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Bioengineering Synovial Fluid with Theoretical and Experimental Models of the Synovial Joint UNIVERSITY OF CALIFORNIA, SAN DIEGO BIOENGINEERING SYNOVIAL FLUID WITH THEORETICAL AND EXPERIMENTAL MODELS OF THE SYNOVIAL JOINT A dissertation submitted in partial satisfaction of the requirements for the degree Doctor of Philosophy in Bioengineering by Megan E. Blewis Committee in charge: Professor Robert L. Sah, Chair Adjunct Professor William D. Bugbee Professor Gary S. Firestein Professor Jeff M. Hasty Professor Marcos Intaglietta 2008 Copyright Megan E. Blewis, 2008 All rights reserved. The dissertation of Megan E. Blewis is approved, and it is acceptable in quality and form for publication on microfilm and in digital formats: Chair University of California, San Diego 2008 iii TABLE OF CONTENTS Signature Page ............................................................................................................iii Table of Contents........................................................................................................iv List of Figures and Tables .........................................................................................ix Acknowledgments.......................................................................................................xi Vita……….................................................................................................................xvi Abstract of the Dissertation...................................................................................xviii Chapter 1: Introduction..............................................................................................1 1.1 General Introduction to the Dissertation ...............................................1 1.2 Synovial Joint Components and Structure ............................................6 1.3 Synovial Fluid: Complex Milieu of Lubricants & Chemical Factors ...9 1.4 Articular Cartilage: Load Bearing Material & Source of Lubricants..10 1.5 Synovium: Semi-Permeable Membrane & Source of Lubricants .......11 1.6 Synovial Fluid Lubricant Homeostasis, Dynamics, and Imbalance....12 iv 1.7 Vision & Application of Synovial Joint Models.................................18 1.8 References ...........................................................................................22 Chapter 2: A Model of Synovial Fluid Lubricant Composition in Normal and Injured Joints...........................................................................................32 2.1 Abstract................................................................................................32 2.2 Introduction .........................................................................................34 2.3 Model...................................................................................................38 2.4 Results .................................................................................................52 2.5 Discussion............................................................................................59 2.6 Acknowledgments ...............................................................................64 2.7 References ...........................................................................................65 Chapter 3: Interactive Cytokine Regulation of Synoviocyte Secretion of Synovial Fluid Lubricants, Hyaluronan and Proteoglycan 4..............75 3.1 Abstract................................................................................................75 3.2 Introduction .........................................................................................77 3.3 Materials and Methods ........................................................................81 3.4 Results .................................................................................................85 3.5 Discussion............................................................................................93 v 3.6 Acknowledgments ...............................................................................98 3.7 References ...........................................................................................99 Chapter 4: Semi-Permeable Membrane Retention of Synovial Fluid Lubricants, Hyaluronan and Proteoglycan 4, for a Biomimetic Bioreactor...............................................................................................105 4.1 Abstract..............................................................................................105 4.2 Introduction .......................................................................................107 4.3 Materials and Methods ......................................................................110 4.4 Results ...............................................................................................118 4.5 Discussion..........................................................................................130 4.6 Acknowledgments .............................................................................135 4.7 References .........................................................................................136 Chapter 5: Biomimetic Bioengineering of Synovial Fluid: a Bioreactor for Generating Functional Lubricant Solutions .......................................143 5.1 Abstract..............................................................................................143 5.2 Introduction .......................................................................................146 5.3 Bioengineering Analysis & Design of Bioreactors ...........................149 5.4 Materials and Methods ......................................................................152 vi 5.5 Results ...............................................................................................160 5.6 Discussion..........................................................................................172 5.7 Acknowledgments .............................................................................177 5.8 References .........................................................................................179 Chapter 6: Conclusions...........................................................................................185 6.1 Summary of Findings ........................................................................185 6.2 Discussion..........................................................................................188 6.3 Future Work.......................................................................................192 6.4 References .........................................................................................195 Appendix A: Microenvironment Regulation of PRG4 Phenotype of Chondrocytes .........................................................................................198 A.1 Abstract.............................................................................................198 A.2 Introduction ......................................................................................200 A.3 Materials and Methods .....................................................................202 A.4 Results ..............................................................................................206 A.5 Discussion.........................................................................................215 A.6 Acknowledgments ............................................................................219 vii A.7 References ........................................................................................220 viii LIST OF FIGURES AND TABLES Figure 1.1: Overall aims of dissertation ........................................................................3 Figure 2.1: In vivo synovial joint components and compartmental model .................37 Figure 2.2: Schematic of model describing SF lubricant dynamics............................39 Figure 2.3: General curves of predicted transient SF lubricant concentration............55 Figure 2.4: Transient predictions of lubricant composition after lavage ....................57 Figure 2.5: Transient predictions of lubricant composition after HA injection ..........58 Figure 3.1: Key components of synovial joints, in vivo and modeled........................80 Figure 3.2: Effects of individual cytokines on HA and PRG4 secretion rates............86 Figure 3.3: Effects of combinations of cytokines on HA and PRG4 secretion rates ..88 Figure 3.4: Effects of selected cytokine combinations on HA structure.....................90 Figure 3.5: Effects of all analyzed cytokine combinations on HA structure ..............91 Figure 3.6: Effects of combinations of cytokines on PRG4 structure.........................92 Figure 4.1: Schematic of bioreactor model for assessing SF lubricant retention......111 Figure 4.2: Synoviocyte proliferation and lubricant secretion on ePTFE.................121 Figure 4.3: Total HA and PRG4 loss with transport across membranes...................122 Figure 4.4: HA loss, as a function of MW, with transport across membranes..........123 Figure 4.5: Membrane permeability to total HA and PRG4 .....................................126 Figure 4.6: Memebrane permeability to HA as a function of MW ...........................127 Figure 4.7: Lubricant mass in BF vs. time fit to exponential curves ........................129 Figure 5.1: Schematic of biomimetic bioreactor model of synovial joint.................148 Figure 5.2: HA characterization in SF, BF, and NF..................................................163 ix Figure 5.3: PRG4 characterization in SF, BF, and NF.............................................164 Figure 5.4: Friction-reducing
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