Restoring diabetes-induced autophagic flux arrest in ischemic/reperfused heart by ADIPOR (adiponectin receptor) activation involves both AMPK-dependent and AMPK-independent signaling.
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2017-01
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Macroautophagy/autophagy is increasingly recognized as an important regulator of myocardial ischemia-reperfusion (MI-R) injury. However, whether and how diabetes may alter autophagy in response to MI-R remains unknown. Deficiency of ADIPOQ, a cardioprotective molecule, markedly increases MI-R injury. However, the role of diabetic hypoadiponectinemia in cardiac autophagy alteration after MI-R is unclear. Utilizing normal control (NC), high-fat-diet-induced diabetes, and Adipoq knockout (adipoq-/-) mice, we demonstrated that autophagosome formation was modestly inhibited and autophagosome clearance was markedly impaired in the diabetic heart subjected to MI-R. adipoq-/- largely reproduced the phenotypic alterations observed in the ischemic-reperfused diabetic heart. Treatment of diabetic and adipoq-/- mice with AdipoRon, a novel ADIPOR (adiponectin receptor) agonist, stimulated autophagosome formation, markedly increased autophagosome clearance, reduced infarct size, and improved cardiac function (P < 0.01 vs vehicle). Mechanistically, AdipoRon caused significant phosphorylation of AMPK-BECN1 (Ser93/Thr119)-class III PtdIns3K (Ser164) and enhanced lysosome protein LAMP2 expression both in vivo and in isolated adult cardiomyocytes. Pharmacological AMPK inhibition or genetic Prkaa2 mutation abolished AdipoRon-induced BECN1 (Ser93/Thr119)-PtdIns3K (Ser164) phosphorylation and AdipoRon-stimulated autophagosome formation. However, AdipoRon-induced LAMP2 expression, AdipoRon-stimulated autophagosome clearance, and AdipoRon-suppressed superoxide generation were not affected by AMPK inhibition. Treatment with MnTMPyP (a superoxide scavenger) increased LAMP2 expression and stimulated autophagosome clearance in simulated ischemic-reperfused cardiomyocytes. However, no additive effect between AdipoRon and MnTMPyP was observed. Collectively, these results demonstrate that hypoadiponectinemia impairs autophagic flux, contributing to enhanced MI-R injury in the diabetic state. ADIPOR activation restores AMPK-mediated autophagosome formation and antioxidant-mediated autophagosome clearance, representing a novel intervention effective against MI-R injury in diabetic conditions.
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Wang, Yajing, Bin Liang, Wayne Bond Lau, Yunhui Du, Rui Guo, Zheyi Yan, Lu Gan, Wenjun Yan, et al. (2017). Restoring diabetes-induced autophagic flux arrest in ischemic/reperfused heart by ADIPOR (adiponectin receptor) activation involves both AMPK-dependent and AMPK-independent signaling. Autophagy, 13(11). pp. 1855–1869. 10.1080/15548627.2017.1358848 Retrieved from https://hdl.handle.net/10161/31639.
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Walter J. Koch
My research interests are centered on the molecular mechanisms involved in the regulation of signaling through cardiovascular adrenergic receptors (ARs) including the specific in vivo interactions between ARs and myocardial G protein-coupled receptor kinases (GRKs). This includes the role of ARs and GRKs in cardiovascular disease. The β-adrenergic receptor kinase (betaARK1) is a prototypic member of the GRK family that targets and phosphorylates agonist-occupied G protein-coupled receptors which causes functional uncoupling and a dampening of signaling, a process termed desensitization. There is a growing body of evidence demonstrating that the actions of GRKs in the heart are extremely important in modulating myocardial adrenergic signaling and hence cardiac function. Of particular interest to my laboratory, are the biochemical and physiological consequences of altering myocardial AR and GRK signaling. To address this, we are utilizing several in vitro and in vivo model systems. Of special interest, are the endogenous alterations that occur in myocardial GRKs and ARs during cardiovascular diseases such as heart failure (HF). We are also interested in the role of adrenergic signaling in the transplanted heart.
Since heart disease accounts for nearly 40% of all deaths annually in this country, it is of importance to learn more of the molecular pathology present in diseased myocardium. For example, in human HF regardless of the cause, a specific constellation of biochemical defects in cardiac tissue has been noted including loss of specific betaAR density and uncoupling of remaining receptors. Moreover, it has been shown that the levels of betaARK1 rise 3-5 fold in human HF, probably contributing to the dysfunction. Interestingly, it has recently been demonstrated that cardiovascular GRK levels (i.e. betaARK1) and activity were also increased in other disorders including myocardial hypertrophy and hypertension. Thus, a primary objective of my laboratory is to investigate the molecular mechanisms involved in myocardial GRK alterations and how these changes relate to adrenergic signaling. This research includes studying and exploring novel protein binding partners of GRKs in the myocardium using a variety of molecular biochemical assays including GRK-affinity columns.
Over the last several years, my laboratory has been investigating the consequences of altering myocardial ARs and GRKs in several in vivo model systems. Original studies were done in transgenic mice where AR and GRK-based transgenes were targeted specifically to the heart. These studies have been done in close collaboration with Dr. Robert Lefkowitz and Dr. Howard Rockman here at Duke University. We have found that altering AR signaling in the hearts of transgenic mice has profound effects on cardiovascular physiology including the findings that overexpression of beta2ARs or a peptide inhibitor of betaARK1 known as the betaARKct, significantly enhances in vivo cardiac contractility. We now have transgenic animals with myocardial-targeted overexpression of several ARs (alpha and beta) or GRKs and are studying the specific in vivo interactions of these molecules in the heart. Moreover, in collaboration with Drs. Lefkowitz and Rockman, we are generating a host of tissue and temporal specific conditional-knockout mice targeting the myocardial betaAR and GRK systems. Another area of transgenic animals being studied exclusively in my laboratory is altered betaAR and GRK activity specifically targeted to arterial smooth muscle in order to determine the role of AR signaling and desensitization in hypertension and other vascular diseases. These mice have recently been generated in our laboratory and are the target of current investigations.
Over the last year, we have been able to specifically study the role of betaAR desensitization and the role of betaARK1 in HF using genetically engineered mouse models of cardiomyopathy and HF. These studies have lead to the findings that inhibition of betaARK1 through myocardial-targeted betaARKct expression has rescued three separate genetic mouse models of cardiomyopathy, preventing the development of HF. The molecular study of these genetic models through DNA array technology is a logical step in the evolution of these studies and this is a new area of focus in the laboratory. Thus, we are analyzing the hearts of mice with altered betaAR and/or betaARK1 signaling by gene (DNA) chips to determine other genes that have been induced or silenced by our transgenes or gene knockouts. We are excited about using this technology in the laboratory taking advantage of our novel mouse models. Specifically, comparing differential gene expression in a failing mouse heart and comparing the genetic pattern in a heart that has been "rescued" by the betaARKct could lead to the elucidation of specific genes involved in the pathogenesis of HF. Importantly, this may also lead to novel therapeutic approaches to treating this disease as well as other cardiovascular disorders.
Our findings in mice that show that we can genetically enhance the functional contractility of the heart form the basis of another major focus of my research program which is the investigation of enhancing the in vivo function of the compromised heart via acute genetic manipulation using gene therapy. My laboratory heads the Cardiovascular Gene Therapy Program at Duke University Medical Center. The ability to manipulate betaAR density or receptor desensitization in the diseased heart is of great interest since it may provide unique inotropic support and improve existing therapeutic strategies. Gene transfer to the heart in vivo is a powerful approach to study the specific role of GRKs and adrenergic desensitization in both normal and diseased myocardium. Currently, we are employing various in vivo gene delivery techniques using adenoviruses, in order to effectively express specific betaAR and GRK-based transgenes which may produce alterations in myocardial signaling and global cardiac function. We also have established several surgical models of compromised heart function to use in these gene transfer studies including experimental HF in rabbits and pigs, and cardiac transplantation models in rats and rabbits. In addition to studying the feasibility of gene therapy approaches to HF, the Cardiovascular Gene Therapy Program here at Duke has developed a molecular gene therapy strategy to prevention of pathological vascular smooth muscle intimal hyperplasia such as coronary artery restenosis after angioplasty. Our first clinical trial employing adenoviral-mediated gene therapy for restenosis is tentatively planned for the end of 2002.
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