POLY(VINYL ALCOHOL) NANOCOMPOSITE HYDROGELS FOR INTERVERTEBRAL DISC PROSTHESES (Spine title: PVA Nanocomposite Hydrogels for IVD Prostheses) (Thesis format: Monograph) by Elaine Y L Wong Biomedical Engineering Graduate Program A thesis submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy The School of Graduate and Postdoctoral Studies The University of Western Ontario London, Ontario, Canada © Elaine Y L Wong 2012 THE UNIVERSITY OF WESTERN ONTARIO School of Graduate and Postdoctoral Studies CERTIFICATE OF EXAMINATION Supervisor Examiners Dr Wankei Wan Dr Brian Amsden Supervisory Committee Dr Don Hewson _ Dr John de Bruyn Dr Jeffrey Hutter Dr John Medley Dr James Johnson Dr Amin Rizkalla The thesis by Elaine Y L Wong entitled: Poly(Vinyl Alcohol) Nanocomposite Hydrogels for Intervertebral Disc Prostheses is accepted in partial fulfillment of the requirements for the degree of Doctor of Philosophy Date _ Chair of the Thesis Examination Board ii ABSTRACT Spinal fusion is currently the gold standard for surgical intervention of intervertebral disc (IVD) diseases leading to neck and back pain failing conservative treatments However, fusion removes motion between the vertebrae and can result in adjacent level degeneration Total disc replacement (TDR) is an emerging treatment alternative that preserves motion, but materials found in clinically available devices bear little resemblance to the properties of the native IVD Poly(vinyl alcohol) (PVA) hydrogels are biocompatible, have mechanical behaviour similar to natural tissues, and properties that can be tuned by varying polymer concentration and physical crosslinking through freeze-thaw cycling Furthermore, their properties can be modified with the addition of nanofillers In the present study, PVA hydrogels and its nanocomposites containing Laponite and bacterial cellulose (BC) were investigated in compression and crossing-paths wear for potential application in cervical TDR While increases in PVA concentration increased stiffness and decreased time-dependent response in neat PVA hydrogels, viscous response increased with nanofiller addition BC addition also increased stiffness of the hydrogels without large changes in water content To measure wear in the hydrogels, a technique using three-dimensional ultrasound imaging was developed Wear volume and depth decreased with decreasing water content, while fatigue wear was eliminated with the addition of nanofillers in crossing-paths wear Finally, a two-component PVA hydrogel demonstrated that compression properties could be tailored by mimicking the natural IVD structure These results indicated that various parameters could be used to optimize the properties of PVA and PVA-nanocomposite hydrogels for applications in cervical TDR Keywords: hydrogel, poly(vinyl alcohol), Laponite, bacterial cellulose, nanocomposites, mechanical properties, strain rate dependence, crossing-paths wear, three-dimensional ultrasound imaging, intervertebral disc iii ACKNOWLEDGEMENTS First of all, I would like to sincerely thank my supervisor, Dr Wankei Wan, for his inspiration in the field of biomaterials and medical devices I am grateful for his all his ideas, helpful advice, guidance, encouragement and support that culminated with the completion of this thesis I am also thankful for the direction my advisory committee has given to my project, especially Dr John Medley for his helpful insight in wear testing and for the loan of the wear tester The following people have contributed their expertise and assistance to the work in this thesis: Dr Jim Lacefield for the use of the ultrasound equipment at Robarts and his knowledge in ultrasound imaging and analysis, the staff at the Biotron for their advice and training on sample preparation and SEM, Clayton Cook from the University Machine Services for help with designing various parts and moulds, Michael Roach from 3M for advice on adhesives and providing us with a sample, Dr Leonardo Millon for instruction and advice on PVA and mechanical testing, Darcy Small and Dr Kenneth Wong for the TEM images, Dr Donna Padavan for help on SEM, Dr Karen Kennedy for assistance in the preparation and testing of the IVD prototype, undergraduate students Rachel Brown and Ghaleb Sater for their work in the laboratory, Andrew Norman, Jordan DeMello and Xinsheng Li for providing the bacterial cellulose, and Dr Jian Liu for performing EDX on the cellulose samples Thank you also to other colleagues in my lab and BME for their friendship through the years I am truly blessed to have enjoyed valuable friendships outside of the lab: friends from the King’s community for keeping me rooted and for the wonderful gifts of music and fellowship; and my relatives and friends, especially Anabela, Amanda, Calvin, Karen, Sarah, Wailan, and my EngSci family, who have touched my life and stuck with me through thick and thin I am humbled by your presence in my life I wish to honour my parents, to whom this thesis is dedicated, for supporting me from the moment I hurriedly entered the world All of this would not have been possible iv without their love, care and sacrifices to provide me with the opportunities they did not have Finally, to my brilliant husband, I would like to acknowledge and thank him for his unwavering patience and for lending his competency in computer graphics There are no words worthy to express my gratitude for his love and unfaltering faith in me v Dedicated to my parents vi TABLE OF CONTENTS CERTIFICATE OF EXAMINATION _ ii ABSTRACT _ iii ACKNOWLEDGEMENTS _ iv TABLE OF CONTENTS _ vii LIST OF TABLES _ xii LIST OF FIGURES xiv LIST OF APPENDICES _ xx LIST OF ABBREVIATIONS xxi LIST OF SYMBOLS _ xxiii CHAPTER INTRODUCTION 1.1 Background and Motivation 1.2 Objectives CHAPTER LITERATURE REVIEW _ 2.1 Prevalence of Neck and Back Pain _ 2.2 Intervertebral Discs _ 2.2.1 Intervertebral Disc Anatomy _ 2.2.2 Degenerative Disc Disease 2.2.3 Intervertebral Disc Mechanics _ 2.2.3.1 Compressive Stress-Strain Behaviour and Strain Rate Dependence 10 2.2.3.2 Stress Relaxation and Creep _ 12 2.2.3.3 Mechanical Properties of IVD Components _ 13 2.2.3.4 Summary 15 2.2.4 Current and Emerging Treatments _ 15 2.3 Cervical Artificial Discs 17 2.3.1 BRYAN Disc _ 19 2.4 PVA Hydrogels _ 21 2.4.1 Physically Crosslinked PVA 21 2.4.2 Characterization of PVA Hydrogel Structure _ 23 vii 2.4.3 Mechanical Properties _ 2.4.3.1 Unconfined Compression _ 2.4.3.2 Stress Relaxation, Creep and Dynamic Mechanical Properties 2.4.4 Effects of aging and salt on PVA hydrogels 2.4.5 Biocompatibility _ 25 25 27 29 31 2.5 Nanofillers and Nanocomposites 2.5.1 Laponite 2.5.2 Bacterial Cellulose _ 2.5.3 PVA Hydrogel-Based Nanocomposites _ 32 33 35 36 2.6 Hydrogel-based Artificial IVD _ 39 2.7 Wear Testing _ 2.7.1 Lubricant _ 2.7.2 Wear and Friction of Hydrogels _ 2.7.3 Characterization of Wear 2.7.4 Crossing-Paths Wear 40 41 42 43 45 2.8 High Frequency 3D Ultrasound Imaging _ 46 2.9 Motivation for Thesis 47 CHAPTER MATERIALS AND METHODS 48 3.1 Materials 48 3.2 Preparation of PVA and PVA-Nanocomposite Hydrogels 3.2.1 Preparation of PVA Solutions _ 3.2.2 Preparation of PVA-Laponite Solutions _ 3.2.3 Preparation of PVA-BC Solutions _ 3.2.4 Pouring Solutions in to Moulds 3.2.5 Freeze-Thaw Cycling _ 3.2.6 Aging in Water and Solutions _ 49 49 50 50 52 52 52 3.3 Structure Studies 3.3.1 Scanning Electron Microscopy 3.3.1.1 Critical Point Drying _ 3.3.1.2 Scanning Electron Microscopy _ 3.3.2 Differential Scanning Calorimetry _ 53 53 53 54 54 3.4 Mechanical Testing 3.4.1 Unconfined Compression 3.4.2 Stress Relaxation _ 3.4.3 Creep 3.4.3.1 Creep Modelling 55 55 56 57 57 viii 3.4.4 Cyclic Compression Testing 59 3.5 Crossing-Path Wear 60 3.5.1 Three-Dimensional Ultrasound for Characterization of Wear 63 3.5.2 Scanning Electron Microscopy 64 3.6 Two-Component Hydrogel IVD Structure 3.6.1 Mould Design _ 3.6.2 Two-Component Hydrogel Disc _ 3.6.3 Compression Testing of Two-Component Hydrogel Disc _ 64 65 65 66 3.7 Statistics _ 66 CHAPTER COMPRESSION PROPERTIES OF PVA AND PVA-NANOCOMPOSITE HYDROGELS 67 4.1 Composition and Structure of Hydrogels _ 4.1.1 Laponite 4.1.2 Bacterial Cellulose _ 4.1.3 Water Content of Solutions and Hydrogels 4.1.4 Porous Structure of Hydrogels 4.1.5 Aging of Hydrogels in Water and PBS 4.1.5.1 Decrease in Mass and Volume _ 4.1.6 Crystallinity of Hydrogels 68 69 69 70 71 73 74 76 4.2 Unconfined Compression of Hydrogels 4.2.1 PVA Concentration _ 4.2.2 Nanofiller Addition _ 4.2.3 Effect of Aging in Water and PBS _ 4.2.4 Strain Rate Dependence _ 77 78 79 81 84 4.3 Stress Relaxation and Creep _ 87 4.3.1 Stress Relaxation _ 87 4.3.2 Creep 89 4.4 Cyclic Compression Testing _ 95 4.5 Discussion _ 95 4.5.1 Structure of PVA and PVA-NC Hydrogels 95 4.5.2 Compression Properties 100 4.5.3 Strain Rate Dependence 105 4.5.4 Stress Relaxation and Creep _ 106 4.5.5 Cyclic Compression Testing _ 109 4.5.6 Comparison to the Natural IVD and Application to IVD Device Design _ 110 4.5.7 Proposed PVA Hydrogel IVD Design _ 115 ix 4.6 Concluding Remarks 119 CHAPTER TRIBOLOGY TESTING OF PVA AND PVA-NANOCOMPOSITE HYDROGELS 120 5.1 Crossing-Paths Wear Testing _ 120 5.2 Volume and Depth of Wear Track _ 121 5.3 Wear Track Surfaces and Substructure 5.3.1 Surface of Unworn Areas _ 5.3.2 Wear Track Surfaces _ 5.3.2.1 PVA Hydrogels 5.3.2.2 PVA-NC Hydrogels 5.3.3 Subsurface Structure _ 123 126 126 128 129 131 5.4 Discussion 133 5.5 Concluding Remarks 147 CHAPTER DESIGN OF A MULTI-COMPONENT PVA HYDROGEL-BASED CERVICAL IVD PROSTHESIS 149 6.1 IVD Prototype Composition 149 6.2 Unconfined Compression 151 6.2.1 Strain Rate Dependence 151 6.3 Stress Relaxation and Creep 155 6.4 Discussion 6.4.1 Prototype Design and Performance 6.4.2 Parameters for Optimization of Properties 6.4.3 Implementation of a Hydrogel-Based IVD Device 157 157 166 169 6.5 Concluding Remarks 171 CHAPTER CONCLUSIONS AND FUTURE WORK 173 7.1 Conclusions _ 173 7.2 Future Work _ 175 BIBLIOGRAPHY _ 178 APPENDIX A PBS PREPARATION 201 APPENDIX B ENERGY-DISPERSIVE X-RAY SPECTROSCOPY OF PHOSPHORYLATEDBACTERIAL CELLULOSE _ 204 APPENDIX C PROCEDURE FOR FITTING OF UNCONFINED COMPRESSION DATA _ 207 x 210 Appendix D SAMPLE HOLDERS FOR CROSSING-PATHS WEAR TESTER 211 212 Appendix E MATLAB PROGRAM FOR EXTRACTION OF ULTRASOUND IMAGES 213 Saving B-Mode Images Planes in a 3D Ultrasound File (.rdb) as a Series of TIFF Image Files Adapted from code written by Z Khan %rdb_to_tiff.m clear filename='filename.rdb'; fid=fopen(filename,'r'); frames=427; pathname = strrep(filename,'.rdb','\'); mkdir(pathname); for pos=1:frames fseek(fid,pos*512*512,'bof'); %find position of each 512x512 frame im = fread(fid,[512,512],'uint8'); im = uint8(im); im = im'; imname=[strrep(filename,'.rdb','') '_' num2str(pos) '.tif']; imwrite(im,[pathname imname],'TIFF') end 214 Appendix F COPYRIGHT PERMISSIONS 215 Figure 2.1b Formal permission not required for reproduction in thesis/dissertation Figure 2.4 216 217 218 Figure 2.5 219 220 221 Figure 2.6 222 CURRICULUM VITAE ELAINE Y L WONG EDUCATION 2012 Doctor of Philosophy (PhD) in Biomedical Engineering The University of Western Ontario, London, Ontario Poly(vinyl alcohol) Nanocomposite Hydrogels for Intervertebral Disc Prostheses Supervisor: Dr Wankei Wan 2003 Bachelor of Applied Science (BASc) in Engineering Science University of Toronto, Toronto, Ontario Biomedical Option, Professional Experience Year Program AWARDS 2005 – 2008 Western Graduate Research Scholarship (WGRS) ($6600 per year) 2003 – 2005 Special University Scholarship RELATED WORK EXPERIENCE 2005 – 2010 Graduate Teaching Assistant in Biomaterials Engineering Biomedical Engineering Graduate Program The University of Western Ontario Supervisors: Dr A Rizkalla and Dr W.K Wan 223 2003 – 2004 Graduate Teaching Assistant in Industrial Organic Chemistry Department of Chemical and Biochemical Engineering The University of Western Ontario Supervisor: Dr Paul Charpentier 2001 – 2002 Research Assistant Corporate Research and Development ShawCor Ltd Supervisor: Dr Dennis Wong CONFERENCE PRESENTATIONS Wong, E.Y.L., Brown, R., Ma, R., Xiong, D.S., Wan, W.K (2009) Poly(vinyl alcohol) hydrogels and its nanocomposites for applications in artificial intervertebral disc and cartilage replacement Poster Presentation 28th Annual Meeting of the Canadian Biomaterials Society, Kingston, Ontario Wong, E.Y.L., Brown, R., Wan, W.K (2009) Poly(vinyl) alcohol nanocomposite hy- drogels for medical applications in compression Poster Presentation 27th Annual Meeting of the Canadian Biomaterials Society, Quebec City, Quebec Wong, E.Y.L., Wan, W.K (2009) Poly(vinyl alcohol) hydrogels for biomedical applica- tions in compression Western Research Forum, London, Ontario Wong E.Y.L., Wan, W.K (2007) Mechanical properties of poly(vinyl alcohol)-Laponite nanocomposite hydrogels Poster Presentation 26th Annual Meeting of the Canadian Biomaterials Society, London, Ontario 224 Wong, E.Y.L., Kennedy, K.L., and Wan, W.K (2006) Mechanical Properties of Opti- cally Transparent Poly(Vinyl Alcohol) by the Freeze-Thaw Method Poster Presentation 25th Annual Meeting of the Canadian Biomaterials Society, Calgary, Alberta ... by Elaine Y L Wong entitled: Poly(Vinyl Alcohol) Nanocomposite Hydrogels for Intervertebral Disc Prostheses is accepted in partial fulfillment of the requirements for the degree of Doctor of Philosophy... _ 2.2 Intervertebral Discs _ 2.2.1 Intervertebral Disc Anatomy _ 2.2.2 Degenerative Disc Disease 2.2.3 Intervertebral Disc Mechanics... optimize the properties of PVA and PVA -nanocomposite hydrogels for applications in cervical TDR Keywords: hydrogel, poly(vinyl alcohol), Laponite, bacterial cellulose, nanocomposites, mechanical properties,