Regulation of Cellular Cholesterol Through Oxysterol-Membrane Interactions

描述(由申请人提供)：严格控制细胞和血浆胆固醇对细胞的正常运作至关重要，因为过量的游离胆固醇对细胞有害，并与动脉粥样硬化和心脏病有关。细胞内胆固醇的动态平衡是由酶促形成的含氧胆固醇衍生物(称为氧化甾醇)来调节的。氧化甾醇可以抑制胆固醇从头合成和脂蛋白胆固醇摄取相关基因的表达，并作为核受体的内源性配体，激活胆固醇分解代谢和外流途径。虽然氧化甾醇在胆固醇稳态的急性调节中的重要性已为人所知，但氧化甾醇发挥作用的确切分子机制仍有待阐明。我的中心假设是，氧化甾醇通过非对映体选择性的膜无序效应来发挥其胆固醇调节作用。为了验证这一假设，我将(1)合成已知的25-HC拮抗剂LY295427的天然形式(NAT)和对映体(ENT)形式，(2)检测NAT和ENT-LY295427-膜相互作用的对映选择性以及LY295427对25-HC膜无序特性的影响，以及(3)确定LY295427抑制25-HC介导的培养细胞中的类固醇稳态反应的对映选择性。该项目的长期目标是阐明维持细胞胆固醇平衡的分子机制，并为如何利用膜结构的变化传递调节信号提供洞察力。拟议的研究可能有助于确定操纵细胞处理胆固醇和治疗动脉粥样硬化的药理靶点。 公共卫生相关性：高水平的血浆脂蛋白胆固醇(如低密度脂蛋白)与动脉粥样硬化和心脏病有关。了解细胞和全身胆固醇水平的调节机制将为开发降低胆固醇的治疗方法提供新的途径，并可能减少心血管疾病。

DESCRIPTION (provided by applicant): Tight regulation of cellular and plasma cholesterol is crucial to proper cellular functioning, as excess free cholesterol is toxic to cells and is associated with atherosclerosis and heart disease. Cellular cholesterol homeostasis is regulated by enzymatically formed oxygenated cholesterol derivatives called oxysterols. Oxysterols can suppress expression of genes responsible for de novo cholesterol synthesis and lipoprotein cholesterol uptake, and serve as endogenous ligands for nuclear receptors, which activate cholesterol catabolic and efflux pathways. While the importance of oxysterols in the acute regulation of cholesterol homeostasis is known, the precise molecular mechanisms through which oxysterols exert their effects remain to be elucidated. My central hypothesis is that oxysterols exert their cholesterol-regulatory effects through non- enantioselective membrane-disordering effects. To test this hypothesis, I will (1) synthesize both the natural (nat) and enantiomeric (ent) forms of LY295427-a known antagonist of 25-HC, (2) examine the enantioselectivity of nat- and ent-LY295427-membrane interactions and the effect of LY295427 on the membrane-disordering properties of 25-HC, and (3) determine the enantioselectivity of LY295427 inhibition of 25-HC-mediated sterol homeostatic responses in cultured cells. The long-term objectives of this project are to elucidate the molecular mechanisms underlying the maintenance of cellular cholesterol balance, and to provide insight into how alteration in membrane structure can be used to relay regulatory signals. The proposed studies may help to identify pharmacological targets for manipulation of the cellular handling of cholesterol and treatment of atherosclerosis. PUBLIC HEALTH RELEVANCE: High levels of plasma lipoprotein cholesterol (e.g., in the form low-density lipoproteins (LDL), are associated with atherosclerosis and heart disease. Understanding the mechanisms through which cellular and whole-body cholesterol levels are regulated will provide new approaches to developing cholesterol-lowering treatments, and may reduce cardiovascular disease.