多途径探讨蛹虫草多糖TY258促胆固醇逆向转运的分子机制

胆固醇逆向转运（RCT）是机体抗动脉粥样硬化（AS）的重要机制之一。申请团队前期发现：蛹虫草多糖（CMPS）粗品可显著降低C57BL/6小鼠血脂水平，促进RCT，但机制不明。多糖结构是决定其活性的物质基础。申请团队采用分离纯化技术结合胆固醇外排活性筛选实验发现：纯品TY258促RAW264.7泡沫细胞3H-胆固醇外排效果良好。在已明确TY258结构和预实验的基础上，我们提出：TY258为CMPS的主要活性成分，其可通过促进RCT降低血脂。胆固醇酯转运蛋白（CETP）与脂代谢密切相关，但小鼠缺乏该蛋白。本项目拟采用CETP转基因小鼠和细胞模型，运用同位素示踪等关键技术，结合分子生物学手段，多途径探讨TY258促 RCT的分子机制，并在国际公认的AS模型“载脂蛋白E敲除小鼠”上验证其效果，期待为锁定TY258降脂的调控靶点提供依据，为开发构-效关系明确的多糖降脂药物提供理论支撑。

Atherosclerosis (AS) induced by hyperlipidemia is the basic pathology process of cardiovascular and cerebrovascular diseases, and reverse cholesterol transport (RCT) is one of the most important lipid-lowering mechanisms. Our preliminary experiments indicated that the crude extract of polysaccharides from Cordyceps militaris (CMPS, 50 mg/Kg) could decrease plasma lipid levels and promote macrophage RCT in C57BL/6 mice, however, the underlying mechanisms are not clear. Structure determines the function of polysaccharide. We separated and purified CMPS and bioassay guided screening in vitro revealed TY258, one of the purified polysaccharides, exhibited the best effect on promoting the HDL3 mediated and 3H-labelled cholesterol efflux from RAW264.7 macrophages. Now we have elucidated the fine structure of TY258 by 1 and 2 dimensional nuclear magnetic resonance (1D, 2D-NMR) and methylation analysis. Based on our previous work, we put forward the assumption that TY258 is the major effective component of CMPS, and it could regulate plasma lipids by promoting RCT. Cholesteryl ester transfer protein (CETP) is closely associated with lipid metabolism, while mice are lack of this protein. In the present study, we will apply CETP transgenic (CETP-tg) mice model and cell model to clarify the regulatory effect of TY258 on RCT and the underlying mechanisms by a series of key technologies, such as isotopic tracing and molecular biological techniques (RT-PCR, Western blot, Gene silencing, etc). In the last step we plan to verify the protective effect of TY258 on the internationally recognized AS model of apolipoprotein E knockout (apoE-/-) mice. The expected results could provide regulating targets of TY258 for lipid-lowering and AS protection and offer theoretical foundation for clinical transformation of TY258. This would be beneficial to exploit polysaccharide lipid-lowering drug with specific structure-activity relationship and own intellectual property rights for our country.