East Tennessee State University Digital Commons @ East Tennessee State University Electronic Theses and Dissertations Student Works 8-2018 Metabolic Plasticity in the Cellular Stress Response Ying Li East Tennessee State University Follow this and additional works at: https://dc.etsu.edu/etd Part of the Biochemistry Commons Recommended Citation Li, Ying, "Metabolic Plasticity in the Cellular Stress Response" (2018). Electronic Theses and Dissertations. Paper 3467. https://dc.etsu.edu/etd/3467 This Dissertation - Open Access is brought to you for free and open access by the Student Works at Digital Commons @ East Tennessee State University. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of Digital Commons @ East Tennessee State University. For more information, please contact [email protected]. Metabolic Plasticity in the Cellular Stress Response A dissertation presented to the faculty of the Department of Biomedical Sciences East Tennessee State University In partial fulfillment of the requirements for the degree Doctor of Philosophy in Biomedical Sciences by Ying Li August 2018 Gary Wright, Ph.D., Co-Chair Jonathan Peterson, Ph.D., Co-Chair Kenneth Ferslew, Ph.D. Krishna Singh, Ph.D. Stacy Brown, Ph.D. Douglas Thewke, Ph.D. Keywords: hypoxic heart, cardioprotection, HIF-1α, glucose metabolism, metabolite, stable isotope, metabolomics, CTRP3, receptor, lipid metabolism ABSTRACT Metabolic Plasticity in the Cellular Stress Response by Ying Li Changes to the metabolism of the cardiomyocyte are driven by complex signaling pathways in order to adjust to stress. For instance, HIF-1α is classically known to upregulate glycolytic metabolism to compensate for oxygen deficiency. Other important effects upon glucose metabolism, which we investigate here more extensively, were also observed. Hearts derived from mice with the cardiac-restricted expression of a stabilized form of HIF-1α are remarkably ischemia stress-tolerant. Here, stable isotope- resolved metabolomic analyses were utilized to investigate glucose cardiometabolism remodeling by HIF-1α during ischemia. We found that 13C-lactate accumulation was significantly elevated in HIF-1α expressing hearts while paradoxically glycogen was maintained to a remarkable extent during an ischemic time course. These findings suggested an unexpected source of glucose in HIF-1α hearts during global ischemia. Accordingly, the presence of gluconeogenesis in hearts was evaluated. Indeed, gluconeogenic intermediates (i.e. m+3) including glucose-6-phosphate [m+3], fructose- 6-phosphate [m+3], and fructose 1,6-bisphosphate [m+3] were observed at significantly elevated levels in the ischemic HIF-1α heart. Collectively, these data establish the surprising finding that HIF-1α supports active gluconeogenesis in the heart during ischemia. 2 As less is known regarding the effects of CTRP3 we first tested whether CTRP3 overexpression would protect the ischemic heart. Our data indicate that CTRP3 failed to confer ischemic tolerance in heart ex vivo. However, we were able to show that CTRP3 protected the liver from lipid-induced stress and prevented hepatic lipid accumulation. To further investigate the mechanisms of hepatic protective effect mediated by CTRP3, we identified the receptor and established that CTRP3 increases oxygen consumption in response to lipid overloaded. In summary, these data indicate that targeted metabolic rearrangements within cardiomyocyte/hepatocyte holds promise for the alleviation of common pathological conditions. 3 TABLE OF CONTENTS Page ABSTRACT ...................................................................................................................... 2 LIST OF TABLES ............................................................................................................. 7 LIST OF FIGURES ........................................................................................................... 8 Chapter 1. INTRODUCTION ..................................................................................................... 9 HIF-1α ....................................................................................................................... 9 CTRP3 ..................................................................................................................... 12 Overall Hypothesis and Specific Aims .................................................................... 16 2. HIF-1α IN HEART: EVIDENCE OF GLUCONEOGENESIS IN MYOCARDIUM DURING ISCHEMIA .............................................................................................. 18 Abstract ................................................................................................................... 18 Introduction ............................................................................................................. 19 Experimental Procedures ........................................................................................ 21 Results .................................................................................................................... 25 Discussion ............................................................................................................... 32 References .............................................................................................................. 35 3. C1Q/TNF-RELATED PROTEIN 3 (CTRP3) FUNCTION AND REGULATION ...... 38 Abstract ................................................................................................................... 38 Introduction ............................................................................................................. 38 History of CTRP3 .................................................................................................... 40 Initial discovery……………………………..…………………………………………..40 4 Structure……………………………………..…………………………………………..40 Regulation……………………………..…………………….…………………………..47 Tissues expressed………...…………….....…………………………………………..49 Metabolism, Metabolic disease and CTRP3 ........................................................... 51 Overview……………………………..…………………………………………………..51 Human studies………………………….....…………………………………………....55 CTRP3 and Cardiovascular Disease ...................................................................... 57 Inflammation ............................................................................................................ 61 Growth, Reproduction, and Tumorigenesis ............................................................. 64 Summary and Conclusion ....................................................................................... 65 References .............................................................................................................. 67 4. IDENTIFICATION OF PUTATIVE RECEPTORS FOR THE NOVEL ADIPOKINE CTRP3 USING LIGAND-RECEPTOR CAPTURE TECHNOLOGY ....................... 76 Abstract ................................................................................................................... 76 Introduction ............................................................................................................. 77 Experimental Procedures ........................................................................................ 79 Results .................................................................................................................... 85 Discussion ............................................................................................................... 93 Summary and Conclusion ....................................................................................... 93 References .............................................................................................................. 98 5. SUMMARY & FUTURE DIRECTIONS ................................................................ 105 HIF-1α ................................................................................................................... 105 Future Studies with HIF-1α ................................................................................... 107 CTRP3 ................................................................................................................... 107 5 Future Studies with CTRP3 ................................................................................... 108 Conclusion ............................................................................................................ 109 REFERENCES ............................................................................................................. 110 VITA.............................................................................................................................. 115 6 LIST OF TABLES Table Page 3.1 Abbreviations ............................................................................................................ 39 3.2 Complete Summary of the In Vitro Functions of CTRP3 .......................................... 43 3.3 CTRP3 Expression In Vitro ...................................................................................... 50 3.4 Metabolic Effects of CTRP3 In Vivo ......................................................................... 52 3.5 Cross-Sectional Studies Regarding CTRP3 Levels ................................................. 54 3.6 Summary of Cardiovascular Effects of CTRP3 Treatment ....................................... 58 3.7 CTRP3 and Inflammation ........................................................................................