动脉粥样硬化（AS）是一种慢性炎性疾病，过去认为饱和脂肪酸是诱导炎症反应促发AS的因素，我们前期研究发现中链脂肪酸之一辛酸作为饱和脂肪酸却改善血液胆固醇代谢、促进胆固醇逆转运、增加胆汁酸及粪胆固醇排出，是抗AS因素，还发现辛酸抑制肥胖小鼠小肠TLR4的mRNA表达。TLR4介导的炎症反应与AS密切相关。我们假设辛酸经TLR4及其通路抑制AS的炎症反应。通过与硬脂酸、棕榈酸、n-3脂肪酸对照，观察辛酸对apoE-/-小鼠模型的AS炎症反应及TLR4通路中相关蛋白表达的影响；以小鼠RAW264.7细胞株为研究对象，采用siRNA沉默TLR4等实验技术探讨分子机制；最后采用免疫共沉淀结合GC-MS检测法探讨辛酸对细胞膜TLR4的作用方式。本项目将为研究辛酸抗炎、抗AS的作用及机制提供实验依据。

Atherosclerosis (AS) is a chronic inflammatory disease. Saturated fatty acids were thought to induce inflammation in AS. Our previous studies found that caprylic acid, one of medium-chain fatty acids promoted reverse cholesterol transport, increased fecal bile acid and cholesterol discharge, was a protective factor for AS. We also found that caprylic acid could suppress toll like receptor 4(TLR4) mRNA expression in the small intestine of high-fat diet induced obese C57BL/6J mouse. Since TLR4 pathway is closely related to inflammation and AS, we hypothesize that caprylic acid could inhibit inflammation of AS by TLR4 and its signaling pathway. We will observe the inhibition of AS inflammation of caprylic acid in apoE-/- mouse models, and the related protein expression of TLR4 signaling pathway, compared with stearate acid and.palmitic acid and n-3 fatty acid. TLR4 gene-siRNA were carried out to investigate the molecular mechanisms in cultured RAW264.7 cells strains. Finally, immunoprecipitation and GC-MS were used to explore the model of combining of caprylic acid and TLR4 molecular in membrane. This study will provide a new proof for medium chain fatty acids in inhibitory effect of inflammation and AS.

动脉粥样硬化（AS）是一种慢性炎性疾病，本项目在课题组前期研究基础上，探讨中链脂肪酸中的辛酸通过TLR4分子途径抑制机体炎症反应进而防治AS的形成。本项目动物实验以雄性4-6周龄apoE-/-小鼠50只为研究对象，给予含2%浓度辛酸、癸酸、硬脂酸和亚麻酸的高脂饲料喂养16周，同时设高脂饲料喂养的空白对照组，分别观察动脉粥样斑块形成情况、血清脂代谢指标、炎性因子表达以及动脉壁上TLR4途径相关分子的基因和蛋白表达的变化。细胞实验选择RAW264.7细胞株，采用不同浓度辛酸及棕榈酸、DHA干预12h、24h、48h，同时设空白对照组及LPS损伤组，通过测定细胞裂解液中炎性因子及TLR4途径相关分子mRNA表达，以确定辛酸的最佳作用时间和浓度。siRNA-TLR4及TLR4质粒转染实验分别进行炎性因子及TLR4途径相关分子mRNA和蛋白表达的测定。研究结果包括：辛酸可以改善高脂饮食诱导的apoE-/-小鼠动脉壁斑块的形成，并降低血清胆固醇水平，下调血清和动脉壁上TLR4途径相关分子的mRNA表达和蛋白表达，包括降低血浆TNF-a和MCP-1水平，下调动脉壁TLR4、MyD88、TAK1、IKKa/b、NK-kB和TNF-a的mRNA表达。细胞实验结果显示：100μmol/L辛酸终浓度作用24小时时对LPS损伤的RAW264.7细胞的炎症反应修复最佳，表现在该组细胞裂解液中TNF-a、MCP-1等水平最低，TLR4途径分子如MyD88、NK-kB和TNF-a等mRNA及蛋白表达也最低；siRNA-TLR4沉默后，辛酸组的这种抗炎作用减弱；而质粒转染实验的mRNA和Westernblotting结果均显示，辛酸组降低MyD88、NK-kB和TNF-a等水平的作用增强。结论：辛酸可抑制机体炎症反应，作用类似于n-3脂肪酸，其机制可能为通过TLR4/MyD88信号通路降低炎性分子表达，主要是NK-kB和TNF-a，进而防治小鼠AS形成。本项目研究结果为中链脂肪酸抗炎、抗AS的作用及机制提供了实验依据，并为开发防治AS的功能性油脂提供了新途径和新思路。