of Group III and IV muscle

afferents is attenuated or absent below the NLI. Furthermore, how this effects heart rate

and mean arterial pressure also depends on the level of injury. A study by Wecht et al.

(2013) examined the prevalence of blood pressure and heart rate abnormalities in a group

of tetraplegics, high-level paraplegics, and low-level paraplegics. Results indicated that

bradycardia was more common in the tetraplegic group, while tachycardia was more

common in both of the paraplegic groups; systolic hypotension was more common in the

tetraplegic and high-paraplegic groups compared to the low-paraplegic group; and

systolic hypertension was far more common in the low-paraplegic group. Furthermore,

orthostatic intolerance was more apparent in the tetraplegic group. These data illustrate

that blood pressure regulation is impaired to a greater extent the higher the injury. In
 13

contrast, these data also illustrate the compensatory increase in heart rate observed in

low-level paraplegics, likely due to the sympathoinhibition below the NLI leading to a

reduced venous return and stroke volume. This last point is corroborated by Currie,

West, and Krassioukov (2016) who reported a significantly lower average stroke volume

and cardiac output among Paralympic athletes with tetraplegia compared to paraplegia

(p<0.05).

Blood distribution during exercise. This reduction in sympathetic

vasoconstriction also impairs redirection of blood flow to active tissue during exercise

(i.e. to the arms during an arm cranking exercise), ultimately reducing oxygen delivery

and work capacity (Theisen, 2012; Thijssen, Steendijk, & Hopman, 2009). Hostettler et

al. (2012) compared stroke volume and cardiac output responses to arm crank and wheel

chair exercise in a group of cervical spinal cord injured males compared to able bodied

males, and observed no significant increase in stroke volume during either exercise in the

cervical SCI group. Cardiac output did increase in the cervical SCI group, but remained

significantly lower than the able bodied individuals. Vaile, Stefanovic, and Askew

(2016) attempted to circumvent this by applying compression to the lower limbs during

exercise. In this study 10 male wheelchair rugby athletes with cervical SCI's performed

two separate exercise sessions, one while wearing medical grade compression gear and

the other with no compression gear. Cardiac output was not measured in this study, but

the authors did report a greater increase in upper limb blood flow (+10.77±8.24 mL*min-
1
and 6.21±5.73 mL*min-1 for upper and lower limb blood flow, respectively) from pre to

post exercise when compression gear was worn compared to the control session. This
 14

suggests that the pressure applied to the legs compensated for the lack of venous tone and

restricted excess venous distension, likely leading to an increase in stroke volume. In

addition to the effect of sympathoinhibition on blood pressure regulation and blood flow,

there is also evidence that peripheral factors such as sympathetic reflex activity

(Rummery, Tripovic, McLachlan, & Brock, 2010) and the nitric oxide pathway (Brown,

Celermajer, Macefield, & Sander, 2016; Venturelli et al., 2014) are modulators of

vascular function below the NLI.

In relation to impaired blood distribution during exercise; immobility of the lower

limbs during period of inactivity leads to venous hemostasis, which can result in the

formation of a deep vein thrombosis (DVT). DVTs can eventually dislodge and travel to

the heart or lungs, which may result in myocardial infarction or pulmonary embolism.

Evidence suggests that individuals with higher NLIs are at a higher risk of forming a

DVT (Waring & Karunas, 1991). Individuals at high risk for DVTs are generally treated

with anticoagulant and antiplatelet medications, and are encouraged to avoid long periods

of inactivity.

Autonomic dysreflexia. The maintenance of peripheral control of vascular

function combined with supraspinal inhibition can elicit a potentially dangerous

phenomenon, termed autonomic dysreflexia (AD). AD is a condition associated with

dysautonomia, and is characterized by a pathological increase in blood pressure and a

compensatory drop in heart rate. AD is caused by a noxious or painful stimulus below

the NLI that initiates a reflex arc with the intact CNS, resulting in sympathetic

vasoconstriction. This vasoconstriction begins to increase blood pressure, activating the

 15

carotid baroreceptors. This activates the pressure control centers of the brain that send an

inhibitory signal down the CNS to reverse the reflex vasoconstriction and simultaneously

reduce heart rate. However, the inhibitory signal never passes through the NLI, allowing

blood pressure to continue to rise while heart rate decreases. Common stimuli that elicit

AD are bladder distension and fecal impaction. In extreme cases, athletes with a SCI

may intentionally elicit AD for the performance enhancing effect of increasing blood

pressure. Termed “boosting”, this is a method of circumventing the impairment in blood

delivery to exercising muscle observed in athletes with SCI; however, it is prohibited by

the International Paralympic Committee (IPC). One method of preventative treatment for

AD is bladder denervation. In a study by Hohenfellner et al. (2001) 8 subjects with

diagnosed detrusor hyperreflexia or AD underwent bladder denervation, and symptoms

were eliminated in each subject. Other preventative or long-term treatments include

ventral sacral root stimulation (to improve voiding which is a cause of AD),

pharmacotherapy (i.e. Terazosin) (Vaidyanathan et al., 1998), and educating patients on

avoiding potentially noxious stimuli. Acute treatment of AD includes identifying the

noxious stimulus and removing it if possible, sitting the patient in an upright position,

administration of fast-acting antihypertensives such as nifedipine or nitroglycerin paste,

and monitoring the patient’s blood pressure and heart rate continuously until they return

to normal (Consortium for Spinal Cord Medicine, 2001). There is also evidence to

suggest that risk of AD is lower in individuals with low-level iSCI compared to with

high-level complete SCI (Helkowski, Ditunno, & Boninger, 2003).

 16

Motor function

Motor function is directly impacted following a SCI. Supraspinal muscle

activation is impaired which reduces force output and motor recruitment. As is the case

with most of the effects of an SCI, the impact on motor function is dependent on the NLI

and completeness of the injury. The severity of an iSCI can range from significant

functional impairment to almost no impairment (Roberts et al., 2016). A study by

Fehlings and Tator (1995) used a rat-model of SCI to examine the effect of injury

severity on residual neurological function an axon counts. To control for injury severity,

investigators used 5 different compression forces to administer clip injuries in 30 adult

female Wistar rats, and compared results between each compression force and to a

control group (only received a laminectomy). Results from the study showed that

neurological function (as assessed by the inclined plane method) and axon count were

negatively and linear correlated to the compression forces used in the clip injuries (-0.92,

p<0.001). Neurological function was also significantly positively correlated to axon

counts, and the integrity of the rubrospinal, vestibulospinal and raphespinal nonpyramidal

tracts. Another study by Kim, Eng, and Whittaker (2004) tested the relationships

between functional ability (gait speed, 6-minute walk test distance, and ambulation) and

strength of the muscles acting on the hips, knees, and ankles. Results from this study

indicated that the strength of the muscles acting on the hip was significantly correlated to

functional outcomes. Results also indicated that the hip flexors explained the most

variance in gait speed, and the hip extensors explained the most variance in ambulatory
 17

capacity. These results illustrate the importance of maintaining muscular strength of the

muscles acting on the hip.

Treadmill training. Therapist-assisted treadmill training is a commonly used

modality in SCI rehabilitation. However, more novel approaches to locomotor training

have been developed in recent years. Stevens, Caputo, Fuller, and Morgan (2015)

examined the effect of an 8 week, 3 times per week underwater treadmill training

program on motor performance and muscular adaptations in 11 subjects with iSCI. In

this protocol water was used as an unweighting system allowing subjects to balance

during each walking trial, eliminating therapist-fatigue as a limiting factor. Throughout

the 8-week intervention walking speed was increased by 10% biweekly. After the

intervention, six-minute walk test distance (97m ± 80 to 177m ± 122) and daily step

activity (593 ± 782 to 1310 ± 1258) significantly improved, as did lower-extremity leg

strength, balance, and walking speed. Stevens and Morgan (2015) also observed a

decrease in heart rate during submaximal exercise following 8 weeks of underwater

treadmill training, pointing to possible cardiovascular benefits.

Another method of treadmill training is body weight supported treadmill training

(BWSTT). BWSTT uses a suspension system to support a predetermined percentage of

the patient's body weight. This type of training has been shown to improve six-minute

walk test distance and gait function in individuals with iSCI (Leahy, 2010; Lucareli et al.,

2011), and in some cases increase EMG activity through sensory feedback (Dobkin,

Harkema, Requejo, & Edgerton, 1995). A study by Giangregorio et al. (2005) examined

the effect of a 24-week body weight supported treadmill training program on bone
 18

mineral density (BMD) and muscle cross sectional area (CSA) in a sample of 5 subjects

with iSCI. BMD of the proximal femur, distal femur, proximal tibia, and lumbar spine,

and muscle CSA at the mid-femur were compared at baseline, and after 24 weeks of

training. Results from this study indicated that muscle CSA increased between 3.8% and

56.9% in all subjects. However, BMD decreased at all sites following 24 weeks of

training, ranging from -1.2% to 26.7%. These data suggest that body weight support may

be a sufficient stimulus to attenuate, and even reverse, muscular atrophy; but not to

attenuate losses in BMD.

A third form of treadmill training is robotic assisted treadmill training RATT,

which uses mechanical exoskeletons that support the weight of the subject while also

assisting through the range of motion. Similar to underwater treadmill training and

BWSTT, RATT has been shown to improve walking speed, gait, and motor function in

individuals with iSCI (Fleerkotte et al., 2014). RATT has also been shown to improve

cardiorespiratory endurance in some cases. Gorman et al. (2016) examined the effect of 3

months of RATT compared to an at-home stretching program on peak oxygen

consumption (Vo2peak) during robotic assisted treadmill walking and arm crank

ergometry separately. Subjects were divided into two groups, one that participated in the

RATT program first, and one that participated in the at-home stretching program first.

After the first 3 months, the group that had participated in the stretching program was

then transitioned to the RATT program. Vo 2peak was assessed at baseline, after 6 weeks

of training, and after training. Results indicated that the RATT program significantly

improved Vo2peak during that same modality, but not during arm-crank exercise. These
 19

results suggest that improvements in locomotor function, muscular adaptations, or

peripheral factors in the legs following RATT can lead to enhanced cardiorespiratory

fitness level during a walking task. This is supported by Sczesny-Kaiser et al. (2015)

who reported improvements in 10 minute walk test speed (0.25 m*s -1 ± 0.05 to 0.5 m*s-1

± 0.07), 6 minute walk distance (86m ± 20.86 to 149.73m ± 20.32), Timed Up and Go

test score (56.35 ±10.06 to 38.65 ±7.2), and cortical excitability following a 3 month

RATT program.

Over ground walk training. Over ground walk training differs from treadmill

training by requiring the subject to move the body through space, as opposed to

supporting and stabilizing the body during locomotor activity. This presents unique

challenges to individuals with iSCI. Hayes, Chvatal, French, Ting, and Trumbower

(2014) examined the motor complexity of the lower body during over ground walking in

subjects with iSCI compared to able bodied control subjects. The investigators

specifically examined the number of motor modules activated and level of activation

between the two groups at different cadences, and found that individuals with iSCI had

significantly less active motor modules during walking compared to control subjects.

Module composition and activation were also altered in individuals with iSCI, indicating

that muscle coordination is impaired following iSCI. As with treadmill training, over

ground walking can be performed with therapist-assistance, a body weight support

system, or with an exoskeleton. The benefits of over ground walking are similar to those

observed with treadmill walking (Del-Ama, Gil-Agudo, Pons, & Moreno, 2014;

Senthilvelkumar, Magimairaj, Fletcher, Tharion, & George, 2015).

 20

Functional electrical stimulation. Functional electrical stimulation (FES)

training is a training rehabilitation method that uses external electrical impulses to

stimulate movement in the paralyzed limbs. In most cases the muscle is stimulated

through the peripheral nerve (as opposed to the muscle itself) and in a sequence that best

matches the natural muscular contractions during a pedaling or walking motion.

Thrasher, Ward, and Fisher (2013) tested the effect of a 40-session FES cycling

intervention on fatigue and power output in a group of subjects with complete and

incomplete SCI. In this study, 11 subjects completed 40 hours of FES cycling at a

cadence of 45 rpm and a constant resistance. Mean power output was recorded

throughout each cycling trial, and fatigue index and knee extension peak torque were

assessed at baseline and after the trial. Results indicated that mean power output

increased throughout the training program, peak knee extensor torque significantly

increased from pre to post, and fatigue index was significantly reduced from pre to post

in subjects with iSCI. Another study by Yasar, Yilmaz, Goktepe, and Kesikburun (2015)

reported similar findings when they observed significant improvements in total motor

scores (79.9 ± 11.8 to 84.6 ± 8.6, pre to post, respectively), FIM scores (109.8 ± 19.9 to

116.5 ± 13.3), and MAS scores (2.1 ± 0.3 to 0.9 ± 0.7 for the rectus femoris and 1.7 ± 0.4

to 0.7 ± 0.4 for the hamstrings) in a group of subjects with iSCI after a 6 month, 3 times

per week FES cycling program. Ultimately, these studies and others support the use of

FES training as part of a rehabilitation program for individuals with iSCI (Fornusek,

Davis, & Russold, 2013; Kuhn, Leichtfried, & Schobersberger, 2014; Sadowsky et al.,

2013).
 21

Blood Flow Restriction Training

Blood flow restriction (BFR) training, also known as ischemic or Kaatsu training,

is a method of training in which pressure is used to occlude venous flow and reduce

arterial flow to working muscles (Libardi et al., 2015). One of the theories behind this

form of training is that trapped metabolites begin to accumulate in the working muscle

and after repeated exposure elicit the same muscular adaptations known to result from

high intensity exercise (Goto et al., 2005; Loenneke, Fahs, et al., 2011). Others have

postulated that the muscular benefits of BFR training are due to reactive hyperemia,

preferential recruitment of type II fast twitch muscle fibers, hypoxemia induces changes

to anaerobic and aerobic metabolism, and activation of myogenic stem cells (Kon et al.,

2012; Lang et al., 1998; Meyer, 2006; Takarada, Nakamura, et al., 2000). The use of

BFR training has expanded over many populations, including healthy adults, the elderly,

and even those with neurological disorders (Gorgey et al., 2016; Salvador et al., 2016);

and has consistently promoted improvement in muscular strength, size, and function. The

following sections will discuss BFR training, primarily in individuals without a SCI.

Acute responses to blood flow restriction training

Light intensity resistance training with BFR has been shown to elicit similar acute

responses to high intensity resistance exercise without BFR, and in some cases these

responses are even augmented in BFR exercise. For example, Reeves et al. (2006)

compared the hormonal responses to light intensity (30% 1RM) BFR training, traditional

exercise (70% 1RM) and occlusion without exercise in a group of healthy individuals. In

both exercise sessions subjects performed 3 sets of unilateral bicep curls to failure
 22

followed by 3 sets of unilateral knee extensions to failure. In the occlusion only session

subjects rested while the arm and leg were occluded for the same time period as in the

previous 2 sessions. An occlusion pressure of 20 mmHg less than the systolic blood

pressure was used for the arm, and 40 mmHg was added to that pressure for leg

occlusion. Results indicated that lactate accumulation was significantly elevated

following the light BFR condition and the traditional exercise condition, but was not

different between groups. Growth hormone significantly increased in the light BFR

condition, but did not change in the traditional exercise or occlusion only conditions.

This increase in growth hormone following light BFR resistance exercise has also is

supported by other investigations (Madarame, Sasaki, & Ishii, 2010; Sato, Yoshitomi, &

Abe, 2005), as is the increase in lactate accumulation (Loenneke et al., 2016; Takarada,

Nakamura, et al., 2000) and in some cases protein synthesis rate (Fujita et al., 2007).

Surface integrated EMG (iEMG) has also been observed to increase during light-

BFR exercise compared to non-BFR exercise in healthy adults. Yamada et al. (2004)

examined iEMG during BFR exercise of varying intensities in a group of six young

healthy males. Subjects performed unilateral knee extension at 30%, 50%, and 70%

maximal voluntary contraction (MVC) with and without 100 mmHg of occlusion

pressure. Results indicated that iEMG and mean frequency were greater during 30% and

50% 1RM with occlusion than without occlusion. Increases in muscle activation during

BFR exercise were also observed by Fatela, Reis, Mendonca, Avela, and Mil-Homens

(2016) during low intensity knee extensions (28.1% increase in RMS of the rectus

femoris with 80% arterial occlusion). Yasuda et al. (2009) compared muscular activation
 23

between moderate blood flow restriction (160 mmHg), complete blood flow occlusion

(300mmHg), and no blood flow occlusion during bicep curls. A group of healthy

subjects performed 3 different exercise trials. The first trial included 30 consecutive

contractions, the second trial included 3 sets of 10 contractions, and the third trial began

with 30 consecutive contractions and continued to 3 sets of 15 contractions. In all 3

experimental trials, 20% of the 1RM was selected as the workload. In the first two

experimental trials, muscle activation progressively increased in moderate (by

approximately 200 - 300 µV*sec-1) and complete (by approximately 300 µV*sec-1)

occlusion to a greater extent than without occlusion; and MVC was reduced more with

complete occlusion compared to moderate occlusion. In the third experimental trial,

fatigue index and muscular activation were greater in both the moderate and completely

occluded conditions, and were comparable between the two. However, less work was

performed in the occlusion condition due to the subjects' inability to complete the

protocol with 300 mmHg. The authors suggested that this was a result of a mismatch

between an extreme increase in muscular activation (iEMG) and a significant reduction in

energy supply (blood flow), which would ultimately inhibit muscular work. However,

that can only be speculated as the investigators were unable to record muscular activation

during the complete occlusion protocol. The argument could also be made that exercise

during very high occlusion pressures (250 mmHg and more) are often limited by

mechanical restraints and discomfort. This area of BFR research, specifically the effect

of different occlusion pressures, should be expanded in future studies. However, with

that stated, these results do suggest that moderate blood flow occlusion can be sufficient
 24

enough to elicit increases in muscular activation while maintaining the ability to perform

the exercise.

Hemodynamic responses such as elevated heart rate, mean arterial pressure,

systolic blood pressure, and decreased stroke volume have also been observed during

BFR resistance training and treadmill walking (Staunton, May, Brandner, & Warmington,

2015; Takano, Morita, Iida, Kato, et al., 2005); and oxygen consumption has been shown

to be higher during submaximal aerobic exercise with BFR than without (Sakamaki-

Sunaga, Loenneke, Thiebaud, & Abe, 2012). A study by Sugawara, Tomoto, and Tanaka

(2015) examined the blood pressure, heart rate, stroke volume, and cardiac output

responses to a single session of BFR walking. In this study, 15 adults completed 2

separate exercise session, each consisting of 5 bouts of 2 minutes of treadmill walking at

2 mph. In one session blood flow was occluded by applying 160 mmHg of external

pressure around the proximal portion of the thigh, and the other session was performed

without BFR. Beat by beat blood pressure, total peripheral resistance (TPR), heart rate,

stroke volume, and cardiac output were collected throughout each exercise session.

Results indicated that heart rate significantly increased during exercise in both conditions,

but increased more during BFR walking. The same result was observed with systolic

blood pressure (43% ± 5 and 11% ± 4 for BFR walking and non BFR walking,

respectively), and the increases in stroke volume and TPR were lower during BFR

walking. These results suggest that BFR elicits a tachycardic response that can drive

blood pressure during a period of lower peripheral resistance.

 25

Chronic adaptations to blood flow restriction training

Regular resistance training with blood flow restriction has been shown to elicit

muscular improvement that can enhance daily function and potentially prolong

independence in some populations. Shinohara, Kouzaki, Yoshihisa, and Fukunaga

(1998) examined the effect of 4 weeks of repeated isometric BFR knee extensor exercise

on MVC and rate of torque development. Five healthy subjects performed single-leg

isometric BFR exercise 3 days a week for four weeks, and each session included 3

minutes of 2-second isometric contractions with 3 seconds of relaxation between

repetitions. The training load was set at 40% of the baseline MVC. The opposite leg

performed the same exercise, only without BFR. Following the 4-week training period,

MVC increased by 26% in the BFR trained leg, and rate of torque development also

increased following BFR training. Increases in MVC and rate of torque development

were significantly greater than increases observed in the control leg. Takarada, Tsuruta,

and Ishii (2004) also observed increases in isometric and isokinetic torque, along with

significant increases in muscle cross sectional area after 8 weeks of very light intensity

(20% MVC) bilateral knee extensor exercise with BFR. Likewise, Ishii, Madaame,

Odagiri, Nagunama, and Sinoda (2005) observed significant increases in cross sectional

area after body-weight circuit training with BFR. There is also evidence to suggest that

BFR added to a traditional physical therapy program following ACL graft reconstruction

can augment improvements in knee extensor strength and cross sectional area (Ohta et

al., 2003).
 26

Aerobic training with BFR has also been reported to elicit similar muscular and

functional improvements as resistance training with BFR in healthy adults. Abe et al.

(2006) examined the effect of a regular (2 times per day, 6 days per week for 3 weeks)

light-intensity (50m/min) walk training program with BFR compared to a similar walk

training program without BFR when 200 mmHg was applied at the proximal thigh.

Following the BFR training program, thigh (muscle and bone) CSA increased, as did

muscle volume and strength (isometric MVC and maximal strength). These changes

were not observed in the non-BFR group. Similar adaptations to BFR walk-training have

been reported elsewhere (Abe et al., 2009; Sakamaki, M, & Abe, 2011). Ozaki et al.

(2011) tested this effect in a group of elderly sedentary women, and not only observed

improvements in muscle size and strength after regular low intensity (45% of heart rate

reserve) BFR walk training, but also in Timed Up and Go scores (a measure of functional

ability). Salvador et al. (2016) examined the effect of 4 weeks of BFR walk training in a

20 year old Paralympic sprinter who had right-sided hemiplegic cerebral palsy. Results

indicated that BFR training improved 400 meter run performance, 100 meter run

performance, and MVC in the affected leg increased by 19% while the non-affected leg

increased only 9% (thereby reduced functional asymmetry). This indicates that BFR

training could be effective even in limbs with limited neural innervation. However, there

is a void of research examining this type of training in individuals with iSCI. In one

select study, BFR combined with FES training of wrist extensors was observed to

significantly increase the cross sectional area (CSA) of the wrist extensors in a 9 subjects

with SCI, and this increase in CSA was 17% greater than that observed following FES
 27

training without BFR (Gorgey et al., 2016). Subjects in this study were 9 men who had

impairment scores ranging from ASIA B through ASIA D (almost no impairment), had

an injury above C8 (and therefore were tetraplegic), and were at least 2 months post

injury. Training included 6 weeks of twice weekly exercise that lasted for 30 minutes

each session. Exercise included FES handgrip exercise of the right forearm, and FES

handgrip exercise combined with BFR (130% systolic) of the right arm. Subjects

completed 4 sets of 10 repetitions in each arm at a 5:5 on-off ratio. The stimulation was

set to 20Hz, a 450 µs pulse duration, and an amplitude necessary for wrist extension

without finger extension (validated prior). While these results promote the feasibility of

BFR exercise in this population, the small muscle mass used limits the generalizability to

large-muscle group exercise such as walking and lower body resistance training. Other

than improvements in muscular strength and size, regular BFR walk training has also

been observed to increase maximal aerobic capacity and muscular endurance (Park et al.,

2010).

Potential mechanisms

There is some debate regarding the potential mechanisms for the adaptations to

regular BFR training. For one, the increases in cardiorespiratory endurance may be due

to increased mitochondrial density in response to prolonged exposure to hypoxemia, as

has been observed following prolong exposure to hypoxia (Jacobs et al., 2016).

However, Larkin et al. (2012) found that prolonged knee extension exercise with BFR

increased vascular endothelial growth factor (VEGF) and other angiogenic factors. This

could explain an increase in capillary density and tissue perfusion and also contribute to
 28

improvements in aerobic capacity, as well as muscular endurance. Improvements in

muscular strength and size could be mediated by a number of factors, such as preferential

recruitment of Type II muscle fibers. Based on the size principle, the increases in

muscular activation during submaximal exercise with BFR are indicative of large motor

unit (Type II) recruitment (Moritani, Sherman, Shibata, Matsumoto, & Shinohara, 1992).

Ultimately Type II fiber recruitment during low intensity exercise could influence

increases in strength and hypertrophy. Fujita et al. (2007) also observed greater S6

Kinase 1 phosphorylation during BFR exercise compared to non-BFR exercise, which is

indicative of Type II fiber recruitment. However, some have reported that this effect

does not occur when both exercises (BFR and non-BFR) are performed to fatigue

(Wernbom, Jarrebring, Andreasson, & Augustsson, 2009), and suggest that BFR elicits

afferent activity that inhibits the motor signal. None of the studies mentioned here

examined the chronic effects of BFR training on preferential Type II fiber recruitment.

Another potential mechanism for improving muscular strength with BFR training is

fatigue. Yasuda, Fujita, Ogasawara, Sato, and Abe (2010) examined the effect of bench

press training with BFR on chest muscle hypertrophy and strength compare to a control

group. Results indicated that that following 2 weeks of 30% bench press 1RM, triceps

muscle thickness, and pectoralis major muscle thickness increased in the BFR group, but

not the control group. This suggests that the muscles above the level of occlusion can

benefit from the fatigue of the muscle that are ischemic, which may be a mechanism by

which functional scores (such as the Timed Up and Go test) are improved following

regular BFR training. Finally, accumulation of metabolites is another potential

 29

mechanism for muscular benefits observed with BFR training. The accumulation of

metabolites is purported to stimulate peripheral adaptations and anabolic factors

(Loenneke, Wilson, & Wilson, 2010). For example, when cuff occlusion was applied to

a group of patients immediately after ACL-reconstruction muscle atrophy was reduced by

9.4% compared to 20.7% in a group that did not receive occlusion therapy (Takarada,

Takazawa, & Ishii, 2000). In this study exercise was not used as a stimulus, rather it was

simply blood flow occlusion, which eliminated a number of the previously proposed

mechanisms, with the exception of hypoxemia, which also influences metabolite

accumulation. Ultimately, these mechanisms likely work together to promote the

adaptations to BFR training discussed here.

Safety issues

Just like any specialized training modality, BFR training has a number of safety

considerations. These generally revolve around the cardiovascular system. One of these

issues is the dysregulation of blood flow in the peripheral vascular system. The primary

goal of BFR exercise is to occlude venous flow and restrict arterial flow to the exercising

muscle. This has the potential to cause excess venous distension and the pooling of blood

in the lower body (if performing lower body BFR exercise). A decrease in venous return

due to the occlusion pressure could cause a drop in stroke volume, which would then

elicit a rapid increase in heart rate. This is supported by Renzi, Tanaka, and Sugawara

(2010) who observed an increase in heart rate during BFR compared to non-BFR exercise

with a simultaneous drop in stroke volume and systemic arterial compliance; however, is

also disputed by other research that comparing BFR exercise to high intensity exercise
 30

(Libardi et al., 2017). This could also be potentiated by the trapping of metabolites below

the level of occlusion that could trigger a metaboreflex mediated increase in heart rate.

While it is normal for heart rate to increase with exercise, individuals with a history of

abnormal tachycardic episodes should be carefully monitored during BFR exercise.

Another issue related to the cardiovascular system is dysregulation of blood pressure.

Evidence suggests that mean arterial pressure (MAP) and TPR are both reduced during

low-intensity BFR compared to high intensity non-BFR exercise (Loenneke, Wilson,

Wilson, Pujol, & Bemben, 2011). However, when matched for exercise intensity, MAP

tends to be higher. For example, Takano, Morita, Iida, Asada, et al. (2005) compared

MAP between low intensity (20%) resistance exercise with and without BFR and found

that MAP increased from 98 ± 18 to 127 ± 12 with BFR compared to an increase from 88

± 9 to 113 ± 27 without BFR. Therefore, individuals with hypertension should be closely

monitored during BFR exercise, and avoid any maneuvers that could exacerbate the

increase in blood pressure (e.g. the Valsavla). A third consideration related to the

cardiovascular system that is very pertinent to individuals with iSCI is the potential for

development of blood clots. Stopping blood flow in the venous system could potentially

increase the risk for developing a Deep Vein Thrombosis (DVT). Research on the risk of

DVT development with training is still somewhat limited however, some evidence

suggests a very minimal risk ( 0.055%) (Clark et al., 2011; Nakajima et al., 2006;

Nakajima et al., 2007). In the spinal cord injured population, the risk of all-cause DVT is

suggested to be between 10% and 30% during the acute phase of an SCI, but less than 2%

during the chronic phase (Hagen, Rekand, Gronning, & Faerestrand, 2012; Teasell et al.,
 31

2009). The risk for DVT formation is likely population specific, and to date there is no

research on the risk of DVT formation with BFR training in the spinal cord injured

population.

Need For Research

There is mounting evidence supporting the use of BFR exercise as a method for

improving muscular strength, size, and overall function in the healthy and elderly

populations. However, there is a void of research that examines the potential benefits of

BFR training in individuals with SCI. One of the more substantial findings regarding this

exercise modality is that it can elicit similar muscular adaptations to high intensity non-

BFR exercise at very-low intensities (20-30% 1RM and 50 m*min-1). That fact alone

suggests that it could be very beneficial for individuals with iSCI as this population

suffers from a functional limitation that is mediated by impaired neuromuscular

recruitment; and this functional limitation also limits the intensity of exercise that can be

performed during physical rehabilitation. This may be one reason why physical

therapists often see a plateau in strength and functional gains with traditional physical

rehabilitation methods. Therefore, BFR exercise might allow someone with an iSCI to

achieve gains in strength, size, and function normally observed following high intensity

exercise, even if they can only perform low intensity exercise. One could also speculate

that if muscular adaptations could be potentiated through BFR exercise, this could

increase the intensity that this population is able to achieve during regular exercise, which

could then enhance physical rehabilitation outcomes in a step-wise manner. Ultimately,

BFR exercise might allow individuals with iSCI to maximize their training adaptations
 32

and improve overall function. However, before we can truly test the effects of BFR

training in a large sample of subjects with iSCI, we first need to determine if it is feasible

and safe in this population.

There are a number of risks from BFR training that are specific to the individuals

with iSCI. One of these is the risk of AD. BFR requires an external pressure to be

placed over the proximal region of the arm or thigh. This could be enough of a noxious

stimulus to elicit a period of AD. Another risk that is specific to individuals with SCI is

the development of a DVT. DVT risk is already elevated following an SCI or iSCI due to

a lack of mobility in the legs, which reduces movement of blood through the venous

system. Since this risk is already elevated in individuals with SCI's, it is important for

research using BFR in individuals with SCI's to closely monitor DVT risk before and

after exercise.

There is also a risk that these individuals won't be able to perform BFR exercise

due their existing strength impairments. Previous research suggests that BFR exercise is

more difficult than non-BFR exercise, which may make it too difficult for someone who

has relatively weaker muscles. Therefore, it is imperative that the safety and feasibility

of BFR exercise be tested in individuals with iSCIs so that future research can begin

examining its efficacy.

Purpose and Hypothesis

There are two primary aims of this study. The first aim is to determine the

feasibility and safety of BFR exercise for individuals with iSCI. This will be done by

monitoring signs of AD and DVT risk during walking with BFR and lower body
 33

resistance exercise (knee extension) with BFR. We hypothesize that individuals with

iSCI will be able to perform both exercises with only moderate reductions in work

output, and while exhibiting no significant risk for AD or development of DVT.

The second aim of this study will be to explore the potential efficacy of BFR exercise in

individuals with iSCI. This will be achieved by comparing the result of BFR exercise

(walking and knee extension) to non-BFR (traditional) exercise in a sample of iSCI

subjects. Specifically, we will examine EMG activity and tissues oxygenation of the

rectus femoris, whole body lactate accumulation, MAP, HR, and perceived effort during

walking and lower body resistance exercise with and without blood flow restriction. We

hypothesize that whole blood lactate accumulation, HR, and EMG activity during

submaximal exercise will be higher during BFR exercise (both walking and knee

extension) compared to non-BFR exercise; and that muscular tissue oxygenation of the

rectus femoris will decrease during BFR exercise compared to non-BFR exercise. These

results would be consistent with previous research on BFR exercise in the general

population. One hypothesis that is inconsistent with previous research is that we

anticipate MAP to increase during BFR exercise compared to non-BFR exercise. This

hypothesis is grounded in the existing dysregulation of blood flow associated with iSCI,

and the exercise intensities that will be used. Previous research has compared low

intensity BFR exercise to high intensity non-BFR exercise. The subjects in this

investigation will be self-selecting their intensity, and that intensity will be maintained

between each trial. Therefore, the exercise stimulus is the same and we anticipate an

increased in the EPR response due to augmented metabolite accumulation below the level
 34

of occlusion. Normally this would be countered by the fact that venous return decreases,

and therefore so does the SV and MAP. However, in this population SV is already

impaired and would likely not be different enough between sessions to significantly

affect MAP. Simply stated, we speculate that there would be an augmented EPR response

during BFR exercise that would cause a HR driven increase in MAP.

A third purpose of this dissertation is to use the data from this study and the

literature discussed therein to develop a grant application for a long-term study focusing

on BFR training for individuals with SCI. If this training method proves to be safe and

feasible for individuals with iSCIs, it may then have substantial implications for

enhancing muscular adaptations to physical rehabilitation and ultimately functional

abilities. The first aim of this study would be to determine the appropriate exercise

intensity and occlusion pressure to use during BFR walking and BFR resistance exercise.

The second aim of this study would be to determine if BFR exercise could be performed

safely over a long period of time, with an emphasis on the risk of DVT and clot

formation. Finally, this study would determine the training outcomes of regular BFR

exercise in individuals with iSCI. Specifically, this study would examine muscular

characteristics (muscular quality, cross sectional area, torque and force production, fiber

type distribution, and fatigability) and functional outcomes (walking speed, gait,

endurance, timed-up-and-go test, etc.). Ideally, this grant would fund a multi-site study

that would have the potential of reaching a large number of individuals with iSCI and

allow for long term collaborative research between exercise physiologist, physical

therapists, neuroscientists, and other allied health professionals. Results from such a
 35

study could have significant implications for the future of physical rehabilitation from an

incomplete spinal cord injury.

 CHAPTER III

METHODS

This section will begin with a description of the study design, and the individuals

who participated in this study.

Study Design

This study included two separate experiments, each of which followed a repeated

measures design. These experiments occurred on separate days, and all data were

collected at the Louis Stokes' Cleveland V.A. Medical Center in Cleveland, Ohio.

Participants

This study was a feasibility study that is examining a new training method and

there is no current research from which to estimate an effect size. Therefore, data were

collected on 9 subjects in experiment 1, and on 9 subjects in experiment 2. Subjects were

be able to participate in both experiments, however, a three-week washout period was

required between experiments. Subjects were recruited from the Rehabilitation

Therapies, Spinal Cord Injury and Research Center at the Louis Stokes' Cleveland V.A.

Medical Center. Subjects in experiment 1 were diagnosed with chronic (at least 6 months

prior) incomplete spinal cord injuries classified as either ASIA-C or ASIA-D. Subjects in

experiment 2 were also classified as either ASIA-C or ASIA-D, and must have been able

to walk at least 100 feet without assistance. In both experiments, subjects were free of

any recent history (3 years) of diagnosed DVT, and were prescreened for asymptomatic

DVT by a Doppler ultrasound scan. The existence of a DVT or recent history of a DVT

36
 37

precluded participation. Subjects did not have any history of cardiovascular disease

(myocardial infarction, congestive heart failure, coronary artery disease), stroke,

musculoskeletal injuries (non-spinal cord injury related), tissue wounds (i.e. pressure

ulcers), or uncontrolled spasticity.

Protocol

This section describes each experimental protocol, separately.

Experiment 1

Experiment 1 compared the fatigue index, rate of torque development, muscular

activation, and lactate accumulation between BFR and non-BFR resistance exercise in a

group of subjects with iSCI. This experiment included a BFR and a non-BFR resistance

exercise trial, the order of which was counterbalanced. Each trial began with subjects

resting for 5 minutes before resting blood pressure (BP), HR , and whole blood lactate

were collected. Bipolar EMG electrodes were also placed over the vastus lateralis one

third of the way between the patella and the iliac crest to record muscle activation. Near-

infrared spectroscopy (NIRS) electrodes were placed over the rectus femoris to determine

muscle tissue oxygenation. Next, subjects performed the resistance execise protocol,

beginning with the control session. This protocol included 3 sets of 10 repetitions of

knee extension exercise at a self-selected intensity. There is a high degree of variability

in the neuromotor function and force generation in this population, which limits the use

of the guidelines for prescribing resistance exercise intensity for the general population.

Therefore, the same resistance load that was self-selected during the control session was

also used for the BFR session. Perceived effort was also collected after each set of
 38

exercise using a 10 cm visual analog scale (VAS) and compared between each session.

BP, HR, EMG, and muscle tissue oxygenation were collected continously throughout

exercise, and monitored for 15 minutes after exercise. Muscle spasticity and venous

blood flow were collected immediately after the 3rd set of exercise, and whole blood

lacatate was collected at the finger 5 minutes after the end of the third set of exercise.

Whole blood lactate was collected 5 minutes after the cessation of exercise to allow for

blood lactate to reach systemic circulation.

After the first exercise trial subjects rested quietly for at least 15 mintues until BP

and HR returned to near-resting levels (±10 bpm for heart rate, and ±10 mmHg for

systolic and diastolic blood pressure) before beginning the second trial. For the BFR

trial, a pneumatic cuff was placed around the proximal region of the thigh, just distal to

the inguinal crease. The cuff was slowly inflated to 40 mmHg above venous occlusion,

validated at the popliteal artery via Doppler ultrasound (G.E. Logic 7, Doppler

Ultrasound, Milwaukee, WI). The three sets of 10 repetitions were performed again with

the cuff inflated throughout the entire protocol, and immediately released after the end of

the 3rd set. BP, HR, muscle tissue oxygenation, and EMG were recorded throughout the

protocol and for 15 minutes after exercise, muscle spasticity and venous blood flow were

collected immediately after the 3rd set, and a finger stick was used to measure blood

lactate 5 minutes after the cessation of exercise. Together, these protocols allowed us to

determine whether or not BFR reduces the ability of those with iSCI to perform knee

extension exercise and the degree to which muscular strength and endurance may be

influenced by BFR. Three to 4 days following experiment 1, subjects reported back to

 39

the VA medical center where they had a blood sample drawn which was analyzed for D-

Dimer. D-Dimer is a protein fragment that is released during fibrinolysis, and is used as a

marker of blood clot formation. It is possible for factors other than the existance of a

blood clot to increase circulating D-Dimer. Therefore, if the analysis indicated

abnormally high levels of D-Dimer, subjects underwent a second Doppler ultrasound

DVT screening for the presence of a DVT.

Experiment 2

Experiment 2 tested the feasibility of BFR walking by comparing walking

duration with and without the application of BFR. This experiment also tested the safety

of BFR walking by monitoring BP, HR, spasticity, neurpathic pain, and venous blood

flow before, during, and after BFR walking. Subjects entered the lab 3 hours postprandial

and having abstained from any major physical activity for 24 hours. First, subjects rested

for 5 minutes before resting BP, HR, popliteal veinous flow, rectus femoris tissue

oxygenation, and spasticity were assessed. After baseline data were collected, each

subject was secured into a LiteGait body weight support system (LiteGait, Tempe, AZ).

This system attaches around the waist and inguinal region, and allows investigators to

support a predetermined percentage of the subject’s body weight during walking. The

body weight support system was used primarily for fall prevention and therefore did not

support any body weight during walking in this experiment. Once subjects were secured

into the LiteGait system they performed the walking protocol. During the BFR trial, the

cuff was inflated at the beginning of exercise. The walking protocol included 6 2-minute

bouts of walking at a self-selected pace. A 5 minute rest period was given between bouts
 40

3 and 4, during which the occlusion pressure was deflated. A 1-minute rest period was

given between each of the other bouts, during which the occlusion pressure was

maintained. Spasticity and neuropathic pain were assessed a second time immediately

following the cessation of exercise, and BP, HR, and muscle tissue oxygenation were

monitored throughout exercise, and for 15 minutes after exercise. After the 15 minute

recovery period subjects were secured into the LiteGait system a second time and a pair

of pneumatic cuffs were fitted around the proximal regions of the each thigh, just below

the inguinal crease. The cuffs were slowly inflated to 40 mmHg past the point of

complete venous occlusion, which was validated at the popliteal vein via Doppler

ultrasound (G.E. Logic 7, Doppler Ultrasound, Milwaukee, WI). Subjects then

perfromed the same walking protocol previously described. The occlusion pressure was

maintained during each of the 1 minute rest periods, but was relesased during the 2

minute rest period between bouts 3 and 4. Venous blood flow, neuropathic pain, and

spasticity were assessed immediately following exercise, and BP, HR, and muscle tissue

oxygenation was monitored continously throughout execise and for 15 minutes following

exercise. Three to 4 days following experiment 2, subjects reported back to the VA

medical center where they had a blood sample drawn which was analyzed for D-Dimer.

If the analysis indicated abnormally high levels of D-Dimer, subjects underwent a second

Doppler ultrasound DVT screening.

Variables

This section explains how and when each variable was collected.
 41

D-dimer

D-Dimer is a protein fragment that is release during fibrinolysis. It is commonly

used in research as an indicator of the presence of a DVT. In this study, we collected a

sample of blood 1 week after the end of each experiment via a venipuncture blood draw

which was analyzed for D-Dimer levels. Subjects whose D-Dimer levels were above

normal (as indicated by a physician) were screened for a DVT via Doppler ultrasound.

This second screening was necessary, because it is possible that D-Dimer can be elevated

due to factors other than the presence of a DVT. The compression method of Doppler

ultrasound DVT screening was used in this investigation. During this screening, a

practitioner compresses the Common Femoral Vein (2 cm proximal to 2 cm distal to the

junction with the Greater Saphenous vein), the Deep and Superficial Femoral Veins, and

the Popliteal Vein (most distal 2cm and the base of the trifurcation of the Anterior Tibial

Vein, the Posterior Tibial Vein, and the Peroneal Vein). Clips were taken of the

compression of these veins and sent to the radiology lab, where they were interpreted and

diagnosed by a licensed radiologist.

Venous blood flow

Blood velocity was collected in 1 minute clips at the popliteal vein of least-

affected leg (or the dominant leg if motor scores are equal) via Doppler ultrasound (GE

Logic 7, GE Healthcare, Milwaukee, WI). Diameter was also assessed during each

minute used to calculate venous blood flow. In experiment 1, venous blood flow was

compared at baseline, immediately following walking exercise, and at the, 5th, 10th, and

15th minute of recovery. In Experiment 2, venous blood flow was compared at baseline,
 42

immediately after the 3rd set of knee extension, and the 5th, 10th, and 15th minute of

recovery.

Spasticity

Spasticity was assessed via the Modified Ashworth Scales (MAS; Appendix C).

This scale is commonly used in research, and is a comprehensive and validated measure

of joint spasticity (Akpinar et al., 2017; Min et al., 2012; Santos, Santos, Krueger,

Nogueira-Neto, & Nohama, 2016). Spasticity was assessed at the knee and ankle of each

leg and an aggregate score was computed and compared at baseline and immediately

following the 3rd set of knee extension in experiment 1, and at baseline and immediately

following walking exercise for experiment 2.

Neuropathic pain

Neuropathic pain was assessed via the Neuropathic Pain Scale (NPS; Appendix

B) before and after each knee extension trial in experiment 1, and before and after each

walking trial in experiment 2. The NPS is a comprehensive and validated assessment

tool used for assessment of neuropathic pain (Fishbain et al., 2008; Rog, Nurmikko,

Friede, & Young, 2007).

Muscle activation

EMG mean amplitude and mean frequency (root mean squares; rms) were

collected during each MVC and throughout each exercise trial. EMG activity was

compared during the MVC of BFR and non-BFR knee extension, during BFR and non-

BFR knee extension exercise, and during BFR and non-BFR walking.
 43

Perceived effort

Perceived effort was collected with a 10 cm visual analog scale (VAS). Subjects

were given a writing utensil to mark a perpendicular line along the VAS to indicate their

level of perceived effort. This was collected immediately following each set of knee

extension (3) and after the 3rd and 6th walking bout in both the BFR and non-BFR trials.

Blood pressure and heart rate

BP and HR were monitored by a continuous blood pressure monitor (Nexfin,

BMEYE, Amsterdam, The Netherlands) throughout exercise as indicators of potential

AD risk. For experiment 1, BP and HR were compared at baseline, during each set of

knee extension exercise (3), and during the 1st, 5th, 10th, and 15th minutes of recovery.

In experiment 2, BP and HR were compared at baseline, during the 6th and 12th minute

of walking, and during the 1st, 5th, 10th, and 15th minutes of recovery.

Whole blood lactate

Whole blood lactate was collected at baseline and post exercise in each trial via a

finger-stick. Changes in blood lactate were compared between in each trial.

Muscle tissue oxygenation

Muscle tissue oxygenation will be collected using near-infrared spectroscopy

(NIRS; Oxymon MK III, Artinis, Zetten, The Netherlands). This will be collected at

baseline, continuously throughout exercise, and for 15 minutes after exercise. Data will

be collected at a frequency of 1 Hz and averaged in 1 minute clips. In experiment 1,

Muscle tissue oxygenation will be compared at baseline, during each set of knee

extension (3), and during the 1st, 5th, 10th, and 15th minutes of recovery. In experiment
 44

2, muscle tissue oxygenation will be compared at baseline, during the 6 th and 12th minute

of walking, and at the 1st, 5th, 10th, and 15th minutes of recovery.

Statistical Analyses

This section details who data were analyzed following each experiment.

Experiment 1

A 2 (time; pre-, post-exercise) by 2 (condition; BFR, non-BFR) repeated

measures ANOVA was used to test for any significant interactions, or main effects of

time and condition for spasticity, neuropathic pain, and venous blood flow. A 7 (time;

baseline, 6th and 12th min of walking, and 1st, 5th, 10th, and 15th minute of recovery) by

2 (condition; BFR and non-BFR) repeated measures ANOVA was used to test for any

significant interactions, or main effects of time and condition for BP, HR, and muscle

tissue oxygenation. Any significant differences in any of the previously listed variables

were examined further with post-hoc analyses using an LSD correction. Paired samples

t-tests were used to test for significant differences between walking duration and distance

walked between the two trials.

Experiment 2

A 2 (time; pre-, post-exercise) by 2 (condition; BFR and non-BFR) repeated

measures ANOVA was used to test for any significant interactions, or main effect of time

and condition for whole blood lactate and muscle spasticity. A 3 (sets) by 2 (condition;

BFR and non-BFR) repeated measures ANOVA was used to test for any significant

interactions, or main effects of time and condition for muscle activation and perceived
 45

effort during submaximal exercise. An 8 (time; baseline, 1st, 2nd, and 3rd set, and 1st,

5th, 10th, and 15th min of recovery) by 2 (condition; BFR and non-BFR) repeated

measures ANOVA was used to test for any significant interactions, or main effects of

time and condition for BP, HR, and muscle tissue oxygenation. Any significant main

effects were further explored with post-hoc analyses using an LSD correction. Finally, a

paired-samples t-test was used to test for any significant differences in fatigue index

between exercise trials.

 CHAPTER IV

THE FEASIBILITY OF BLOOD FLOW RESTRICTION EXERCISE IN

INCOMPLETE SPINAL CORD INJURED PATIENTS

Spinal cord injuries (SCI) are common neurological disorders that disrupt the

propagation of action potentials along the spinal cord resulting in motor dysfunction,

paralysis, autonomic disorders, and other symptoms which impact daily activities and

quality of life (Kawanishi & Greguol, 2013). The severity of an SCI depends on the

location of the injury along the vertebral column and the completeness of the injury

through the spinal cord. An incomplete spinal cord injury (iSCI) can occur anywhere

along the spinal column, and is characterized by residual sensory or motor function at the

lower portion of the spinal cord (Roberts et al., 2016). The loss of lower limb function

varies widely between different iSCIs: some individuals may be limited to weak

nonfunctional muscle contractions while others can still ambulate. Rehabilitation

programs for individuals with iSCIs are heavily focused on restoring as much motor

function as possible, with an emphasis on locomotor function (Dietz & Fouad, 2014).

Other rehabilitation goals include increasing muscular strength, muscular endurance, and

work capacity; all with the ultimate goal of improving quality of life and obtaining (or

maintaining) independence (Kozlowski & Heinemann, 2013). However, the efficacy of

these rehabilitation programs is challenged by the limited motor function, and therefore

limited exercise intensity, performed by iSCI patients. Therefore, an exercise modality

that maximizes muscular adaptations to light intensity exercise would be ideal in this

population.

46
 47

Blood flow restriction (BFR) training is a form of exercise training that uses

external pressure to occlude venous blood flow from, and limits arterial flow to, the

exercising muscle. Previous reports indicate this type of training increases muscular

strength, endurance, cross sectional area (CSA), and even aerobic exercise capacity at

very low (20-30% maximal exertion) intensities (Abe et al., 2010; Abe et al., 2006;

Barcelos et al., 2015). This affect is attributed to a combination of metabolite

accumulation during exercise, preferential type II fiber recruitment, hypoxemia induced

adaptations to mitochondrial volume and density, elevated growth hormone, and

activation of myogenic cells (Goto et al., 2005; Kon et al., 2012; Lang et al., 1998;

Loenneke, Fahs, et al., 2011; Meyer, 2006; Takarada, Nakamura, et al., 2000). These

benefits have been observed in normal healthy adults (Abe et al., 2010), athletes

Luebbers, Fry, Kriley, & Butler, 2014), elderly individuals (Ozaki et al., 2011) and those

with skeletal-muscle injuries (Ohta et al., 2003). To date there is limited information on

the efficacy of BFR training in those with iSCI, with only Gorgy et al (2016) reporting a

17% greater increase following combined functional electrical stimulation (FES) and

BFR wrist extensor exercise compared to FES alone in 9 individuals with incomplete

tetraplegia.

While the addition of blood flow restriction training to the rehabilitation program

of an individual with an iSCI might present some challenges, the potential benefits could

be substantial. Improving muscular fitness and aerobic capacity in this population may

lead to enhanced motor function and improved independence. Therefore, the purpose of

the present study was to determine the feasibility of BFR knee extension exercise, and to
 48

compare the physiological responses to BFR and non BFR knee extension exercise, in

individuals with iSCI. Another aim of this study was to explore the feasibility of BFR

walking in a sub-set of individuals with iSCI's that still maintain the ability to ambulate.

We hypothesized that individuals with iSCIs would be able to perform knee extensor and

walking exercises with BFR with only moderate reductions in repetitions performed and

distance walked, respectively. We also hypothesized that muscle activation, lactate

accumulation, pain, perceived effort, and the pressor response would all be augmented

with BFR exercise, while local tissue oxygenation would be attenuated. Finally, because

these individuals have incomplete injuries rather than complete injuries, we hypothesized

that they would be able to perform BFR exercise without exhibiting any signs of

Autonomic Dysreflexia (AD) or development of Deep Vein Thrombosis (DVT).

Methods

This study included 2 separate protocols; one a knee extension protocol (n=9) and

the other a walking protocol (n=3). All subjects completed the knee extension protocol,

while only a sub-set of subjects were recruited for the walking protocol. All subjects

were recruited from the Louis Stokes' Veterans Affairs Medical Center in Cleveland,

Ohio, and all provided written informed consent. In order to participate in this study,

subjects must have been diagnosed with a chronic (>6 months) incomplete spinal cord

injury and clear of any existing or recent history of DVT (3 years), recent myocardial

infarction and/or significant hypertension (>160 systolic or >100 diastolic). Level of

injury varied between subjects, and is presented along with other subject characteristics

in Table 1. The protocols used in this study were approved by the Louis Stokes' VA
 49

Medical Center's Institutional Review Board (IRB Log#17005-H05), and conformed to

the standards set forth by the Declaration of Helsinki.

Experimental protocol 1

Participation in protocol 1 included three total visits to the medical center (Figure

1). The first visit included a bilateral leg DVT screening performed by an ultrasound

technician within the medical center’s radiology department. This served as a pre-

screening, and anyone who presented with an existing DVT was excluded from

participation in this study.

The subsequent experimental visit included 2 separate exercise conditions in

counterbalanced order; one active control condition and one BFR condition. Upon

entering the laboratory, subjects were fitted with an pneumatic pressure cuff around the

most proximal region of the least affected leg (DE Hokanson, Inc., Bellevue, WA), and a

beat-by-beat blood pressure monitor around the right wrist and middle finger (Nexfin,

Edwards LifeSciences Corp., Irvine, CA ). Electromyography (EMG) electrodes were

also placed two-thirds of the way between the anterior spina illiaca and (lateral to) the

patella on the vastus lateralis (ADInstruments Inc., Colorado Springs, CO), and near-

infrared-spectroscopy optodes were placed on the rectus femoris, medial from the EMG

electrodes (Hamamastu, Hamamastu City, Shizouka Pref., Japan). The acquired EMG

signal was sampled at 2kHz, amplified (x1000) and bandpass filtered (13 and 1000 Hz).

Lastly, a three-lead ECG (ADInstruments, Colorado Springs, CO) was used to collect

continuous heart rate data and a goniometer was affixed to the knee joint to record knee

extension and flexion movement. Once fitted with all data acquisition equipment, cuff
 50

pressures required to occlude the femoral vein and artery were verified at the popliteal

vein and artery, respectively, via Doppler ultrasound (GE Healthcare, Milwaukee, WI).

To accomplish this, the popliteal vein was identified using a GE logiq 7

Doppler/ultrasound equipped with a 9L linear array operating at14 MHz (Ultrasound) and

5MHz (Doppler). The cuff pressure was then slowly increased until venous blood flow

was occluded. Once venous flow was occluded, the probe was shifted to image the

popliteal artery, and pressure was slowly increased until arterial flow was completely

occluded. The venous occlusion pressure is presented as a percentage of arterial

occlusion pressure in Table 1, which can be used as a benchmark for comparisons to

previously published data.

The appropriate resistance load (weighted ankle cuffs) was then selected for each

subject. Resistance load was selected by consulting with the subject's chief physical

therapist, reviewing the loads used during the subject's current physical therapy program,

and testing single repetitions until a load was identified that could be performed for 3 sets

of 10 repetitions with moderate difficulty. The same resistance was used throughout both

exercise bouts. Next, knee extensor and flexor tone were assessed by a licensed physical

therapist using the Modified Ashworth Scale (MAS; Appendix C). After tone was

assessed, subjects performed 5 minutes of seated controlled-rate breathing to a

metronome (15 breaths per minute) to obtain steady ECG tracings for subsequent analysis

of the low frequency to high frequency (LF/HF) ratio (Table 1). The LF/HF ratio was

used as an indicator of baseline sympathovagal balance (Grimm, DeMeersman,

Almenoff, Spungen, & Bauman, 1997). A baseline lactate reading was then collected,
 51

followed by exercise. In the BFR bout, the cuff was inflated to 125% of the venous

occlusion pressure immediately prior to the start of the first set of exercise. Each exercise

bout included 3 sets of 10 repetitions at the previously selected resistance with 90

seconds of rest between sets. Repetitions were performed at a constant controlled rate

(52 bpm; 2:1 concentric to eccentric ratio) via a metronome. In the BFR condition, cuff

pressure was released immediately after completion of the third set. Perceived pain was

collected using a 10cm visual analog scale (VAS) just prior to exercise (after cuff

inflation in the BFR condition), and immediately after each set of exercise. Perceived

difficulty was also collected using a 10cm VAS after each set of exercise. Lactate and

knee flexor and extensor tone were collected post-exercise in each condition, followed by

a 15-minute recovery period during which subjects rested quietly. Mean arterial pressure

(MAP) and heart rate (HR) were allowed to return within 10% of baseline values between

the 2 conditions.

In order to determine the risk of DVT formation following BFR exercise, D-dimer

levels were assessed within a week following the experimental trial via a 5.0 mL blood

draw. D-dimer is a protein fragment that serves as a maker of fibronolysis. Therefore,

elevated D-dimer levels can be suggestive of thrombosis. However, since D-dimer can

be affected by a number of additional factors it may not be an independent predictor of

recent DVT formation. Therefore, any subjects who presented with an abnormally high

quantitative D-dimer reading (>500 ng/mL) underwent a second bilateral leg ultrasound

DVT scan (n=2) to either confirm or rule out the existence of a formed DVT. This

concluded participation in protocol 1.

 52

Experimental protocol 2

The second protocol was conducted 2 weeks after completion of the first protocol,

and included 2 separate visits to the medical center (Figure 1). The first visit was the

experimental session, and included 2 separate bouts of exercise (BFR and control) in a

counterbalanced order. Similar to the first protocol, subjects were first fitted to all data

collection equipment before venous and arterial occlusion pressure were validated at the

popliteal vein and artery, and resting knee flexor and extensor tone were assessed.

Subjects then performed 5 minutes of seated controlled-rate breathing (15 breaths per

minute) to a metronome, followed by a baseline lactate assessment. Next, subjects were

secured into an adult treadmill harness (LiteGait, Tempe, AZ). This treadmill harness

was used as a fall-prevention measure, and did not support any of the subjects’ body

weight during walking. Subjects then performed either BFR or control walking. Both

walking conditions included 6 two-minute bouts of walking, each separated by a 60

second rest period with the exception of bouts three and four, which were separated by a

5-minute rest period (Figure 2). In the BFR condition, the cuff was inflated to 125% of

venous occlusion pressure just prior to the start of the 1 st walking bout and deflated

immediately following the end of the 3rd walking bout; and then re-inflated just before the

start of the 4th walking bout and deflated immediately following the end of the 6 th

walking bout. Blood pressure was monitored continuously during exercise, and each trial

was separated by a 15-minute recovery period.

Three to 4 days following the completion of the experimental session, subjects

returned to the medical center for a blood-draw that was analyzed for quantitative D-
 53

dimer levels. Any subject whose D-dimer readings were abnormally high (>500 ng/mL)

were referred to the radiology department for a follow-up bilateral leg ultrasound DVT

scan.

Statistical analysis

In order to elucidate the hemodynamic kinetics across the entire exercise session

and within each exercise bout independently, HR, MAP, systolic blood pressure (SBP),

diastolic blood pressure (DBP), oxygenated hemoglobin saturation (Oxy), deoxygenated

hemoglobin saturation (Doxy), total hemoglobin (TotalHb), and tissue oxygenation index

(TOI) were all analyzed using 2 separate repeated measures analyses of variance

(ANOVA) in both protocols. The first of these analyses compared these variables

between conditions and across the entire protocol (baseline, exercise, minutes 8-10 of

recovery, and minutes 13-15 of recovery). The second analysis compared these variables

between conditions, and across each acute bout of exercise (baseline, and the last 10

seconds of each set of exercise and each subsequent rest period). Pain was also compared

between conditions and across time (baseline, sets 1 and 2), as were perceived difficulty

(sets 1, 2, and 3), lactate accumulation, and muscle tone (baseline and post exercise).

Any significant differences were analyzed further using a post-hoc comparison using a

Fisher Least Significant Difference (LSD) correction, and the total number of

contractions performed were compared between conditions using a paired-samples t-test.

Finally, incidence of acute AD and DVT formation, as well as any differences in distance

walked or walking speed, were reported for each individual subject for protocol 2.
 54

Results

All subjects were able to perform the desired number of sets (3) and repetitions

(30) for both exercise conditions in protocol 1, and without exhibiting any signs or

symptoms of acute AD. Two subjects showed signs of elevated D-dimer following the

BFR trial, which were then followed up by ultrasound DVT Scans, both of which

returned negative. There were no significant interactions, nor effects of time or condition

for pain, perceived effort, lactate accumulation, muscle tone, or EMG.

Hemodynamics

When examining changes across each experimental condition, MAP and SBP

significantly changed across time (F3,15=8.081 and F3,15=7.817, respectively; p<0.05);

however, they were not different between conditions. This difference is explained by

significant increases in MAP (+15.4±8.3 mmHg; Figure 3) and SBP (+20.6±12.2 mmHg;

Figure 5) from baseline to exercise which was maintained through 8-10 minutes of

recovery (+7.9±6.5 mmHg and +12.4±9.4 mmHg, respectively; p<0.01), and a return to

baseline following exercise. The same was observed for DBP (F3,15=7.375), which

significantly increased by 12.8±6.9 mmHg from baseline to exercise, and remained

elevated by 5.6±5.7 mmHg through the first recovery (p<0.02 for all comparisons). HR

was only significantly different at the first recovery compared to baseline (F3,24=3.330; -

2±2 bpm; p<0.04).

When examining each acute exercise bout, results indicated significant main

effects of condition (F1,8=136.11, p<0.01) and time (F6,30=3.876, p<0.01) for MAP.

MAP significantly increased during the second and third bouts of knee extension in the
 55

control condition compared to baseline (+22.5±23.7 mmHg and +23.6±19.6 mmHg,

respectively), and during the first and second bouts of knee extension in the BFR

condition (+19.8±17.1 mmHg and +17.7±16.5 mmHg, respectively; p<0.02 for all

comparisons; Figure 4). MAP also remained elevated compared to baseline through the

second and third rest periods in the control condition (+10.8±12.9 mmHg and +13.9±10.3

mmHg, respectively; both p<0.04).

Results indicated significant main effects of condition (F1,8=35.981, p<0.01) and

time (F6,30=3.797, p<0.01) for SBP. SBP significantly increased during the second and

third bout of knee extension in the control condition compared to baseline (+28.3±25.9

mmHg and + 32.3±22.6 mmHg, respectively), and remained elevated throughout their

respective rest periods (+16.6±21.0 mmHg and +19.6±18.6 mmHg for set 2 rest and set 3

rest, respectively; all p<0.05; Figure 6). In the BFR condition, SBP significantly

increased during the first and second set of knee extension compared to baseline

(+25.9±19.9 mmHg and +22.5±20.6 mmHg, respectively, p<0.02; Figure 6), but was not

significantly different from baseline during either of the rest periods. Only a significant

main effect of time was observed for HR (F6,30=4.117, p<0.01), which is explained by a

significant increase in HR during the first exercise bout (+3.3±3.0 bpm, p<0.02)

compared to baseline. No differences were observed for DBP.

Oxyhemoglobin

Results indicated a significant condition by time interaction (F3,21=6.315, p<0.01)

for Oxy. This was explained by an increase in Oxy during exercise in the BFR condition

and a simultaneous decrease in Oxy during exercise in the control condition (+12.3±96.7
 56

NU in BFR and –56.2±82.37 NU in control, p<0.04; Figure 7). There was also a

significant condition by time interaction when Oxy was compared across each acute bout

of exercise (F6,30=5.539, p<0.01). This was explained by a decrease in Oxy during each

set of exercise in control condition (-74.2±96.2 NU, -85.5±99.4 NU, and -93.0±99.0 NU

for sets 1, 2, and 3 respectively; Figure 8), a significant increase in Oxy from exercise to

rest for all bouts of exercise in the control condition and bouts 2 and 3 in the BFR

condition, and a significantly lower Oxy across all sets of exercise in the control

condition compared to the BFR condition (peak difference= 88.60±73.2 NU during the

first rest period; all p<0.05).

Deoxyhemoglobin

Results indicated a significant condition by time interaction for Doxy

(F3,21=13.652, p<0.01). This is explained by a significant increase in Doxy during

exercise compared to baseline in the BFR condition (+105.4±76.7 NU), and a

significantly higher Doxy during BFR compared to control exercise (105.4±76.7 NU and

6.1±28.8 NU in BFR and control respectively, all p<0.01; Figure 9). There was also a

significant condition by time interaction when Doxy was compared across each acute

bout of exercise (F6,48=11.026, p<0.01). This was explained by significant increases in

Doxy during rest periods compared to baseline in the control condition (+20.9±27.5 NU,

+25.6±33.6 NU, and +24.7±31.6 NU, respectively), a significant increase during the first

rest period in the BFR condition that was maintained throughout the entire exercise bout

(peak difference= 153.5±86.8 NU during the second rest period), a significant increase in

Oxy from exercise to rest for all bouts of exercise in the control condition and bouts 1
 57

and 2 in the BFR condition, and significantly greater Doxy during each set and rest in the

BFR condition compared to the control condition (peak difference= 127.8±74.7 NU

during the second rest period, p<0.05 for all comparisons; Figure 10).

Total hemoglobin

Results indicated a significant condition by time interaction for TotalHb when

compared across each experimental condition (F3,21=13.854, p<0.01), which is

contributed to a significant difference between each condition during exercise (-

50.0±113.2 NU and 100.43±173.5 NU during exercise in the control and BFR conditions,

respectively; all p<0.05; Figure 11). Results also indicated a significant condition by

time interaction when TotalHb was compared across each acute bout of exercise

(F6,48=8.872, p<0.05). TotalHb significantly increased from each set of exercise to its

subsequent rest period in both conditions (Peak differences= +117.23±70.2 NU and

+98.1±67.4 NU for the first and third rest period in Con and BFR, respectively), and a

significantly higher TotalHb in the first set of BFR knee extension through the third set of

BFR knee extension compared to the control condition (P<0.05 for all comparisons;

Figure 12).

Tissue oxygenation index

Results indicated a significant condition by time interaction for TOI when

compared across the entire session (F3,21=6.571, p<0.01; Figure 13), which is attributed to

a significantly greater decrease in TOI during BFR exercise compared to control exercise

(-2.4±1.8% and -6.5±3.0% for control and BFR exercise, respectively; all p<0.01). When

compared across each acute bout of exercise, results indicated a significant condition by
 58

time interaction for TOI (F6,48=6.887, p<0.01). TOI significantly decreased from the first

rest period throughout the exercise bout compared to baseline in the control condition

(peak difference= -3.2±2.4 % during the second set), a significant decrease in TOI

throughout the entire exercise bout compared to baseline in the BFR condition (peak

difference= -8.0±3.5% during the third set), and a significantly lower TOI across the

entire exercise bout in the BFR condition compared to control (p<0.03 for all

comparisons; Figure 14).

Protocol 2

In protocol 2, 2 out of the 3 subjects were able to perform both walking trials

(BFR and control) with no difference in distance walked and without exhibiting any signs

or symptoms of acute AD or DVT formation. However, the third subject presented with

significant functional limitations and was unable to complete either of the walking trials

in their entirety. Specifically, this subject walked for a total of 317 seconds during the

control condition (performed first), and only 193 seconds during the BFR condition

(performed second). Also, while the subject did not report any pain in either condition,

there was a moderate increase in reported difficulty between the two conditions (5.3 cm

and 5.6 cm for the first bout and 9.3 cm and 9.7 cm for the last bout in control and BFR,

respectively). Furthermore, spasticity marginally decreased following BFR exercise in

this subject. Finally, this subject did not present with any signs or symptoms of acute AD

or DVT formation.
 59

Discussion

The purpose of the present study was to determine the safety and feasibility of a

single session of BFR exercise in a sample of individuals with chronic iSCIs, with special

attention paid to the risk of acute AD and DVT formation. We also aimed to determine

how muscle activation, lactate formation, hemodynamics, and local tissue oxygenation

respond to a single session of BFR knee extension exercise compared to controlled

exercise. To our knowledge this is the first investigation into the effects of large muscle

mass blood flow restricted exercise in individuals with incomplete spinal cord injuries.

Ultimately, our data suggest that a single session of BFR exercise can be safely

performed in individuals with incomplete spinal cord injuries. Information gained from

such an investigation may provide a foundation for future research focused on the

potential benefits of blood flow restricted exercise in this population, which has the

potential for enhancing recovery and improving quality of life.

Safety and pain

Our main hypothesis was confirmed by our data, as no subjects experienced any

signs of acute AD or DVT formation, or any decrements in total work, in response to

BFR knee extension exercise. To our knowledge, we are the first to report on the risks of

AD and DVT with BFR exercise in individuals with iSCI. It is important to note that

while we are confident that acute BFR exercise can be safely performed in individuals

with iSCI, this investigation was conducted in a controlled environment under the direct

supervision of licensed physical therapists and other clinical staff. Therefore, we

recommend that any such exercise be performed in the same controlled environment with
 60

continuous blood pressure and heart rate monitoring. We also feel that a follow-up study

is necessary to determine the long-term safety and efficacy of BFR exercise in this

population. While we found that a single session of BFR exercise does not elicit DVT

formation, we cannot make inferences regarding the effects of multiple sessions of BFR

exercise.

We also did not observe any differences in pain or perceived difficulty between

BFR or control knee extension exercise. This deviated from our hypothesis and from

previously published literature reporting increases in pain and difficulty with BFR

exercise (Fitschen et al., 2014; Rossow et al., 2012). However, it is important to note the

difference in cuff pressure used in this study and in previous research. In a conservative

effort, we only used an occlusion pressure of 125% of the venous occlusion pressure in

the present study. This was sufficient in occluding venous flow at rest, which is the basis

of BFR exercise. In contrast, other research in the general population uses a variety of

cuff pressures, including near- or supra-systolic cuff pressures. This could potentially

have influenced the lack of a difference in pain and perceived difficulty between BFR

and control exercise. Another potential influencing factor would be the lack of afferent

feedback due to the injury itself. Depending on the level of injury, the ability to feel pain

or discomfort in the exercising leg may be diminished, thereby eliminating any effect

BFR exercise might have on pain or difficulty. These data support the use of BFR

exercise within a physical rehabilitation program, as pain and difficulty level are both

limiting factors to any exercise modality. However, it is important to note that

individuals who lack afferent feedback from the exercising muscle, specifically
 61

nociceptive responses, would be unable to recognize when an occlusion pressure is too

great. This is another reason that the authors recommend that any BFR exercise in this

group be performed in a controlled environment and be supervised by a licensed therapist

or physician.

Physiological variables

In contrast to previous research, we did not observe any significant differences in

muscle activation between BFR and control exercise (Loenneke et al., 2015; Yasuda et

al., 2014). This might be explained by an injury-related limited muscular activation in

these subjects, effectively placing a ceiling on muscular activation. For example, as

muscle fibers fatigue in able-bodied individuals, EMG activity increases to recruit more

unfatigued fibers. In the individuals with iSCIs, the ability to recruit more fibers is likely

limited. The magnitude of this limitation would be dependent on the level and

completeness of each subject’s injury, and therefore is likely exposed to a very high

degree of inter-individual variation. Similar to muscular activation, the pressor response

was not augmented during BFR as we had hypothesized. Instead, MAP (Figure 3) and

SBP (Figure 5) both increased in response to exercise in both conditions with no main

effect of condition, as was HR. Our reasoning for hypothesizing that BFR exercise

would elicit a greater blood pressure response was due to an elevated activation of Group

IV muscle afferents in response to an elevated metabolite accumulation, and greater

fatigue induced central command. However, as evident by our reported lack of any

significant difference in whole blood lactate between conditions, BFR exercise did not

seem to elevate whole body metabolite accumulation. These results conflict with prior
 62

research reporting a potentiation of the pressor response (indicated by increases in blood

pressure) in matched-workload BFR exercise (Brandner, Kidgell, & Warmington, 2015;

Bunevicius et al., 2016; Poton & Polito, 2015), which may be attributable to a lower

absolute exercise intensity used in this study. The absence of a difference in lactate

between the two conditions also conflicts with some prior research, (Lundberg, Olofsson,

Ungerstedt, Jansson, & Sundberg, 2002) and is supported by others (Loenneke, Thrower,

Balapur, Barnes, & Pujol, 2012). This may also be due to a limited exercise intensity, as

well as a limited exercise volume performed in this protocol. However, it is important to

note that whole blood lactate may not reflect all of the lactate accumulated in the muscle

itself, especially in individuals with impaired circulation. Ultimately, these data suggest

that light-load BFR exercise does not elicit a significantly higher cardiovascular or

metabolic strain compared to traditional exercise in individuals with iSCIs.

Another interesting finding from the present study was that the application of

BFR elicited an increase in local tissue perfusion. This was evident by increases in Oxy

(Figures 7 and 8), Doxy (Figures 9 and 10), and TotalHb (Figures 11 and 12) with BFR

exercise compared to control exercise. This suggests that the pressure used in this study

(125% of venous occlusion pressure) was sufficient in occluding venous blood flow

while maintaining arterial flow, and supports the assumption of localized metabolite

accumulation. This effectively created a “reserve” of oxygenated blood in the occluded

limb. However, this reserve was depleted during each set of exercise in the BFR

condition, suggesting that the action of the muscle pump was sufficient in forcing blood

through the occluded vein. This heightened perfusion in response to BFR exercise may
 63

also influence cell swelling, which is one of many proposed mechanisms of BFR induced

hypertrophy (Lang et al., 1998; Low, Rennie, & Taylor, 1997). Future researchers may

consider exploring the effects of BFR exercise and possible cell swelling in this

population. Additionally, muscle tissue oxygenation index (TOI) significantly decreased

during the BFR condition, indicating a greater oxygen depletion and deoxygenated

hemoglobin accumulation during BFR exercise. During the recovery period, however,

perfusion and TOI both returned to near-baseline values. Taken together, these data

indicated that the application of 125% of venous occlusion pressure effectively restricts

blood flow during exercise, and elicits a significant reduction in overall tissue

oxygenation.

Limitations

As is the case with any project, this study has certain limitations. One limitation

to this study is that the exercise workloads were set subjectively, and not as a percentage

of maximal effort. Therefore, it is difficult to know how these exercise intensities

compare to those in previous published studies in the general population. This may have

also contributed to a lack of any significant difference in muscular activation and lactate

accumulation. Future research may consider exploring the effect of varying exercise

intensities in this population, as has been done in the general population. Another

limitation to the current study is that our sample carried a high degree of variability,

specifically related to injury level and completeness of injury. However, we opted to

include a range of injury levels and degrees of injury in order to determine the safety of

BFR exercise in a representative sample of individuals with iSCIs. Future researchers

 64

may consider comparing the safety, feasibility, and physiological responses to BFR

exercise between different levels of injury and ASIA classifications.

In conclusion, results from this study suggest that a single session of controlled

BFR exercise can be safely performed by individuals suffering from iSCIs without added

cardiovascular strain or heightened pain. However, we recommend that any such

exercise be performed in a supervised and controlled environment while research

continues to expand in this area, especially during walking exercise and when working

with individuals with significant functional impairments.

 CHAPTER V

GRANT SUBMISSION: BLOOD FLOW RESTRICTION TRAINING IN

INDIVIDUALS WITH INCOMPLETE SPINAL CORD INJURIES

In accordance with the Neilsen Foundation requirements, the following four

sections comprise a "biosketch" that has been formatted to the standards of the National

Institutes of Health.

Personal Statement

Over the past three years my area of research has focused on the acute effects of

exercise on cardiovascular function and blood flow characteristics in healthy individuals

and individuals with complete and incomplete spinal cord injuries. I have also had the

opportunity to conduct research in the areas of environmental physiology, cognitive

function, and metabolism. My most recent projects include research on the acute effects

of cuff size and pressure on blood flow characteristics and fatigue during BFR exercise in

healthy individuals; and the safety and feasibility of a single set of BFR exercise in

individuals with incomplete spinal cord injuries. My research methodologies generally

include the use of Doppler ultrasound imaging, near-infrared spectroscopy (NIRS), and

other basic physiological measurements such as electromyography (EMG), beat-by-beat

blood pressure analysis, and muscular function assessments. I am currently authored on a

peer-reviewed publication in Aerospace Medicine & Human Physiology, and have

multiple other first-author publications under review in a variety of refereed journals. In

65
 66

addition, I have presented at multiple conferences per year for the last three years,

including the American College of Sports Medicine regional and national conferences,

regional exercise physiology conferences, and university symposia. Finally, I have also

conducted research at the Louis Stokes' Cleveland VA Medical Center under Dr. John

McDaniel, and in collaboration with licensed physical therapists.

Positions and Honors

I am currently a PhD candidate at Kent State University, where I also conduct

research focused on vascular function and teach courses in exercise physiology as part-

time faculty. As a result of my research at Kent State, I have received the honors of most

outstanding graduate presenter at the 2016 Northeastern Ohio Exercise Science

Conference and most outstanding presenter at the 2017 Kent State University Graduate

Student Symposium. I also held appointments at Walsh University, where I taught

courses in anatomy and physiology, and at the Louis Stokes' Cleveland VA Medical

Center, where I conduct research under the direction of Dr. John McDaniel. The research

being conducted at this institution is directed at improving the quality of life of

individuals with complete and incomplete spinal cord injuries.

Contributions to Science

As a result of my research at Kent State University, I have been able to contribute

to the understanding of blood flow regulation during and following exercise. One of my

recent projects helped to elucidate the independent influences of metabolic cost and force

production on post exercise hypotension. Specifically, we found that higher

metabolically demanding concentric exercise potentiates the post exercise hypotensive

 67

response to a bout of moderate intensity resistance exercise, while less metabolically

demanding eccentric exercise elicits a period of elevated blood pressure. This expands

our understanding of the factors influencing post exercise blood pressure regulation. This

paper is currently under peer review. Another project from our laboratory focused on

improving lower leg blood flow during exercise in individuals with complete spinal cord

injuries. In this project, individuals with complete spinal cord injuries performed arm

crank exercise with period of combined passive limb movement. This study found that

while passive limb movement and upper body exercise both increased femoral blood

flow, the additive effective of the combined arm exercise with passive limb movement

was negligible.

Additional Information: Research Support

Throughout my time at Kent State University, I have also aided a number of other

projects not related to vascular function or spinal cord injuries, and I have helped prepare

IRB submissions for research projects. These projects have included: 1) determining the

effect of inspiratory resistance on cognitive function and cycling performance in varying

levels of normobaric hypoxia, 2) determining the influence of moderate cycling during

exposure to moderate normobaric hypoxia on cerebral oxygenation, perfusion, and

cognitive function, and 3) understanding the effects of varying cuff widths and pressures

on blood flow and fatigue during lower body and upper body blood flow restriction

exercise. Finally, I have helped mentor multiple graduate students throughout their

studies and the research process.

 68

Overall Objective

This section will detail the primary objectives of the proposed investigation.

Specific aims

This study has two specific aims that together will achieve the singular goal of

determining the efficacy and feasibility of blood flow restriction (BFR) exercise in

individuals with incomplete spinal cord injuries (iSCIs). Each of the specific aims is

discussed in detail below.

Specific aim 1. The first specific aim of this study is to compare muscle function

and fatigue characteristics between a single session of BFR and non-BFR exercise.

Specifically, the first aim of this study is to compare the effects of BFR and non-BFR

exercise on iEMG, corticomotor evoked potentials (CMEP), peripheral nerve motor

evoked potentials (PMEP), central and peripheral voluntary activation (CVA and PVA,

respectively), and muscular work between individuals with high level iSCIs (C2-C6), low

level iSCIs (C7-L3), and healthy age-matched controls. Understanding how BFR

exercise impacts fatigue in this population may provide insight regarding its acute effects

on the neuromuscular system and its potential efficacy. For our first specific aim, we

hypothesize that a single session of BFR exercise will potentiate peripheral fatigue while

having no effect on corticomotor excitability compared to control exercise in individuals

with iSCIs.

Specific aim 2. The second scientific aim focuses on the long-term safety and

efficacy of BFR exercise in individuals with iSCIs. Specifically, we intend to monitor

the risk of acute autonomic dysreflexia (AD) and deep vein thrombosis (DVT) formation,
 69

and compare measures of muscular strength, size, and functional outcomes before and

after a 6-week BFR exercise program. For our second specific aim, we hypothesize that

6 weeks of BFR exercise will result in increases in muscular strength, cross sectional

area, endurance, and improved functional outcomes compared to 6 weeks of non-BFR

exercise in individuals with iSCIs. We also hypothesize that no subjects will present with

signs or symptoms of acute AD or DVT formation in response to BFR exercise.

Introductory statement and background

This section will provide a brief background and introduction to both protocols

proposed under this investigation.

Protocol 1. In the general population, BFR exercise has been shown to accelerate

peripheral fatigue, as indicated by progressively increasing muscular activation during

submaximal exercise (Yasuda, Loenneke, Ogasawara, & Abe, 2013), reduced force

production (Cook, Clark, & Ploutz-Snyder, 2007), and decreased muscular work

(Yasuda, Fukumura, Iida, & Nakajima, 2015) compared to non-BFR exercise.

Corticomotor excitability has also been shown to increase for up to sixty minutes

following continuous BFR exercise compared to non-BFR exercise of the same intensity,

which is suggested to be due to an increase in Group III and IV afferent activation

(Brandner, Warmington, & Kidgell, 2015). Taken together, these data suggest that the

application of BFR during low intensity exercise causes significant muscular fatigue,

resulting in an increase in cortical activation of the active muscle in order to produce the

same force.
 70

Individuals with iSCIs are generally limited to low-intensity exercise due to

diminished neuromotor function. Due to this, peripheral fatigue is attenuated following

exercise in individuals with iSCIs compared to healthy age-matched controls (Lin, Chen,

Luh, Wang, & Chang, 2012). This suggests an imbalance between central and peripheral

fatigue, which might be corrected with the application of BFR. Additionally, reduced

inhibitory activity the GABAergic neuron in high-level iSCIs reduces second-order

inhibition of efferent motor activation, evident by a decreased cortical silent period and

end latency during exercise induced fatigue (Nardone et al., 2013). This is a similar

effect as that observed with BFR exercise in the general population. However, the effect

of BFR exercise on measures of central and peripheral fatigue has not yet been

investigated in individuals with iSCIs.

Protocol 2. In the general population, BFR exercise has been shown to elicit

muscular adaptations to low-intensity resistance and walking exercise that are similar to

those observed following high-intensity exercise. These included increases in muscular

strength, endurance, size, and function (Abe et al., 2010; Abe et al., 2006; Barcelos et al.,

2015; Loenneke et al., 2016; Shinohara et al., 1998). Some of the proposed mechanisms

for this effect include cell swelling, increased growth hormone signaling, increased

peripheral fatigue, preferential type II fiber recruitment, increased mitochondrial density,

angiogenesis mediated increases in perfusion, and increased metabolite accumulation

(Fujita et al., 2007; Larkin et al., 2012; Loenneke et al., 2010; Moritani et al., 1992;

Wernbom et al., 2009).

 71

As stated previously, individuals suffering from iSCIs experience reduced

voluntary activation, which limits exercise intensity; thereby limiting physical therapy

outcomes. For this reason, an exercise modality that maximizes muscular adaptations to

low intensity exercise, such as BFR exercise, would be ideal in this population.

However, there are some safety considerations regarding BFR exercise in individuals

with iSCIs. For one, the occlusion pressure and related pain response has the potential to

elicit acute AD. Recently, our laboratory has conducted a pilot study in which 9

individuals with iSCIs performed a single set of knee extension exercise with and without

BFR. No subjects presented with acute AD or dangerous elevations in blood pressure in

either of the trials. Another safety consideration for BFR in this population is the risk of

DVT formation. In the same study subjects were screened for a pre-existing DVT at

baseline, and for elevated D-dimer (a protein-fragment marker of fibrinolysis) following

BFR exercise; and a follow-up ultrasound DVT scan was performed to rule out DVT if

D-dimer levels were elevated (>500 ng/mL). No subjects presented with post-exercise

DVT formation. While these data do support the safety of acute BFR exercise in

individuals with iSCIs, to our knowledge there are no studies focused on the safety of

chronic BFR training in individuals with iSCI.

Preliminary data

In this section, preliminary data are presented for each of the proposed protocols.

Specific aim 1. Currently, there is very limited research on the effects of BFR

exercise on muscle function and fatigue in individuals with incomplete spinal cord

injuries. In fact, to our knowledge our laboratory has conducted the only investigation
 72

into the feasibility of large-muscle mass BFR exercise in individuals with iSCIs. This

study was primarily focused on the safety of BFR exercise in these individuals, and

therefore the only muscle-function related variable that was collected was iEMG. No

significant differences were reported in iEMG between BFR and non-BFR exercise in

these individuals. This contrasts with BFR research in healthy populations that generally

report progressive increases in EMG across repetitions of BFR exercise (Yasuda et al.,

2014). This suggests that the peripheral fatigue-mediated increase in central activation

may not be present in individuals with neuromotor impairment. However, more data are

needed to understand how peripheral and central fatigue are impacted by BFR in

individuals with iSCIs.

Specific aim 2. Currently, there is only one study that reports on the benefits of

regular BFR training in individuals with spinal cord injuries. Gorgey et al. (2016)

examined the effect of six weeks of twice weekly combined BFR and functional

electrical stimulation (FES) wrist extensor exercise in 9 males with incomplete

tetraplegia. The increase in CSA was 17% greater following combined BFR and FES

exercise than FES exercise alone. While this investigation promotes the use of BFR in

individuals with SCIs, this investigation used a small muscle mass, and was conducted

with combined FES rather than voluntary exercise. The proposed investigation would

expand on this line of inquiry, and be the first to report on the regular use of BFR

exercise during large-muscle mass exercise in a group of individuals with iSCIs.

Regarding safety, our laboratory recently completed a feasibility study examining the risk

of acute AD and DVT formation in response to one bout of unilateral knee extension
 73

exercise with the application of 125% venous occlusion pressure; and found that BFR did

not elicit any significant risk of acute AD or DVT formation.

Rationale

This section describes the overall importance of the proposed investigation.

Specifically, this sections explains the potential short term, and long term benefits of the

proposed investigation.

Significance

In recent years, investigators started exploring new methods of improving

physical rehabilitation from complete and incomplete spinal cord injuries. These include

water-based physical therapy (Stevens et al., 2015), body weight supported treadmill

training (Cayot, Lauver, Silette, & Scheuermann, 2016; Giangregorio et al., 2005),

robotic assisted treadmill training (Del-Ama et al., 2014; Gorman et al., 2016), and more.

In general, these therapies helped to improve function, gait, and restore independence in

individuals with iSCIs. However, these therapies are limited in intensity due to impaired

neuromotor control. Therefore, an exercise modality that can maximize outcomes from

limited exercise intensities would be of critical importance in this population. The

ultimate goal of this investigation is to explore the safety and efficacy of BFR exercise as

a method of enhancing the muscular outcomes of low-intensity exercise in individuals

with iSCIs. The significance and potential impact of each specific aim of this

investigation is discussed below.

Scientific aim 1. Understanding fatigue is important to understanding the effects

of a mode of exercise. For example, research indicates that exercise related fatigue is
 74

primarily centrally mediated (i.e. neural drive) as opposed to peripherally mediated (i.e.

localized muscle fatigue) in individuals with iSCIs (Lin et al., 2012). Research also

suggests that elements related to peripheral fatigue, such as mechanical distortion of the

muscle fiber, metabolite accumulation, and muscle damage are necessary for exercise

related improvements in muscular force production and CSA (Rooney, Herbert, &

Balnave, 1994; Toigo & Boutellier, 2006; Vandenburgh & Kaufman, 1979). Therefore,

heightened central fatigue may limit muscular adaptations to exercise in individuals with

iSCIs. In contrast, BFR exercise has been shown to accelerate peripheral fatigue

(Brandner, Warmington, et al., 2015; Cook et al., 2007), which could potentially correct

this imbalance in central and peripheral fatigue in individuals with iSCI. This first aim of

this investigation will explore the impact of BFR exercise on central and peripheral

fatigue in individuals with high- and low-level iSCIs. Results from this study may be

useful for future research focused on the implementation of BFR exercise in iSCI therapy

programs.

Scientific aim 2. The second specific aim of this study focuses on the potential

long-term safety and efficacy of BFR training in individuals with iSCIs. As mentioned

earlier, in recent years investigators and clinicians have begun exploring the efficacy of

modified exercise therapies in individuals with complete and incomplete spinal cord

injuries. While many of these therapies have proven effective in restoring a moderate

level of function, they are subject to limited exercise intensity. BFR exercise may be a

method of circumventing that limitation, and significantly restoring muscle strength, size,

and overall function in individuals with iSCIs. There are some safety considerations
 75

associated with BFR exercise in individuals with iSCIs, which is also a focus of the

second aim of this investigation. Ultimately, results from this study may support the

application of BFR training as a method for enhancing muscular adaptations to physical

rehabilitation in individuals with chronic iSCIs.

Relevance to the Neilsen Foundation’s mission

The main goal of this investigation is to explore a new and potentially improved

method of exercise therapy for individuals suffering from iSCIs. The results from this

investigation may ultimately aid in enhancing recovery from an iSCI and improve quality

of life. This is very relevant to both the Neilsen foundation's mission of "supporting both

programs and scientific research to improve the quality of life for those affected by and

living with a spinal cord injury," and vision for individuals who care for or live with

spinal cord injuries to "live full and productive lives as active participants in their

communities." Ultimately, we hope that the results of this investigation provides a

foundation for research and clinical practice that can eventually improve the lives of

individuals living with iSCIs across the global community.

Research Plan

This section describes the research plan, including how and when data will be

collected, for the proposed investigation.

Scientific aim 1. The first aim of this investigation will include two separate

experimental protocols performed in counterbalanced order (Figure 15). In addition to

the two experimental protocols, subjects will undergo a bilateral leg ultrasound DVT scan
 76

to rule out the an existing DVT. Each session will follow the same experimental

protocol, with the exception of the type of exercise being performed. In each session,

subjects will arrive to the physiology lab three hours postprandial and having abstained

from caffeine and alcohol for eight hours, and major physical activity for the previous

twenty-four hours. Upon entering the laboratory, subjects will be comfortably seated in

an isokinetic dynamometer (Humac Norm, CSMi, Stoughton, MA). Following

appropriate hair removal and skin abrasion, 8 mm EMG electrodes will be placed over

the rectus femoris (midway between the spina iliaca superior and the superior portion of

the patella) and the vastus lateralis (two thirds of the way between the anterior spina

iliaca superior and the lateral side of the patella) on the subject's dominant leg. EMG

signals will be grounded on the ipsilateral patella. Subjects will also be prepped with a 3-

lead ECG (ADInstruments, Colorado Springs, CO), a beat-by-beat blood pressure

monitor (Nexfin, Edwards LifeScences, Irvine, CA), and near-infrared spectroscopy

optodes (Oxymon MKIII, Artinis, Elst, the Netherlands) superior and medial to the

patella on the vastus medialis oblique. Subjects will then perform three isometric

maximal voluntary contractions (MVCs) of the dominant leg. The peak force will be

recorded, and used for calculation of exercise intensity. Next, a tight-fitting lycra cap

will be place over the top of the subject's head in reference to the Nasion-Inion line and

the pre-auricular point. A 9 mm figure-eight magnetic coil (Magstim 2002 stimulators,

Magstim Co Ltd, Spring Gardens, Whitland, UK) will be used to stimulate a maximal

isometric contraction of the knee extensors (assessed by the isokinetic dynamometer) of

the subject's dominant leg via the contralateral primary motor cortex (M1). When an
 77

optimal transcranial magnetic stimulation (TMS) intensity and location on the

contralateral M1 are identified, that location will be marked on the lycra cap. An

appropriate TMS intensity will also be identified by repeatedly stimulating knee extensor

contractions at the contralateral M1 with graduating intensity (starting <100 µV) until a

plateau in knee extension force is observed. Finally, a 10 cm wide pneumatic cuff (DE

Hokanson, Bellevue, WA) will be secured over each the proximal portion of each thigh.

This will complete preparation for data collection in each visit.

Once all subjects are prepped for data collection, baseline MVCs, motor evoked

potentials (MEPs), and central and peripheral voluntary activation (CVA and PVA,

respectively) will be collected. This will require subjects to perform three separate

sustained (three second) isometric MVCs of the dominant leg, while a superimposed

twitch is applied during the third second via TMS. MEP will be reported as both the peak

force and peak iEMG following TMS activation. MVC will be reported as the peak force

of the first two seconds; and central voluntary activation will be calculated as 100*(1-

(superimposed twitch / MEP)). CMEP, MVC, iEMG, and CVA will each be reported as

an average of the 3 trials. Next, the magnetic coil will be moved from the contralateral

M1 to the femoral nerve of the dominant leg. Subjects will perform three more sustained

isometric contractions followed by a superimposed twitch, which will be applied at the

femoral nerve. Peripheral motor evoked potentials (PMEP) and PVA will both be

calculated and reported as averages over the 3 separate trials. In order to account for any

possible order effect, the order of central or peripherally applied magnetic stimulation

will be counterbalanced between subjects (however, it will remain consistent within

 78

subjects). Next, subjects will either perform BFR or control exercise (counterbalanced by

condition; Figure 15). Occlusion pressure will be set at 125% of venous occlusion

pressure, validated at the popliteal vein via Doppler Ultrasound (GE Logic 7, GE

Healthcare, Milwaukee, WI). Each exercise session will consist of 3 sets of 10

repetitions of isokinetic bilateral knee extension at 120° per second. Torque will be

recorded for each repetition by the isokinetic dynamometer, and reported as an average

for each set. Following exercise, MVC, CMEP, PMEP, CVA, and PVA will be recorded

a second time. This will conclude each experimental visit. Three to 7 days following

each experimental visit, subjects will return for a 5.0 mL blood draw that will be

analyzed for quantitative D-dimer levels. Any subjects who present with abnormally high

(>500 ng/mL) will undergo a second bilateral leg ultrasound DVT scan.

Scientific aim 2. The second aim of this investigation is a training study, and

includes bi-weekly outcome assessments. This research protocol for this aim is presented

in Figure 16. Prior to participation in this study, each subject will be prescreened for an

existing DVT. Once a DVT is ruled out, baseline data will be collected for each subject.

This will include a walking gait analysis (performed by a licensed physical therapist),

isometric and isokinetic (1 per second) MVCs of each leg, timed-up-and-go (TUG)

scores, 6-minute walk test scores, and MRI assessments of quadriceps and hamstrings

CSA. Isometric and isokinetic knee extensor torques will be assessed with an isokinetic

dynamometer, and muscle CSA will be evaluated in 2 mm thick slices taken at the widest

portion of the thigh. After all baseline data are collected, subjects will participate in 6

weeks of twice-weekly exercise training. This exercise training will be supervised by a

 79

licensed physical therapist, and will include a combination of gait training (over ground

and treadmill walking) and lower-body resistance exercise. Lower-body resistance

exercise will include knee extension, flexion, and leg press; and each exercise will be

performed with the goal of achieving 3 sets of 10 repetitions separated by 90 seconds

rest, and the exercise load will be increased by 5% when subjects are able to comfortably

perform the desired repetitions. Both groups (high level iSCI and low level iSCI) will be

stratified by age into a control or experimental group. The control and experimental

groups will perform the same exercises, with the only difference being the application of

BFR in the experimental group. Occlusion pressure will be applied with the use of an

automated pressure cuff set to 125% of venous occlusion pressure (validated at the

popliteal vein via Doppler ultrasound), which will be on a mobile cart that can move with

each subject around the fitness facility. Cuffs will be inflated at the onset of each set of

resistance exercise and each bout of walking exercise, and will be deflated during rest

periods between sets and between exercises. D-dimer will be assessed at the end of each

week, and bilateral leg ultrasound DVT scans will be performed for any subject whose D-

dimer level is returned above 500 ng/mL. TUG tests, 6-minute walk tests, gait analyses,

and isometric and isokinetic muscle function testing will be performed 24-48 hours after

the final exercise session each second week (weeks 2, 4, and 6). MRI assessment of

quadriceps and hamstrings CSA will also be performed pre and post training.

Human subjects

This study includes two separate experimental protocols. The first protocol will

follow a 2 group by 2 condition by 4 time point multifactorial design; and the second
 80

protocol will follow a 4 group by 2 condition by 4 time point multifactorial design.

Based on recent pilot data collected in collaboration with the Louis Stokes' Cleveland VA

Medical Center’s Spinal Cord Injury and Disease Center, 16 subjects would need to be

recruited in order to achieve statistical power (1-β= .90) for the first protocol, and 32

subjects will need to be recruited for the second protocol. Therefore, 60 subjects will be

recruited for participation in this study to account for subject drop-out (20 for the first

protocol, and 40 for the second protocol). Groups will be stratified into individuals with

high-level iSCIs (C2-C6) and low-level iSCIs (C7-L3); and these groups will be further

stratified into age-matched control and experimental groups in the second protocol.

However, subjects will be encouraged to participate in both experimental protocols,

which would reduce the requirement for subject recruitment.

Each of the control groups will participate in 6 weeks of twice per week physical

rehabilitation with no modifications to their exercise program. The experimental groups

will also participate in 6 weeks of twice per week physical rehabilitation with the addition

of BFR. All subjects will have an MRI scan of the widest portion of the thigh for

assessment of rectus femoris, vastus lateralis, and biceps femoris CSA at baseline and

week 6. D-dimer will be assessed at the end of each week, and a bilateral leg ultrasound

DVT scan will be performed if any readings are abnormally high (>500 ng/mL).

Subject recruitment and inclusion

Subjects will be recruited from surrounding medical centers and fitness facilities

that cater to spinal cord injuries. To start, we will reach out to contacts at the Louis

Stokes' Cleveland VA Medical Center where there is an existing subject pool. This is a
 81

full-service medical center where we have existing relationships with the radiology

department; which would allow us to schedule MRI screenings for each of our subjects.

We will also reach out to existing contacts at a local spinal cord injury specific wellness

center. From there, we intend to contact local medical centers and physical therapy

programs for subject recruitment.

All subjects in this study will be diagnosed with chronic incomplete spinal cord

injuries. A pre-screening bilateral leg ultrasound DVT scan will be performed on each

subject to rule out an existing DVT. Subjects will also have no recent history of MI or

DVT (< 3 years), no existing peripheral artery disease, no existing coronary artery

disease, uncontrolled hypertension, or other musculoskeletal injuries that would preclude

participation in regular exercise.

Data collection

In the proposed study, data will be collected by a variety of methods. Quadriceps

and hamstrings muscle cross sectional area (CSA) will be evaluated using MRI. To get

the most accurate measurement of muscle CSA, muscles will be imaged at the widest

portion of the thigh. MRI scanning will occur at the Louis Stokes' Cleveland VA

Medical Center. Bilateral leg DVT scans will be performed by the investigators using the

compression method, and will be performed via Doppler Ultrasound (GE Logic 7, GE

Healthcare, Milwaukee, WI). Central and peripheral voluntary activation and

corticomotor excitability assessments will be performed by trained investigators using

transcranial magnetic stimulation (TMS; Magstim 2002 stimulators, Magstim Co Ltd,

Spring Gardens, Whitland, UK). To acquire a MEP, a magnetic coil will be placed over
 82

the optimal site of the contralateral M1 to elicit activation of the knee extensors. To

assure consistency in TMS activation, each subject will wear a tight fitting lycra cap that

will be positioned in reference to the Nasion-Inion line and the pre-auricular point, and

marked at the desired location for magnetic stimulation. Central MEP will be recorded as

the force and iEMG produced during transcranial stimulation while resting, and

peripheral MEP will be recorded as the force and iEMG produced during femoral nerve

stimulation while resting. Central and peripheral voluntary activation will both be

assessed by applying TMS during a maximal voluntary contraction; and will be

calculated as 100*(1-(superimposed twitch / MEP)). Surface EMG will collected from

the vastus lateralis and rectus femoris using 9 mm gel electrodes, and will be amplified

(1000), bandpass filtered (13 Hz and 1000 Hz), and sampled at 2 kHz during voluntary

and evoked contractions. EMG and ECG will both be collected and analyzed offline with

a multi-channel data acquisition system (ADInstrument, Colorado Springs, CO).

Oxyhemoglobin saturation, deoxyhemoglobin saturation, total hemoglobin, and tissue

oxygenation will also be collected (NIRS, Oxymon MK III, artinis, Elst, the Netherlands)

and streamed into the data acquisition system. Isometric extensor torque will be

evaluated using an isokinetic dynamometer (CSMi Humac Norm; Stoughton, MA). Beat

by beat blood pressure will be collected with an automated continuous blood pressure

monitoring system (Nexfin, Edwards LifeSciences, Irvine, CA). Functional outcomes

will include the average speed during a 6-minute walk test, the TUG test, and a gait

analysis performed by a licenses physical therapist.

 83

Potential risks and protection against risks

Individuals with iSCIs have an elevated risk of DVT formation due to impaired

mobility of the lower limbs. There is a potential for this risk to be exaggerated during

BFR exercise. However, it is important to note that DVT risk is significantly reduced

from the acute to the chronic stages of a spinal cord injury (Hagen et al., 2012; Teasell et

al., 2009) and there have been minimal ( 0.055% from a sample of 12,600) reported

incidents of DVT in the general population (Clark et al., 2011; Nakajima et al., 2006;

Nakajima et al., 2007). Furthermore, this risk is the one of the foundations for the second

specific aim of this study (understanding the safety of regular BFR training). Therefore,

subjects will have 5.0 mL blood drawn at the end of each week which will be analyzed

for D-dimer, and a bilateral leg ultrasound DVT scan will be performed if any readings

are returned above 500 ng/mL. In addition to DVT risk, there is also risk that the

occlusion pressure or associated pain response to BFR could elicit acute AD. As stated

earlier, previous research from our labs found that 3 sets of unilateral BFR did not elicit

any signs or symptoms of acute AD. However, we intend to monitor beat-by-beat blood

pressure and heart rate during each exercise session. In the event that any subjects elicit

signs and symptoms of acute AD (specifically, significant elevations in blood pressure

with simultaneous bradycardia), the pressure will be removed and patients will be

supported in an upright position in accordance with the Consortium for Spinal Cord

Injury Medicine's clinical practice guidelines for the acute management of autonomic

dysreflexia (Linsenmeyer, 2001). Subjects will also be asked to use the restroom prior to

starting exercise in order to reduce the influence of bladder distension or fecal impaction
 84

on development of acute AD. Similar to DVT risk, the risk of acute AD is also an

outcome measure for the second aim of this study.

Another potential risks associated with this study include possible discomfort and

pain that is normally associated with BFR exercise. However, it is important to note that

the occlusion pressures often used in the general population far exceed the pressures that

we have, and intend to, use in this population. In addition, a recent pilot study from our

lab found that pain was not significantly different between BFR and non-BFR exercise in

our sample of individuals with iSCIs, with BFR set to 125% of venous occlusion

pressure. Therefore, we expect this risk to be minimal. Finally, any exercise, whether

performed under BFR or not, presents the risk of exercise-related injury (i.e. falls, muscle

strains, etc.). In order to minimize this risk, subjects in this study will be exercising in a

very controlled environment under the close supervision of the investigators; who are

experienced exercise physiologists. Specifically, any walking exercise will be performed

with a body-weight support harness for fall prevention, and subjects will receive regular

feedback regarding proper exercise form in order to avoid any musculoskeletal injuries.

Finally, each subject will be given a coded identification number, and all personally

identifiable information will be kept in a locked filing cabinet in a locked office, to which

only the principle investigator will have access.

Potential benefits

Subjects who participate in the experimental conditions in this investigation may

experience improvements in muscular size, strength, and overall function following BFR

exercise. These improvements have the potential for improving overall quality of life and
 85

maintaining independence. Subjects who participate in the control conditions will

continue with their existing physical therapy programs, and may not experience these

added benefits should they exist. However, if this investigation does provide reasonable

evidence that BFR exercise can significantly augment muscular adaptations to low-

intensity exercise in individuals with iSCIs, individuals participating in the control

condition will be transitioned to BFR exercise for an additional 6 weeks. This will assure

that all subjects who participate in this study will have the opportunity to garner the

potential benefits of BFR exercise.

The information that can be gained from this study could be very valuable for

future physical rehabilitation programs. If this investigation were to provide sufficient

evidence that BFR exercise can significantly increase muscular strength, size, and overall

function compared to traditional physical therapy programs, it would support a longer

line of research into the long-term benefits of BFR training in individuals with spinal

cord injuries. Ultimately, the introduction of BFR exercise into regular physical therapy

programs may enhance therapeutic outcomes for individuals suffering from iSCIs.

Safety monitoring

As part of this investigation, our research group will hold weekly research

meeting. As part of this meeting, we will review any adverse events and assure that any

unforeseen safety concerns are appropriately addressed. Should any adverse events

occur, the Kent State University Institutional Review Board will be notified, and the

principle investigator will follow up with the subject. All research activities will then be

suspended by the principle investigator and an emergency research team meeting will be
 86

scheduled. At this meeting, the research team will discuss the event and put in place

additional preventative measures before resuming research activity.

Facilities and Resources

The research supported by this award will be conducted in the Vascular Function

Laboratory housed in the Kent State University Exercise Physiology Program, and in

collaboration with the Louis Stokes' Cleveland VA Medical Center. This Vascular

Function Laboratory houses the majority of the equipment necessary to conduct this

investigation. This includes a Doppler Ultrasound (GE Logic 7, GE Healthcare,

Milwaukee, WI), Near Infrared Spectroscopy (NIRS) unit (Oxymon MKIII, Artinis, Elst,

Netherlands), beat-by-beat blood pressure monitoring (Nexfin, Edwards LifeSciences,

Irvine, CA), pneumatic blood pressure cuffs (DE Hokanson, Bellevue, WA), and a multi-

channel data acquisition system (ADInstruments, Colorado Springs, CO). The Vascular

Function Lab also provides ample space for multiple investigators and wheelchair users

to comfortably maneuver around the lab. Kent State University also houses additional

resources that will be at the investigators’ disposal. These include access to journal

databases, statistical and writing software packages, and a graduate student body who are

actively involved in research. Lastly, all MRI scans will be performed at the Louis

Stokes' Cleveland VA Medical Center. This medical center is one of the largest Medical

Center in the VA network, and has been featured in National Geographic and by the

Discovery Channel for its research on prosthetic hand interfaces for veterans.

Outside of the Kent State community, there are multiple healthcare systems from

which to recruit subjects. These include the Summa Health System, the Cleveland Clinic,
 87

the Louis Stokes' Cleveland VA System, and multiple outpatient physical therapy

programs. Currently, our research team holds relationships with the Louis Stokes'

Cleveland VA Medical Center, the Buckeye Wellness fitness center for individuals with

spinal cord injuries, and other local facilities. Ultimately, the Vascular Function Lab at

Kent State University and the surrounding area offer the resources necessary to complete

the proposed investigation.

Major equipment

This section will describe each piece of major equipment that will be used in this

investigation, including where each is located.

Doppler ultrasound. A GE Logic 7 (GE Healthcare, Milwaukee, WI) Doppler

ultrasound (5 MHz and 14 MHz, respectively) is currently housed in the Vascular

Function Lab at Kent State University. This Doppler ultrasound can be used to image

vessels, muscle tissue, record blood flow velocities, measure vessel and muscle

diameters, and can even be used for analysis of muscle cross sectional area should it be

necessary.

Near infrared spectroscopy. An Oxymon MKIII (Artinis, Elst, the Netherlands)

NIRS system is currently housed in the Vascular Function Lab at Kent State University.

This system includes 2 light transmitters and a single receiver, and gives 2 separate

readings of oxyhemoglobin saturation, deoxyhemoglobin saturation, total hemoglobin,

and tissue oxygenation index. The 2 separate readings (one from each transmitter) are
 88

averaged and reported as a single change score. This system is often used in reporting

tissue oxygenation and perfusion.

Beat-by-beat blood pressure monitoring. A continuous blood pressure

monitoring system (Nexfin, Edwards LifeSciences Corp, Irvine, CA) is currently housed

in the Vascular Function Lab at Kent State University. This system includes a wrist and

finger-cuff unit that reports beat-by-beat systolic, diastolic, and mean arterial pressures;

as well as heart rate.

Isokinetic dynamometer. An isokinetic dynamometer (Humac Norm, CSMi,

Stoughton, MA) is currently housed in the Applied Physiology Laboratories at Kent State

University. This system allows for controlled isometric or isokinetic movement, and can

be used to isolate any joint in the body. This system will also record torque production,

and can specialized protocols can be programmed into its computer to assure consistency

between trials.

Pneumatic occlusion cuffs. A pneumatic pressure cuff system (DE Hokanson

Inc, Bellevue, WA) is currently housed in the Vascular Function Lab at Kent State

University. This system uses a finely controlled air compressor and gauge to

automatically inflate one or two cuffs to a desired pressure, and maintain that pressure

through movement patterns. Included with this system is a range of cuff sizes and

lengths.

Multi-channel data acquisition system. A multi-channel data acquisition

system (ADInstruments Inc., Colorado Springs, CO) is currently housed in the Vascular

Function Lab at Kent State University. This system includes dual-channel EMG, 3-lead
 89

ECG, respiratory belt, strain gauge, and handgrip dynamometer recording. In addition to

that, the NIRS system, the beat-by-beat blood pressure monitoring system, and Doppler

ultrasound audio signal can all be streamed into this system and recorded in real-time.

Finally, this system allows for offline data analysis, and conversion to file formats

compatible with most statistical analysis systems.

Fitness room and equipment. A full-size fitness facility with seven guided

weight-stack machines, a power-rack that supports bench press and squat, free weights,

and resistance bands are currently housed in the Applied Physiology Laboratories at Kent

State University. This facility is accessible by a ground floor for wheelchair users, and is

designated for research purposes.

Other Research Support

The principle investigator does not currently have any other research support from

the Neilsen foundation, or any other grant funding organizations.

 CHAPTER VI

SUMMARY

To our knowledge, no other studies have reported on the safety or feasibility of

performing large muscle mass exercise in individuals living with incomplete spinal cord

injuries. The purpose of this study was to determine if blood flow restricted knee

extension exercise can be performed in individuals with incomplete spinal cord injuries,

if it increases the risk of acute autonomic dysreflexia or deep vein thrombosis formation,

and to explore how muscular activation, oxygenation, and perfusion respond to BFR

exercise.

We reported no incidents of acute autonomic dysreflexia or deep vein thrombosis

in our study sample following unilateral knee extension exercise with blood flow

restriction. We also reported no significant decrease in work performed, pain, or

perceived difficulty between blood flow restricted and unrestricted exercise. In contrast

to the previously reported progressive increase in muscle activation during blood flow

restriction exercise compared to unrestricted exercise in the healthy population, we did

not observe any significant difference in iEMG between knee extension exercise with and

without blood flow restriction. Finally, we observed an increase in tissue oxyhemoglobin

saturation and perfusion with a simultaneous decrease in tissue oxygenation index during

blood flow restriction. Tissue perfusion also undulated during exercise sets, with

Oxyhemoglobin, deoxyhemoglobin, and total hemoglobin saturations decreasing during

muscle contractions and increasing during each subsequent rest period; indicating that the

muscle pump was sufficient in overcoming occlusion pressure during exercise.

90
 91

Ultimately, the data collected from this study suggests that single leg exercise with

moderate (125% of venous occlusion pressure) blood flow restriction does not present a

significant risk of acute autonomic dysreflexia or DVT formation. This provides support

for future research to explore potential training effects of blood flow restricted exercise in

individuals with iSCIs. Accordingly, we are proposing a study that would examine the

effects of six weeks of blood flow restriction exercise on muscle strength, size, and

overall function in individuals with iSCIs. In this same investigation, we also intend to

determine the effects of acute blood flow restriction exercise on muscle function and

fatigue characteristics in individuals with high-level and low-level iSCIs.

 92

Figure 1 An outline of the study design, including both experimental protocols.

 93

Figure 2 An outline of the BFR walking protocol.

 94

Figure 3 Mean arterial pressure (MAP) compared across each experimental trial. *

indicates a significant difference from baseline (p<0.05).

 95

Figure 4 Mean arterial pressure (MAP) compared across each acute exercise bout. *

Indicates a significant difference from baseline (p<0.05).

 96

Figure 5 Systolic blood pressure (SBP) compared across each experimental trial. *

indicates a significant difference from baseline (p<0.05).

 97

Figure 6 Systolic blood pressure (SBP) compared across each acute exercise bout. *

indicates a significant difference from baseline (p<0.05).

 98

Figure 7 Oxyhemoglobin (Oxy) compared across each experimental trial and between

conditions. * indicates a significant difference from baseline, and # indicates a significant

difference between conditions (p<0.05).

 99

Figure 8 Oxyhemoglobin (Oxy) compared across each acute exercise bout and between

conditions. * indicates a significant difference from baseline, ! indicates a significant

difference from the preceding time point, and # indicates a significant difference between

conditions (p<0.05).
 100

Figure 9 Deoxyhemoglobin (Doxy) compared across each experimental trial and between

conditions. * indicates a significant difference from baseline, and # indicates a significant

difference between conditions (p<0.05).

 101

Figure 10 Deoxyhemoglobin (Doxy) compared across each acute exercise bout and

between conditions. * indicates a significant difference from baseline, ! indicates a

significant difference from the preceding time point, and # indicates a significant

difference between conditions (p<0.05).

 102

Figure 11 Total hemoglobin compared across each experimental trial and between

conditions. * indicates a significant difference from baseline, and # indicates a significant

difference between conditions (p<0.05).

 103

Figure 12 Total hemoglobin compared across each acute exercise bout and between

conditions. * indicates a significant difference from baseline, ! indicates a significant

difference from the preceding time point, and # indicates a significant difference between

groups (p<0.05).
 104

Figure 13 Tissue oxygenation index (TOI) compared across each experimental trial and

between conditions. * indicates a significant difference from baseline, and # indicates a

significant difference between groups (p<0.05).

 105

Figure 14 Tissue oxygenation index (TOI) compared across each acute exercise bout and

between conditions. * indicates a significant difference from baseline, ! indicates a

significant difference from the preceding time point, and # indicates a significant

difference between conditions (p<0.05).

 106

Figure 15 A timeline of protocol 1. MVC- Maximal Voluntary Contraction, CMEP-

Central motor evoked potential, CVA- central voluntary activation, PMEP- peripheral

motor evoked potential, PVA- peripheral voluntary activation, BFR- blood flow

restriction.
 107

Figure 16 A timeline of protocol 2. iSCI- incomplete spinal cord injury, BFR- blood

flow restriction, MRI- magnetic resonance imaging of the quadriceps and hamstrings

cross sectional area, Performance measures- Timed-up-and-go, six-minute-walk-test, gait

analysis.
 108

Table 1 Subject Characteristics.

APPENDICES
 APPENDIX A:

AMERICAN SPINAL INJURY ASSOCIATION IMPAIRMENT SCALE

 APPENDIX A:

AMERICAN SPINAL INJURY ASSOCIATION IMPAIRMENT SCALE

111
112
 APPENDIX B:

NEUROPATHIC PAIN SCALE

 APPENDIX B:

NEUROPATHIC PAIN SCALE

114
115
 APPENDIX C:

MODIFIED ASHWORTH SCALE

 APPENDIX C:

MODIFIED ASHWORTH SCALE

117
 APPENDIX D:

CRAIG H. NEILSEN FOUNDATION SPINAL CORD INJURY RESEARCH ON

THE TRANSLATIONAL SPECTRUM PILOT RESEARCH GRANT TEMPLATE

 APPENDIX D:

CRAIG H. NEILSEN FOUNDATION SPINAL CORD INJURY RESEARCH ON THE

TRANSLATIONAL SPECTRUM PILOT RESEARCH GRANT TEMPLATE

119
120
 APPENDIX E:

MANUSCRIPT ABSTRACT
 APPENDIX E:

MANUSCRIPT ABSTRACT

Abstract

Background

Individuals with incomplete spinal cord injuries (iSCIs) suffer from significant

muscle weakness and functional limitations that impact daily life. Improvements in

muscle strength with standard physical therapy is limited. Therefore, an exercise

modality and maximizes adaptations to low intensity exercise would be ideal in this

population.

Purpose

The purpose of this study was to determine the safety and feasibility of blood flow

restriction (BFR) exercise in a sample of individuals with iSCIs, and to compare

physiological responses between BFR and non-BFR exercises.

Methods

Ten subjects with varying levels of iSCI completed a trial of unilateral BFR knee

extension (3 sets x 10 reps) at a self-selected intensity with, and without, BFR (125% of

venous occlusion pressure) in a counterbalanced order. Pain, perceived difficulty, muscle

activation (iEMG), hemodynamics, and tissue oxygenation characteristics were compared

between conditions. A sub-set (n=3) also completed two walking trials, one with BFR

and one without. Each subject was screened for a DVT at the start of the protocol, and

returned for a quantitative D-dimer assessment within a week following the protocol.

Results

122
 123

All subjects were able to complete each BFR trial without any adverse events

(including acute autonomic dysreflexia or DVT formation). No differences were

observed for pain, perceived effort, muscular activation, and lactate between BFR and

control exercise. Mean arterial pressure and systolic pressure both increased with

exercise (18.8% and 17.6% in BFR, and 19.4% and 19.6% in control, respectively;

p<0.05), however, were not different between conditions. Local tissue perfusion and

oxyhemoglobin saturation both increased during BFR exercise (+12.3±96.7 and

+105.4±76.7, respectively); while tissue oxygenation decreased (6.5±3.0%; p<0.05 for all

comparisons).

Conclusion

In conclusion, results from this study suggest that controlled BFR exercise can be

safely performed by individuals suffering from iSCIs without added cardiovascular strain

or heightened pain.
 APPENDIX F:

CRAIG H. NEILSEN FOUNDATION PILOT RESEARCH GRANT SUBMISSION

BUDGET
 APPENDIX F:

NEILSEN FOUNDATION PILOT RESEARCH GRANT SUBMISSION BUDGET

125
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