ROLE OF SPHINGOMYELIN IN LIPOPROTEIN METABOLISM

描述(申请人提供)：这项建议的长期目标是研究神经鞘蛋白(SPH)在血浆中的生理作用，它是仅次于磷脂酰胆碱(PC)的最丰富的磷脂。与其在膜胆固醇代谢和信号转导中的作用相比，SPH在血浆脂蛋白代谢中的作用鲜有人关注，尽管其在动脉粥样硬化和衰老中的浓度显著增加。根据初步数据，PI提出SPH调节血浆脂解活性，逆转胆固醇转运途径，以及脂质过氧化反应，从而防止PC和胆固醇的过度周转和过早降解。PI将检验这样的假设，即SPH由于其结构与PC的相似，不仅竞争性地抑制LCAT，如他之前所展示的那样，还能竞争性地抑制其他能降解PC的脂解酶，如肝脂酶和分泌型磷脂酶A2。他还将通过使用单层技术、酶动力学和SPH分子的结构修饰来研究抑制这些活性的机制(S)。通过调节天然脂蛋白和重组高密度脂蛋白(RHDL)颗粒中SPH的浓度，研究SPH在脂蛋白之间交换游离胆固醇和胆固醇酯(CE)以及不同细胞选择性摄取HDLCE中的作用。由于CE的表面浓度决定了它的转移速率，因此将用[13C]核磁共振研究SPH对CE分配到rHDL表面脂中的影响。PI还将研究膜SPH在SR-B1受体功能中的可能作用。PI提出了一种新的假设，即SPH通过延缓脂质过氧化自由基的传播来抑制脂质过氧化，并且与浮力低密度脂蛋白相比，小而致密的低密度脂蛋白的氧化敏感性增加是由于前者的低SPH/PC比率。他将把各种低密度脂蛋白亚组分和合成脂质体的氧化性与它们的SPH/PC比率联系起来。通过使用SPH的结构类似物，通过氧化定位于脂蛋白表面或核心的荧光探针，以及通过研究SPH对膜流动性和横向扩散速率的影响，来研究SPH抑制脂质过氧化的机制。这些研究结果将为揭示SPH在脂蛋白中的生理功能及其与动脉粥样硬化和炎症的可能关系提供新的见解。

DESCRIPTION (provided by applicant): The long term objective of this proposal is to investigate the physiological role of sphingomyeIin (SPH) in plasma, where it is the most abundant phospholipid next to phosphatidylcholine (PC). In contrast to its functions in membrane cholesterol metabolism and signal transduction, the role of SPH in the metabolism of plasma lipoproteins has received little attention, although its concentration is significantly increased in atherosclerosis and aging. Based on preliminary data, the PI proposes that SPH modulates plasma lipolytic activities, reverse cholesterol transport pathways, and lipid peroxidation reactions, thereby preventing excessive turnover and premature degradation of PC and cholesterol.The PI will test the hypothesis that SPH, by virtue of its structural similarities to PC, competitively inhibits not only LCAT, as he previously showed, but also other lipolytic enzymes that hydrolyze PC, such as hepatic lipase and secretory phospholipases A2. He will also investigate the mechanism(s) involved in the inhibition of these activities, by employing monolayer techniques, enzyme kinetics, and structural modifications of the SPH molecule. The role of SPH in the exchange of free cholesterol and cholesteryl ester (CE) between lipoproteins, and in the selective uptake of HDL CE by various cells in culture will be studied by manipulating the SPH concentration of native lipoproteins and recombinant HDL (rHDL) particles. Since the surface concentration of CE determines its transfer rate, the effect of SPH on the partitioning of CE into surface lipids of rHDL will be investigated by [13C] NMR. The possible role of membrane SPH in the function of SR-B1 receptor will also be investigated.The PI proposes to test the novel hypothesis that SPH inhibits lipid peroxidation by retarding the propagation of lipid peroxy radicals and that the increased oxidative susceptibility of small dense LDL, compared to buoyant LDL, is due to the low SPH/PC ratio in the former. He will correlate the oxidizability of various LDL subfractions and synthetic liposomes with their SPH/PC ratios. The mechanism by which SPH inhibits lipid peroxidation will be investigated by using structural analogs of SPH, by oxidizing fluorescent probes which are specifically localized in the surface or core of lipoproteins, and by studying the effects of SPH on membrane fluidity and lateral diffusion rates.The results from these studies should provide new insights into the physiological functions of SPH in lipoproteins, and its possible relevance to atherosclerosis and inflammation.