线粒体融合蛋白-2 DNA甲基化在同型半胱氨酸致血管平滑肌细胞增殖中的作用

Hyperhomocysteinemia(HHcy) is an independent risk factor of atherosclerosis(AS), and Mitochondrial fusion protein-2(HSG/Mfn2) is a key gene which regulates cell proliferation. DNA methylation is an important way of gene expression regulation, but it has not been reported whether Hcy causes vascular smooth muscle cells(VSMCs) proliferation by the changes of HSG/Mfn2 abnormal DNA methylation. Therefore, this project plans to replicate Hcy-induced VSMCs proliferation of animal and cell models, and to detect the variation of HSG/Mfn2 via real time PCR, verify the function of HSG/Mfn2 by silence and overexpression, observe the proliferation of VSMCs intervened by Hcy, clarify the effects of HSG/Mfn2. It is clarified of the mechanism of HSG/Mfn2 DNA methylation in Hcy-induced VSMCs proliferation through detecting the changes of HSG/Mfn2 DNA methylation by high-throughput MethyLight assay and detecting the changes of regulatory factors such as DNMTs by using real-time PCR. Determining it's a key target for the regulation of DNA methylation through the silence and overexpression by using gene recombinant and RNA interference. This project seeks to clarify the mechanism of HSG/Mfn2, and to provide a experimental basis for Hcy-targeted therapy.

高同型半胱氨酸(Hcy)血症是动脉粥样硬化(AS)独立危险因子，而线粒体融合蛋白-2(HSG/Mfn2)是调控细胞增殖的关键基因，且DNA甲基化是基因表达调控的重要方式，但Hcy是否通过HSG/Mfn2 DNA甲基化异常引起VSMCs增殖未见报道。因此本项目拟复制Hcy引起VSMCs增殖动物和细胞模型，实时PCR等检测HSG/Mfn2的变化，分别过表达和沉默HSG/Mfn2，Hcy干预后，观察VSMCs增殖的变化，明确HSG/Mfn2的作用；高通量MethyLight法检测HSG/Mfn2 DNA甲基化的变化，实时PCR等检测DNMTs等调控相关因子的变化，揭示HSG/Mfn2 DNA甲基化在Hcy引起VSMCs增殖中的作用机制；基因重组和RNA干扰技术分别使DNMT1过表达和沉默，明确其是调控DNA甲基化的关键靶点；本课题旨在阐明HSG/Mfn2的作用机制，为Hcy靶向治疗提供实验依据。

高同型半胱氨酸(Hcy)血症是动脉粥样硬化(AS)独立危险因子，而线粒体融合蛋白-2(HSG/Mfn2)是调控细胞增殖的关键基因，且DNA 甲基化是基因表达调控的重要方式，但Hcy 是否通过HSG/Mfn2 DNA甲基化异常引起VSMCs 增殖未见报道。因此本项目拟复制Hcy 引起VSMCs 增殖动物和细胞模型，实时PCR 等检测HSG/Mfn2 的变化，分别过表达和沉默HSG/Mfn2，Hcy 干预后，观察VSMCs 增殖的变化，明确HSG/Mfn2 的作用；高通量MethyLight 法检测HSG/Mfn2 DNA 甲基化的变化，实时PCR 等检测DNMTs.等调控相关因子的变化，揭示HSG/Mfn2 DNA 甲基化在Hcy 引起VSMCs 增殖中的作用机制；基因重组和RNA 干扰技术分别使DNMT1 过表达和沉默，明确其是调控DNA 甲基化的关键靶点；本课题旨在阐明HSG/Mfn2 的作用机制，为Hcy 靶向治疗提供实验依据。