Impaired insulin (INS) sensitivity is a common feature of disease states such as obesity, hypertension and diabetes. A western diet (WD), especially characterized by excess intake of high fat, high sucrose and carbohydrates, is a major factor in the increased prevalence of hypertension and diabetes. These co- morbidities may be driven by a decrease in INS-mediated vasorelaxation and glucose transport in cardiovascular (CV) and skeletal muscle tissue. In addition to our WD, several other mechanisms, such as enhanced activation of the renin-angiotensin-aldosterone-system (RAAS) and associated abnormalities in INS metabolic signaling, may help explain the linkage between INS resistance and hypertension. There is emerging evidence that enhanced activation of the RAAS may promote INS resistance through the mammalian target of rapamycin (mTOR)/S6 kinase 1 (S6K1) signaling pathway. mTOR, a highly conserved nutrient sensor, modulates INS metabolic signaling through its phosphorylation [(p)] of S6K1, an evolutionarily conserved serine (Ser) kinase. Evidence is mounting that chronic activation of S6K1, by excessive nutrients, promotes INS resistance in fat, liver and skeletal muscle tissue through increased Ser (p) of the critical INS signaling/docking molecule, INS receptor substrate protein 1 (IRS-1), leading to impaired phosphoinositol 3 kinase (PI3-K) engagement and protein kinase B (Akt) stimulation. Our recent work indicates that S6K1 is activated by angiotensin II (Ang II) and aldosterone in CV tissue leading to diminished INS metabolic signaling and biological consequences, such as impaired nitric oxide (NO)-mediated vascular relaxation. This proposal seeks to investigate novel molecular mechanisms by which Ang II, aldosterone and a WD individually and collectively promote INS resistance in CV and skeletal muscle tissue. To evaluate the CV functional effects of INS metabolic signaling, we will utilize our state-of-the-art rodent imaging center. In the INS resistant state, myocardial and skeletal muscle glucose uptake and metabolism is impaired, leading to diastolic dysfunction, attenuated myocardial and skeletal muscle blood flow, and impaired ischemic reconditioning. We have shown that both impaired INS stimulated glucose uptake and diastolic dysfunction are related to impaired systemic and myocardial INS metabolic signaling in models of obesity and increased tissue RAAS expression. For this proposal, we will utilize novel knockout and knockdown strategies, as well as innovative rodent imaging tools, to evaluate the impact of increased S6K1 signaling (Ang II/aldosterone and/or WD) on myocardial function and coronary and skeletal microvascular blood flow responses to INS metabolic signaling. To address Objective 1, we will examine the relationship between Ang II/aldosterone/WD and S6K1 activation and INS signaling in primary cultured endothelial cells, vascular smooth muscle cells and cardiomyocytes. Metabolic signaling results will be correlated to functional measures including NO production, cardiomyocyte glucose transport and diastolic relaxation. To further explore the collective, as well as the independent, roles of Ang II/aldosterone and a WD on S6K1, Objective 2 will focus on in vivo/ex vivo effects in the S6K1-/- and C57BL/6 mice treated with Ang II/aldosterone that produces a slow pressor response and/or a WD. A cohort of animals will be treated with an AT1R blocker or a mineralocortiod receptor in doses determined by telemetry to have no effect on blood pressure in mice. INS resistance will be assessed by hyperinsulinemic and euglycemic clamp, cardiac PET scanning, ex vivo IRS-1 (p) and INS metabolic signaling, and glucose uptake in heart and skeletal muscle. Finally, in vivo INS mediated skeletal muscle arteriolar and ex vivo coronary arteriolar, NO induced relaxation, and in vivo cardiac glucose uptake and diastolic relaxation will be related to ex vivo S6K1 activity and IRS-1 site specific Se vs. tyrosine (p) and the resultant downstream IRS-1/PI3-K/Akt signaling.
Insulin resistance often evolves into Type 2 diabetes and is an important contributor to the development of hypertension. Diabetes and hypertension contribute enormously to the morbidity, mortality, and health care costs of the Veteran population. Elucidation of the biochemical pathways affecting insulin signaling will not only contribute to th identification of access points for potential new drug interventions but will also address the current epidemics of Type 2 diabetes and hypertension. Further, the information gleaned from the proposed studies might also set the stage for future work needed to develop optimal hygienic and medical interventions in order to improve clinical outcomes in current and future Veterans.
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