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I SHORT TERM ELECTRICAL STIMULATION for ISOGRAFT PERIPHERAL NERVE REPAIR and FUNCTIONAL RECOVERY a Thesis Presented to the Gradu

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I SHORT TERM ELECTRICAL STIMULATION for ISOGRAFT PERIPHERAL NERVE REPAIR and FUNCTIONAL RECOVERY a Thesis Presented to the Gradu SHORT TERM ELECTRICAL STIMULATION FOR ISOGRAFT PERIPHERAL NERVE REPAIR AND FUNCTIONAL RECOVERY A Thesis Presented to The Graduate Faculty of The University of Akron In Partial Fulfillment Of the Requirements for the Degree Master of Science in Engineering, Biomedical Concentration Galina Y. Pylypiv May, 2018 i SHORT TERM ELECTRICAL STIMULATION FOR ISOGRAFT PERIPHERAL NERVE REPAIR AND FUNCTIONAL RECOVERY Galina Y. Pylypiv Thesis Approved: Accepted: Advisor Dean of the College of Engineering Dr. Rebecca Kuntz Willits Dr. Donald Visco Committee Member Executive Dean of the Graduate School Dr. Matthew Becker Dr. Chand Midha Committee Member Date Dr. Ge Zhang Biomedical Engineering Department Chair Dr. Brian Davis ii ABSTRACT Electrical stimulation (ES) has previously demonstrated promising effects on peripheral nerve repair through enhanced neurite growth in vitro and shortened recovery time in vivo. In this study, we aimed to evaluate the effect of intraoperative short term ES on a clinically relevant isograft-repair model of a rodent peripheral nerve. In our model, an isograft was used to repair a 13 mm sciatic nerve gap-defect in adult male rats. Intraoperative ES was applied for 10 min at 24 V/m-DC to the experimental group and no stimulation was applied to the control group. We evaluated biweekly functional recovery over 12 weeks for motor function, using the sciatic functional index and external postural thrust. Sensory function was evaluated using a thermal stimulus. Motor nerves are more heavily myelinated and regenerate more quickly, while sensory nerves are less myelinated and have a slower recovery time. Structural repair outcomes were evaluated through histological examination of the sciatic nerves and gastrocnemius muscles at 6 and 12-week time points. The ES group had a significantly better motor recovery than the control group in weeks 4 and 6 after surgery. In addition, the ES group had 7% fewer paw contractures than the control group. Paw contractures form when the flexor muscles are innervated more quickly than extensor muscles, leaving the paw in a chronically curled over position that is representative of a clinical challenge in human nerve recovery. Furthermore, sensory functional testing and histology evaluation confirmed that ES was safe to use, as the ES- treated group had comparable recovery outcomes to the no-ES control group. Our previous iii study showed that 10 minutes of ES was effective in promoting functional recovery through a collagen scaffold bridging a 10-mm nerve defect. Here, we extend our findings by showing that ES can speed up motor recovery in an isograft-repair model, while slowing contracture formation. Demonstrating the benefits of applying short term intraoperative ES in a clinically relevant nerve injury model, creates an early point of translation to improve the current standard of care for peripheral nerve injuries. Future work is necessary to evaluate the sensory functional recovery response to ES and evaluate the mechanisms of ES that may lead to a reduction in contracture formation. iv DEDICATION For the other Galina Pylypiv, my mother, from whom I inherited many qualities. And also for Emmanuel Koh, my fiancé, for his encouragement and shared love for rats. v ACKNOWLEDGEMENTS First, a huge thank you to Dr. Rebecca Willits for inspiring me to continuously grow as a researcher. You have inspired my curiosity, pushed me to pose quality questions and seek answers for my questions. Furthermore, you have demonstrated a good example for me in maintaining a good work-life balance. I will forever cherish our conversations and the advice I have received from you throughout my career. Secondly, I would like to thank the Akron General Medical Center – Dr. William Lanzinger, Dr. Robert Tysklind, and Dr. Carol Fouad for their collaboration and meaningful discussions. Thank you, Carol, for sharing your clinical practice as a surgeon and for the efficient teamwork throughout this project. Next, I would like to thank my committee members for their advice and constructive feedback. Additionally, I would like to thank Dr. James Keszenheimer and Stephen Paterson for their help in validating the performance of the electrical stimulation device used in this thesis work. Thank you to the University of Akron Research Vivarium management: Kelly Stevanov, Michelle Evancho-Chapman, and Beth Kanaga. Thank you to Dr. Walter Horne, the attending veterinarian, and to the undergraduate vivarium staff, Ali Daniliuk and Sofie Cressman. Thank you to the graduate and undergraduate students I have worked alongside with. I had the privilege to work with past and present members of Willits Materials for Tissue Engineering Laboratory: Dr. Jessica Stukel, Diana Liz Philip, Elham Malekzadeh, vi Nikhil Prasad, Carlisle DeJulius, Kaitlyn Mangus, Jacquie Carpenter, Wafaa Nasir, Abrar Alniemi, Alena Casella, McKay Cavanaugh, Sean Sulivan, Olivia Detzel, Bailei Hoyng, and Jilian Savage. A special thank you to Jessica for always being encouraging. Also, a special thank you to Diana Liz Philip for becoming like a sister to me, for allowing me to bounce ideas, and for keeping me updated with the latest memes. Lastly, I would like to acknowledge my long list of immediate family members, who make my life so rich and colorful. Thank you to my wonderful fiancé Emmanuel Koh for being my strongest support! Thank you to my parents Yeugen and Galina, and to my siblings along with their significant others: Illya and Nadiya, Yeugen and Sveta, Andrey in heaven, Jerry and Alana, Nick and Katie, Oksana and Tima, and Aleks. Also, my nieces and nephews: Evaleena, Anita, Jaydon, Benjamin, Daria, Josiah, Solomon, and baby Moses in the belly. And finally, a special shout out to my mom, who remembered every deadline and called me – first, to make sure I was well-fed, and second, to let me know she was always cheering me on! vii LIST OF TABLES Table Page 1. Selection of Circuit Components .............................................................................. 19 2. Voltage Variability During Surgery… ....................................................................... 21 3. Sensory NR Data….................................................................................................... 60 viii LIST OF FIGURES Figure Page 1. Structure of Nerve ........................................................................................................ 2 2. Setup of Electrical Stimulation Device ....................................................................... 17 3. Application of Electrical Stimulation During Surgery… ........................................... 28 4. Walking Track Testing Setup… ................................................................................ 30 5. Paw Measurements for SFI… ..................................................................................... 31 6. Measurements of Contractured Paws ......................................................................... 32 7. Extensor Postural Thrust (EPT) Testing Setup… ....................................................... 33 8. Sensory Testing Setup ................................................................................................ 35 9. Glass Temperature for Sensory Testing… ................................................................. 36 10. Tissue Cutting Schematic ......................................................................................... 42 11. Glass Temperature for Sensory Testing… ............................................................... 51 12. EPT Test Evaluation… ............................................................................................ 54 13. Paw Contractures ..................................................................................................... 56 14. Walking Track Evaluation… ................................................................................... 58 15. Sensory Testing Evaluation… .................................................................................. 61 16. Muscle Evaluation…................................................................................................. 62 17. Qualitative Nerve Histology Evaluation… ............................................................... 63 18. Quantitative Nerve Histology Evaluation… ............................................................. 65 ix TABLE OF CONTENTS Page LIST OF TABLES .................................................................................................... vii LIST OF FIGURES ................................................................................................... ix CHAPTER Page I. INTRODUCTION .................................................................................... 1 1.1 Significance .................................................................................................... 1 1.2 Anatomy and Function of Peripheral Nerves .................................................. 2 1.3 Nerve Response to Damage ........................................................................... 3 1.4 Injury Classification… ................................................................................... 4 1.5 Clinical Repair Options and their Limitations ............................................... 5 1.6 Nerve Environment ........................................................................................ 8 1.7 In Vitro Electrical Stimulation… ................................................................... 9 1.8 In Vivo Electrical Stimulation…
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