Cholesterol Regulation of Endothelial K+ Channels

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DESCRIPTION (provided by applicant): Hypercholesterolemia-induced endothelial dysfunction is a major factor in the pathogenesis of cardiovascular disease but the mechanisms of endothelial injury are not well understood. Our general hypothesis is that suppression of endothelial K+ channels plays a key role in cholesterol-induced endothelial dysfunction. Our studies have shown that an increase in membrane cholesterol strongly suppresses endothelial inwardly-rectifying K+ (Kir) channels, the dominant K+ channels in aortic endothelial cells and putative flow sensors. Furthermore, we have shown that endothelial Kir channels are suppressed by atherogenic lipoproteins in vitro and by plasma hypercholesterolemia in vivo leading to endothelial membrane depolarization. Based on these findings, we hypothesize that cholesterol-induced suppression of endothelial Kir and impairment of its sensitivity to flow result in endothelial dysfunction and the loss of flow-induced endothelium-dependent vasodilatation. The goal of this study is to investigate molecular mechanisms responsible for cholesterol- and LDL-induced suppression of Kir and to determine its role in flow-induced vasodilatation. To address this hypothesis, we propose three specific aims: (1) To elucidate the molecular basis of the sensitivity of endothelial Kir channels to cholesterol by identifying specific lipid-protein (PIP2, lipid-protein interface of the transmembrane domains of the channels) and/or protein-protein (caveolin, scaffolding proteins) interactions responsible for the sensitivity of the channels to cholesterol. (2) To investigate the impact of LDL-induced Kir suppression in endothelial cells derived from different vascular beds and to evaluate potential strategies to rescue the current. Specifically, we will test whether Kir activity can be rescued by HDL, by cholesterol structural analogues and by cholesterol-insensitive Kir mutants. (3) To determine the impact of in vivo plasma hypercholesterolemia on the sensitivity of endothelial Kir to flow and on flow-induced endothelial membrane hyperpolarization and to elucidate the role of Kir channels in flow-induced vasodilatation of conduit and small coronary arteries under different cholesterol conditions. These studies will be performed in a porcine model of diet-induced hypercholesterolemia. We believe that achieving these goals will make a major contribution to the understanding of cholesterol-induced endothelial dysfunction, as well as of ion channel-cholesterol interactions in general. We also believe that these studies may provide the basis for developing new therapeutic strategies. PUBLIC HEALTH RELEVANCE: Cardiovascular disease (CVD) results in 40% of all deaths and results in serious morbidity in both men and women. It is well known that a key early step in the development of CVD is dysfunction of a thin single-cell layer of the inner lining of the blood vessels, called endothelium. Our goal is to determine the mechanisms responsible for the impairment of endothelial function by hypercholesterolemia, a major risk factor for the development of CVD. Our studies focus on hypercholesterolemia-induced loss of the ability of endothelial cells to respond to changes in blood flow, a hallmark of endothelial dysfunction. We intend to investigate the mechanisms by which an increase in plasma and cellular cholesterol suppress endothelial ion channels that are involved in endothelial-dependent control of vascular tone and the ability of endothelial cells to respond to blood flow. We believe that elucidating these mechanisms will contribute to the understanding of endothelial dysfunction that is associated with hypercholesterolemia and provide the basis for new therapeutic approaches.