Diabetic cardiomyopathy is a complex disorder that emanates from the chronic and excessive use of fatty acids to fuel contractile function in diabetic myocardium due to the lack of insulin signaling. However, the nearly exclusive use of fatty acids for fuel in diabetic myocardium results in widespread metabolomic dysregulation that precipitates multiple deleterious alterations in membrane structure and function. Consequences of these membrane-mediated abnormalities in diabetic myocardium include hemodynamic compromise, defective excitation-contraction coupling and mitochondrial dysfunction that collectively conspire to promote the progression of heart failure in diabetic patients. Moreover, the profound alterations in substrate utilization in diabetic myocardium result in the accumulation of multiple dysregulated metabolites that lead to maladaptive alterations in interwoven cardiac myocyte signaling networks. Previously, through genetic, pharmacologic and chemical biological approaches, we have identified three major phospholipases and lipases in myocardium iPLA2? (PNPLA9), iPLA2? (PNPLA8), and iPLA2? (PNPLA2;ATGL) that likely serve as principal mediators of myocardial hemodynamic dysfunction, electrophysiologic alterations and maladaptive remodeling in diabetic myocardium. Recently, we demonstrated that iPLA2g and its downstream signaling metabolites are key regulators of the mitochondrial permeability transition pore which is responsible for necrosis, necroptosis, and electrical instability in diabetic myocardium subjected to ischemia. Accordingly, in Specific Aim 1, we will use the novel cardiac myocyte specific iPLA2g conditional knock out mouse we generated to determine if iPLA2g loss of function attenuates acute ischemic injury, electrophysiologic instability and the maladaptive generation of lipid 2nd messengers in diabetic myocardium. Furthermore, we demonstrated that exposure of mitochondria to calcium ion results in the activation of iPLA2g leading to the release of arachidonic acid, 2-arachidonoyl lysophosphatidylcholine, and the subsequent production of multiple downstream biologically active lipid 2nd messengers. Accordingly, iPLA2g-dependent alterations in lipid 2nd messenger production will be examined employing integrative mass spectrometric platforms we developed in conjunction with the cardiac myocyte specific iPLA2g loss of function mouse.
In Specific Aim 2, we will determine the molecular mechanisms through which acyl-CoA facilitates CaMKII phosphorylation and activation of iPLA2b. The activating phosphosite(s) will be identified, mutated and their mechanistic importance in CaMKII-mediated activation of iPLA2b in diabetic myocardium and diabetic myocardium rendered ischemic will be explored.
In Specific Aim 3, the role(s) of iPLA2z (ATGL;PNPLA2) in catalyzing the bidirectional flux of lipids through triglyceride hydrolysis, transacylation and acyltransferase activities will e determined. The participation of iPLA2z in generating lipid 2nd messengers in diabetic myocardium will be examined using cardiac myocyte specific iPLA2z null mice and the effects of iPLA2z genetic ablation on myocardial function in the diabetic state will be explored. Collectively, these studies are a synergistic multidisciplinary approach to identify the chemical mechanisms mediating diabetic cardiomyopathy.
Diabetes is rapidly increasing in industrialized societies due to an increase in caloric intake that typically contains excessive amounts of fat. The major cause of death from diabetes is heart disease due to heart failure or sudden death. In diabetic patients, the heart uses excessive amounts of fat to fuel contractile function. However, the chronic and excessive reliance of fat to fuel heart contractions results in widespread changes in the structure and function of heart cell membranes. These studies are an integrated approach to understand the mechanisms mediating membrane dysfunction in diabetic hearts to identify novel therapies for heart disease in diabetic patients.
|Liu, Xinping; Moon, Sung Ho; Mancuso, David J et al. (2013) Oxidized fatty acid analysis by charge-switch derivatization, selected reaction monitoring, and accurate mass quantitation. Anal Biochem 442:40-50|
|Yang, Kui; Dilthey, Beverly Gibson; Gross, Richard W (2013) Identification and quantitation of fatty acid double bond positional isomers: a shotgun lipidomics approach using charge-switch derivatization. Anal Chem 85:9742-50|