miR-296-5p调控巨噬细胞极性的分子机制及其在动脉粥样硬化中的作用

MiR-296-5p is highly expressed in the progressive of atherosclerotic plaque, but its roles and mechanisms in atherosclerosis are still not clear. Previously through bioinformatics and preliminary experiments we have found that miR-296-5p inhibited peroxisome proliferator activated receptor delta (PPARδ) targetedly, which plays an important role in modulating macrophage polarization phenotype transformation and thereby affecting the process of atherosclerosis. To this end, the project will investigate more deeply on the direct effects and binding sites of miR-296-5p to PPARδ by report gene systems. In vitro experiments observe the effects of miR-296-5p on the macrophage polarization phenotype, inflammatory cytokine release and PPARδ expression. Furthermore, we investigate the roles of PPARδ on miR-296-5p regulating macrophage polarization phenotype transformation. Finally, we evaluate the regulatory role of miR-296-5p on plaque progression, macrophage polarization and the level of inflammation in vivo at systematic level. The launch of the project will reveal the effects and molecular mechanisms of mir-296-5p targeted inhibition PPARδ, promote macrophage polarization phenotype transformation and atherosclerosis, and provide new intervention avenues and drug targets for in vivo regulation of macrophage polarization phenotype and anti-atherosclerosis as well.

miR-296-5p在动脉粥样硬化（As）斑块进展期高表达，但其在As中的作用及机制尚不清楚。前期我们采用生物信息学及预实验发现miR-296-5p可靶向抑制过氧化物酶体增殖物激活受体δ（PPARδ），而PPARδ可调控巨噬细胞极性表型转换，进而影响As进程。为此本项目拟采用报告基因系统深入探讨miR-296-5p与PPARδ的直接作用和结合位点；体外实验观察miR-296-5p对巨噬细胞极性表型、炎症因子的释放和PPARδ表达的影响；进一步探讨PPARδ在miR-296-5p促巨噬细胞极性表型转换的作用；建立As动物模型，整体水平观察miR-296-5p对斑块进程、巨噬细胞极性、PPARδ表达和体内炎症水平的影响。项目的研究有望揭示miR-296-5p靶向沉默PPARδ、促进巨噬细胞极性表型转换及在As中的作用和分子机制，为体内调控巨噬细胞极性表型和抗As提供新的干预途径和药物作用靶点。

微小RNA（microRNA, miR）在动脉粥样硬化发病过程中扮演重要角色。近期，miR-296-5p被发现在动脉粥样硬化斑块中表达上调，但是它的具体功能尚不清楚。在本研究中，我们观察了miR-296-5p在动脉粥样硬化中的作用，并探索了潜在的分子机制。我们采用西方饮食饲喂ApoE-/-雄性小鼠8周，以构建动脉粥样硬化动物模型，同时经尾静脉注药完成相应处理。结果显示miR-296-5p antagomir（anti-miR-296-5p）可减少ApoE-/-小鼠的斑块负荷，并增加斑块的稳定性。此外，anti-miR-296-5p还能显著降低ApoE-/-小鼠体内的炎症反应，且不伴有血浆总胆固醇和高密度脂蛋白胆固醇水平的改变。在体外实验中，miR-296-5p可以促进巨噬细胞促炎型M1极化，而抑制抗炎型M2极化。通过生物信息学分析和荧光素酶报告基因检测，我们证实转录因子过氧化物酶体增殖激活受体δ（peroxisome proliferator-activated receptor δ, PPARδ）是miR-296-5p的下游靶基因。而且，PPARδ表达在体内外均受miR-296-5p负向调控。最后，救济实验显示miR-296-5p对巨噬细胞极化的调节效应可被PPARδ部分逆转。概而言之，我们的研究揭示了一种新的As发病机制，即miR-296-5p通过靶向抑制PPARδ使巨噬细胞极化由M2向M1倾斜，从而促进机体炎症反应，加速动脉粥样硬化发展。