脂联素（APN）是新近发现的脂肪细胞因子，具有明确的心血管保护作用。申请者前期系列研究已证实：APN可降低血管内皮氧化/硝化应激，改善高脂血症内皮功能；而高脂血浆可抑制APN活性，提示APN活性降低致其内皮保护作用丧失，可能参与高脂血症早期内皮功能障碍的形成。但高脂血浆抑制APN活性的分子机制目前尚一无所知。申请者最新预实验结果提示，高脂血浆中多种细胞因子特别是TNF-α与APN偶联并相互作用可能是抑制其活性的重要因素。本研究拟采用免疫共沉淀、质谱分析及双向凝胶电泳等技术，首先明确高脂血浆中与APN偶联的物质成分及其生化特性，进一步在细胞和整体水平验证其对APN活性的影响，最终确定可阻断高脂血浆对APN偶联抑制的分子靶点，并验证这些干预措施的内皮保护效应。本项目的完成将不仅为高脂血症血管内皮功能障碍提供全新的分子机制，且可望为以提高APN生物活性而保护血管的这一全新策略提供有力的实验证据。

Adiponectin (APN) is a novel adipocyte-derive molecule with strong cardiovascular protective actions. We have recently demonstrated that administration of APN protects vascular endothelial function in hyperlipidemic animals by reducing oxidative/nitrative stress; Additional study further demonstrated that APN might be modified/inhibited by unknown molecules present in hyperlipidemic plasma, suggesting that post-translational modification and functional inhibition of APN contribute to vascular endothelial dysfunction in hyperlipidemia. However, the identity of and mechanisms by which the hyperlipidemic plasma may bind and modify APN function remains completely unknown. Our most recent preliminary experiments suggest that multiple cytokines, particularly TNFα, can bind with APN. The specific aims of the current grant application are 1) defining the identity of those molecules that bind with APN by co-immunoprecipitation, mass spectrum analysis, and 2D gel electrophoresis; 2) determining the effect of this APN-binding/modification on APN biological function in cultured endothelial cells; and 3) identifying the molecular interventions that may block APN modification by hyperlipidemic plasma, and verifying their vascular protective effect in hyperlipidemic animals. Successful completion of these experiments will not only advance our understanding of molecular mechanisms of endothelial dysfunction in dyslipidemia, but may also provide experimental evidence supporting enhancement of APN function as a novel therapeutic strategy in hyperlipidemic patients.