BIOPHYSICAL SIGNIFICANCE OF MITOCHONDRIAL PROPERTIES ON MITOCHONDRIAL FUNCTION: EXPERIMENTAL-COMPUTATIONAL APPROACH by Dong Hoon Song A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy (Mechanical Engineering) in The University of Michigan 2013 Doctoral Committee: Emeritus Professor Ann Marie Sastry, Co-Chair Professor Wei Lu, Co-Chair Associate Professor Christian Lastoskie Assistant Research Scientist Jonghyun Park Professor Martin Philbert © Dong Hoon Song 2013 DEDICATION To my family ii TABLE OF CONTENTS DEDICATION .............................................................................................................. ii LIST OF TABLES ....................................................................................................... vi LIST OF FIGURES .................................................................................................... vii ABSTRACT ................................................................................................................x CHAPTER 1 INTRODUCTION...................................................................................1 1.1. CHALLENGES ............................................................................................1 1.2. PRIOR ART ON MITOCHONDRIAL RESEARCH .....................................3 1.2.1. Role of mitochondria in CNS diseases ............................................3 1.2.2. Mitochondrial membrane potential ..................................................6 1.2.3. Mitochondrial morphology .............................................................9 1.3. SCOPE AND OUTLINE OF THE DISSERTATION .................................. 12 1.4. BIBLIOGRAPHY ....................................................................................... 13 CHAPTER 2 QUANTITATIVE ANALYSIS OF MITOCHONDRIAL PROPERTIES UNDER 1,3-DNB EXPOSURE ................................... 17 2.1. INTRODUCTION ....................................................................................... 17 2.2. METHODS ................................................................................................. 19 2.2.1. Experimental methods................................................................... 19 2.2.2. Image processing and quantification of mitochondrial properties .. 20 2.2.3. Treatment of mitochondrial permeability transition inhibitors ....... 23 2.2.4. Statistical analysis ......................................................................... 25 iii 2.3. RESULTS ................................................................................................... 27 2.3.1. Correlation between mitochondrial morphology and membrane potential in individual mitochondria .............................................. 27 2.3.2. Effects of 1,3-DNB on mitochondrial properties ........................... 31 2.3.3. Effects of mitochondrial permeability transition inhibitors ............ 34 2.4. DISCUSSION ............................................................................................. 36 2.5. CONCLUSIONS ......................................................................................... 38 2.6. BIBLIOGRAPHY ....................................................................................... 40 CHAPTER 3 BIOPHYSICAL SIGNIFICANCE OF THE INNER MITOCHONDRIAL MEMBRANE STRUCTURE ON THE ELECTROCHEMICAL POTENTIAL OF MITOCHONDRIA ........ 43 3.1. INTRODUCTION ....................................................................................... 43 3.2. METHODS ................................................................................................. 46 3.2.1. Structure of the mitochondrion model ........................................... 46 3.2.2. Model descriptions and assumptions ............................................. 51 3.2.3. Post-processing ............................................................................. 56 3.3. RESULTS ................................................................................................... 57 3.3.1. The effect of the crista on electrochemical potential ...................... 57 3.3.2. The effect of the overall shape of a crista ...................................... 59 3.3.3. The effect of the surface area and the surface-to-volume ratio of a crista .......................................................................................... 60 3.3.4. The effect of the crista junction morphology ................................. 61 3.4. DISCUSSION ............................................................................................. 63 3.4.1. Biophysical significance of the crista structure .............................. 63 3.4.2. Membrane-bound proton diffusion coefficient .............................. 64 3.4.3. Model assumptions and limitations ............................................... 67 3.5. CONCLUSIONS ......................................................................................... 69 iv 3.6. BIBLIOGRAPHY ....................................................................................... 72 CHAPTER 4 NUMERICAL STUDY OF INNER MITOCHONDRIAL MEMBRANE DYNAMICS: EFFECTS OF LOCAL pH ON CRISTAE MORPHOLOGIES ............................................................. 76 4.1. INTRODUCTION ....................................................................................... 76 4.2. METHODS ................................................................................................. 79 4.2.1. Local pH and curvature of the lipid bilayer ................................... 80 4.2.2. Area expansion coefficient (αA) ..................................................... 82 4.2.3. Lipid bilayer model ....................................................................... 83 4.2.4. Finite element approximation and energy minimization ................ 87 4.2.5. Initial configurations, pH profiles, and post-processing ................. 88 4.3. RESULTS ................................................................................................... 91 4.3.1. Model verification......................................................................... 91 4.3.2. Formation and dissipation of the cristae-like structure at the GUV scale .................................................................................... 93 4.3.3. The effects of Paff and initial shapes .............................................. 95 4.3.4. Cristae formation at the mitochondrial scale .................................. 98 4.4. DISCUSSION ........................................................................................... 100 4.4.1. Comparison with existing experimental observations .................. 100 4.4.2. Model assumptions and limitations ............................................. 102 4.5. CONCLUSIONS ....................................................................................... 105 4.6. BIBLIOGRAPHY ..................................................................................... 107 CHAPTER 5 CONCLUSIONS AND FUTURE WORK ......................................... 110 v LIST OF TABLES Table 1.1. Prior art on mitochondria and CNS diseases ....................................................5 Table 1.2. Prior art on mitochondrial membrane potential (MMP) ...................................8 Table 1.3. Prior art on mitochondrial morphologies ....................................................... 11 Table 2.1. Correlation between mitochondrial morphology and membrane potential ...... 30 Table 3.1. Parameters for constructing type I geometries ............................................... 50 Table 3.2. Parameters for constructing type II geometries .............................................. 51 Table 3.3. Simulation coefficients and parameters ......................................................... 55 vi LIST OF FIGURES Figure 2.1. Image processing procedures. (a) Image of the experimentally measured point spread function. (b) An example of the image processing result. .......... 20 Figure 2.2. (a) Illustration of the problem in the conventional morphology quantification method. (b) Morphology quantification of a branched mitochondrion achieved by applying the method introduced in this study. .... 22 Figure 2.3. Image processing for the quantification of mitochondrial position. ............... 24 Figure 2.4. Images of the selected mitochondrion showing correlated changes in mitochondrial morphology (MTDR images) and membrane potential (TMRM images). ......................................................................................... 28 Figure 2.5. Simultaneous quantification of mitochondrial properties (mitochondrial membrane potential, aspect ratio, and volume). ............................................ 29 Figure 2.6. Changes in mitochondrial properties after 5 hours of 600 μM 1,3-DNB exposure. ...................................................................................................... 31 Figure 2.7. Response surface of linear regression model for mean aspect ratio. .............. 32 Figure 2.8. Probability of flickering estimated from the probit regression model and experiment. .................................................................................................. 33 Figure 2.9. The effects of the treatments on the mean aspect ratio and the number of mitochondria per cell. (a) The mean aspect ratios of mitochondria from the different treatments. (b) The numbers of mitochondria per cell from the different treatments. ....................................................................................