Cardiovascular diseases are the major cause of death for people over the age of 65 in the U.S. In particular, vascular endothelial cell function markedly declines, contributing to the profound vessel rigidity evident with age. Increased oxidative stress, cellular redox changes, and altered calcium homeostasis may be underlying factors in age-related endothelial cell dysfunction, which in turn, could be caused by mitochondrial decay. However, there is nothing known about mitochondrial function in vascular endothelial cells, the extent or precise nature of mitochondrial decay with age, or the significance of such decay on endothelial cell function. This proposal is intended to fill these significant gaps in endothelial cells with age? B) does mitochondrial decay affect endothelial cell function and vessel tone?, and C) does manipulation of cellular and/or mitochondrial antioxidant levels by addition of ascorbic acid (AA) or lipoic acid (LA) reverse the consequences of mitochondrial decay to endothelial function? We propose to investigate these questions in 3 specific aims: 1) Compare age-related changes in mitochondrial function and oxidant flux in freshly isolated porcine aortic endothelial cells (PAEC) from young and old pigs. These studies will define how aging affects mitochondrial function (bioenergetics, superoxide production and calcium homeostasis) in PAEC. In particular, we will test how mitochondrial function derived reactive oxygen species (ROS) alter cellular antioxidant status and oxidative stress. 2) Assess the impact of mitochondrial decay of PAEC function. We will correlate mitochondrial decay, especially in terms in terms of increased ROS and altered thiol redox status, to PAEC nitric oxide synthase (eNOS) activity and availability of nitric oxide (EDNO). 3) Determine whether manipulation of cellular antioxidant of cellular antioxidant and thiol redox status by addition of AA or LA in vitro (PAEC) or in vivo (guinea pig aortic rings) improves mitochondrial function and endothelial-dependent biological activity. AA or LA will be added to freshly isolated PAEC from young and old pigs to determine whether these agents reverse or """"""""mask"""""""" mitochondrial-dependent alterations to cellular redox status, oxidative stress (Aim 1) and endothelial biological activity (Aim 2). These studies will be augmented by supplemental feeding of guinea pigs with AA and LA to determine whether increased mitochondrial function (LA) and/or increased antioxidant and thiol redox status (AA or LA) may reverse the age-related loss in vascular tone in vivo.
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