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Regulation of Cardiac Hypertrophy and Metabolism by Regulator of G Protein Signalling 2 (RGS2)

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Regulation of Cardiac Hypertrophy and Metabolism by Regulator of G Protein Signalling 2 (RGS2) Western University Scholarship@Western Electronic Thesis and Dissertation Repository 9-1-2017 10:45 AM Regulation of Cardiac Hypertrophy and Metabolism by Regulator of G Protein Signalling 2 (RGS2) Katherine N. Lee The University of Western Ontario Supervisor Dr. Peter Chidiac The University of Western Ontario Co-Supervisor Dr. Qingping Feng The University of Western Ontario Graduate Program in Pharmacology and Toxicology A thesis submitted in partial fulfillment of the equirr ements for the degree in Doctor of Philosophy © Katherine N. Lee 2017 Follow this and additional works at: https://ir.lib.uwo.ca/etd Recommended Citation Lee, Katherine N., "Regulation of Cardiac Hypertrophy and Metabolism by Regulator of G Protein Signalling 2 (RGS2)" (2017). Electronic Thesis and Dissertation Repository. 4929. https://ir.lib.uwo.ca/etd/4929 This Dissertation/Thesis is brought to you for free and open access by Scholarship@Western. It has been accepted for inclusion in Electronic Thesis and Dissertation Repository by an authorized administrator of Scholarship@Western. For more information, please contact [email protected]. Abstract Pathological left ventricular hypertrophy is a maladaptive cardiomyocyte growth response to various cardiovascular conditions such as hypertension, and is a major risk factor for heart failure and stroke. The majority of drugs used to treat cardiovascular diseases target G protein coupled receptors (GPCRs), which are regulated by regulator of G protein signalling (RGS) proteins. RGS2 is a GTPase activating protein which limits Gq- and Gs-mediated signalling, which are known to play major roles in the development of pathological cardiac hypertrophy. In addition to its G protein effects, we have previously shown that RGS2 can also inhibit protein synthesis and cultured cardiomyocyte growth via a region in its RGS domain, RGS2eb, which inhibits the rate-limiting eIF2B/eIF2 step of protein synthesis initiation. Thus, we hypothesized that the in vivo expression of RGS2eb could limit the development of experimentally induced hypertrophy. Herein, we demonstrate that the in vivo cardiomyocyte specific overexpression of RGS2eb prevents the development of cardiac hypertrophy in a transverse aortic constriction (TAC) model of experimental pressure overload, as well as functional loss and the expression of “fetal” genes associated with hypertrophy and heart failure. Although we further hypothesized that the expression of RGS2eb in Rgs2-/- mice (which are highly sensitive to hypertrophy and heart failure following pressure overload) could compensate for the loss of endogenous RGS2, we were limited in our characterizations by poor survival rates in Rgs2-/- animals following TAC surgery. Cardiovascular dysfunction is also known to be directly influenced by obesity and weight gain, which reflects the importance of understanding the mechanisms behind metabolic dysregulation. We have previously demonstrated that Rgs2-/- mice exhibit a lean phenotype on a normal chow diet, but still gain weight on a high fat diet. We thus sought to further characterize the metabolic role of RGS2 via Comprehensive Lab Animal Monitoring System (CLAMS) metabolic cages and gene expression analysis of skeletal rather than adipose tissue. Here we report that loss of RGS2 leads to greater use of carbohydrates and significant increases in basal metabolism, as well as increases in associated metabolism-related genes. Altogether, these studies provide new insights into the role of RGS2 in cardiovascular health and metabolism. Keywords Regulator of G protein signalling 2 (RGS2), RGS2eb, G protein-coupled receptors (GPCRs), eukaryotic initiation factor 2 (eIF2), eukaryotic initiation factor 2B (eIF2B), cardiac hypertrophy, CLAMS metabolic cages, metabolism, lean phenotype. ii Co-Authorship Statement Chapter 3: Lee KN, Lu X, Nguyen C, Feng QP, Chidiac P (2017), Cardiomyocyte specific overexpression of regulator of G protein signalling 2 inhibits cardiac hypertrophy and improves function in response to pressure overload in mice. J Mol Cell Cardiol 108: 194-202. KNL, XL, QPF, and PC designed the experiments. CN generated mouse line, KNL managed all mice colonies. Animal surgeries were conducted by KNL and XL. KNL performed all other experiments and analyzed all data. KNL and PC wrote the manuscript, with revisions and approval from all authors. Chapter 4: Lee KN, Lu X, Gros R, Xiang FL, Feng XP, Chidiac P (2017), Effects of transverse aortic constriction and drug induced experimental pressure overload hypertrophy on Rgs2-/- mice with cardiomyocyte specific overexpression of RGS2eb. In preparation. KNL and PC designed the experiments. KNL, XL, and FLX performed TAC surgeries; KNL and RG conducted osmotic pump implantations. KNL performed all other experiments and analyzed all data. KNL and PC wrote the manuscript. Chapter 5: Lee KN, Gros R, and Chidiac P (2017), Loss of regulator of G protein signalling 2 (RGS2) alters the effects of acute and chronic diet restriction on metabolic outcomes in C57 Bl/6 mice. Submitted to International Journal of Obesity. Manuscript number: 2017IJO01137. KNL, RG, and PC designed the experiments. KNL and RG conducted CLAMS cage experiments. KNL performed all other experiments and analyzed all data. KNL and PC wrote the manuscript, with revisions and approval from all authors. iii Dedication To my parents and grandparents iv Acknowledgments First, I am incredibly thankful to have had Dr. Peter Chidiac as my doctoral supervisor. The positive research environment that he continuously fosters has given me the opportunity to conduct high quality research, explore my own ideas, and develop as an independent scientist. He has, through his immeasurable patience and sage guidance, instilled within me a core set of technical and intangible skills which I will value for the foreseeable future, and for that I am forever grateful. I would also like to thank my co-supervisor Dr. Qingping Feng for his guidance and in-depth knowledge of cardiovascular physiology, without which I could not have completed much of my doctoral work. In addition, I am grateful for the generous amount of time that Dr. Sharon Lu dedicated to train me in surgical and lab techniques, as well as for her surgery skills, which were critical for the delicate procedures that were required in our animal models. I am further thankful for the mentorship and experimental help from my advisory committee member Dr. Robert Gros, who was integral to the successful development of many of my projects. I also thank Dr. Thomas Drysdale, my advisory committee chair, who facilitated and contributed invaluable research advice. I am fortunate to have completed my training within the collegial environment of the Department of Physiology and Pharmacology, and am thankful for the support and advice I have received from various faculty and staff including, among many others, Dr. Frank Beier, Dr. John DiGuglielmo, Murong Liu, Susan McMillan, Dr. Rithwik Ramachandran, Dr. Peter Stathopulos, Chris Webb, and Dr. Lin Zhao. Many friends and past lab members have also made these years a wonderful experience. From the Feng lab, thank you to Dr. Carmen Leung, Dr. Jessica Blom, Anish Engineer, and Tana Saiyin for your friendship and support both inside and outside of the lab. Thank you also to the past and current members of the Chidiac lab, including Patrick Stockwell, Lylia Nini, and especially Dr. Elva Zhao. Dr. Thomas Velenosi, I value the constant support you have given me. I would also like to thank the Heart and Stroke Foundation of Ontario, the Canadian Institutes of Health Research and its Institute of Circulatory and Respiratory Health, the Ontario v provincial government, and the Schulich School of Medicine and Dentistry for the financial support of this thesis. Finally, I would like to thank my family. My grandparents, aunts, and uncles have always supported me, and I am so very grateful for their love. Ultimately, this thesis could not have been possible without my mother and father. My successes stem directly from their unconditional support, their hard work, and the sacrifices they have made for me. I am beyond fortunate to have them as my parents. vi Table of Contents Abstract ................................................................................................................................ i Co-Authorship Statement................................................................................................... iii Dedication .......................................................................................................................... iv Acknowledgments................................................................................................................v Table of Contents .............................................................................................................. vii List of Tables .....................................................................................................................xv List of Figures .................................................................................................................. xvi List of Appendices ......................................................................................................... xviii Abbreviations ................................................................................................................... xix 1 INTRODUCTION ..........................................................................................................1 Regulator of G
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