转录因子Mipu1抑制mTOR表达诱导自噬抗动脉粥样硬化研究

Mipu1(myocardial ischemic preconditioning upregulated protein 1) is a nuclear transcription factor, which displays a powerful effect in protecting from oxidative stress-induced cell injury. Our study indicated that Mipu1 inhibited the formation of foam cell from macrophages and induced macrophage autophagy. This study is to observe the effects of Mipu1 on macrophage autophagy by transmission electron microscope, immunofluorescence and western blot in macrophages isolated from wild mice and Mipu1-/- mice. Chromatin immunoprecipitation (ChIP), luciferase assays, gene transfection and RNA interference were used to determine whether Mipu1 regulates mTOR expression and its role in the formation of foam cell from macrophages. Oil red O staining, ELISA and high performance liquid chromatograph were used to observe the effects of Mipu1 and mTOR inhibition on atherosclerosis(As) formation, inflammatory cytokines and blood lipid level in apoE-/-Mipu1-/-mouse. This research will discover the effect of Mipu1 on induction of autophagy through inhibiting mTOR expression in As formation. Our study will be benefit to understand the molecular mechanism of Mipu1 inhibiting the formation of foam cell from macrophages in As formation, and it will provide new idea for the antiatherosclerosis effect of Mipu1.

心肌缺血预适应上调蛋白1(Mipu1)是一种氧化应激时具有细胞保护作用的核转录因子。本课题组近来发现Mipu1能抑制巨噬细胞泡沫化，诱导巨噬细胞自噬。本项目拟在野生型和Mipu1-/-小鼠来源的腹腔巨噬细胞中采用电镜、免疫荧光和Western blot等观察Mipu1对巨噬细胞自噬的影响；采用染色质免疫共沉淀、荧光素酶报告基因活性检测、基因转染和RNA干扰等探讨Mipu1如何调控自噬相关基因mTOR的表达及其在巨噬细胞泡沫化中的作用；在ApoE-/-和Mipu1-/-双敲除小鼠上复制As模型采用油红O染色、ELISA和高效液相色谱等观察Mipu1及干扰mTOR对As斑块、炎症因子及血脂水平的影响。本研究可明确Mipu1抑制mTOR表达诱导自噬在As发病中的作用。该研究可望阐明Mipu1抑制巨噬细胞泡沫化的分子机制，为Mipu1抗As提供理论依据。

项目的背景.过表达Mipu1可抑制由oxLDL诱导的RAW264.7细胞的泡沫化，且Mipu1可在转录水平调控CD36表达，抑制Mipu1表达可抑制由oxLDL诱导的巨噬细胞自噬。.主要研究内容.Mipu1对巨噬细胞及内皮细胞自噬和炎症反应的影响及机制，以及Mipu1调节自噬在动脉粥样硬化（As）与主动脉夹层（AD）中的作用。.重要结果.1.H2O2可诱导Mipu1表达；Mipu1可上调内皮细胞自噬及减轻氧化应激损伤。.2.缺血再灌注损伤时Mipu1表达上调，Mipu1可拮抗由ox-LDL诱导的人脐静脉内皮细胞损伤。.3.Mipu1通过mTOR抑制巨噬细胞炎症反应和有氧糖酵解。.4.Mipu1通过调控mTOR抑制As和AD形成。.科学意义.本研究可明确Mipu1抑制巨噬细胞泡沫化及巨噬细胞炎症反应的分子机制，可望阐明Mipu1调控自噬在As和主动脉瘤等心血管疾病发病中的作用，为Mipu1在As和主动脉瘤中的作用提供新的理论依据。