长链非编码RNA-PILR充当p53-Parkin分子脚手架调控糖尿病心肌病线粒体自噬的机制研究

Diabetic cardiomyopathy (DCM) lacks a detailed mechanism. The inhibition of mitochondrial autophagy (mitophagy) in DCM leads to the imbalance of mitochondrial dynamic balance. Long chain non coding RNA (lncRNA) plays its role in DCM mitophagy and its mechanism have not been elucidated. We used lncRNA microarray to screen high expression of lncRNA-PILR (p53 interact lncRNA) in myocardial tissue of DCM patients. Si-PILR can promote mitophagy and reduce myocardial damage induced by high glucose, suggesting lnc-PILR is closely related to DCM. Further using of RNA pull down analysis suggested that PILR could be combined with cytoplasmic p53 and Parkin simultaneously. Accordingly, we speculate that PILR may be used as a "molecular scaffold" to mediate the combination of p53 and Parkin to inhibit mitophagy.In this project, the overexpression of lnc-PILR and the interference of lentivirus transfection into DCM mice and high-glucose human AC16 cell lines in vitro were studied to elucidate the molecular mechanism of PILR-p53-Parkin complex inhibiting DCM mitophagy. The present study indicates a new mechanism of LncRNA in pathogenesis of DCM, and provides a theoretical basis for the development and treatment of DCM.

糖尿病心肌病（DCM）发病机制不清。线粒体自噬（mitophagy）和长链非编码RNA（lncRNA）在DCM病理过程中扮演重要角色。前期研究通过lnc芯片筛选出DCM患者心肌高表达lnc-PILR。体外沉默PILR促进高糖心肌细胞mitophagy，减轻高糖引起的细胞损伤，表明lnc-PILR与DCM密切相关；进一步采用RNA pull down揭示lnc-PILR可同时与胞质p53和Parkin结合，推测lnc-PILR充当p53-Parkin复合体的分子脚手架抑制DCM mitophagy。本项目以lnc-PILR过表达及干扰慢病毒转染高糖人AC16心肌细胞及DCM小鼠为研究对象，阐明lnc-PILR在DCM mitophagy中的作用，并探究lnc-PILR与p53-Parkin复合体相互作用抑制DCM mitophagy的分子机制，为DCM发生发展及治疗提供理论基础。

糖尿病心肌病（DCM）发病机制不清。线粒体自噬（mitophagy）和长链非编码RNA（lncRNA）在DCM病理过程中扮演重要角色。通过lncRNA芯片分析筛选出了LncRNA -ENST00000440196.2在DCM心肌组织中高表达，命名为lnc-PILR。体外沉默PILR促进高糖心肌细胞mitophagy，减轻高糖引起的细胞损伤，表明lnc-PILR与DCM密切相关；进一步采用RNA pull down揭示lnc-PILR可同时与胞质p53和Parkin结合，推测lnc-PILR充当p53-Parkin复合体的分子脚手架抑制DCM mitophagy。本项目以lnc-PILR过表达及干扰慢病毒转染高糖人AC16心肌细胞及DCM小鼠为研究对象，通过基因芯片微阵列、双荧光素酶基因报告系统、荧光原位杂交(FISH)、RNA pull down、RNA结合蛋白免疫沉淀(RIP)、心肌细胞培养、免疫荧光技术、激光共聚焦显微镜技术、流式细胞计量术、Western Blot、染色质共沉淀(CHIP)、免疫共沉淀（Co-IP）、Real-Time PCR、质粒转染、慢病毒质粒构建等方法，初步阐明了lnc-PILR在DCM mitophagy中的作用，探究了lnc-PILR与p53-Parkin复合体相互作用抑制DCM mitophagy的分子机制。综上，该研究发现了p53和parkin通过一个共享的分子脚手架 lnc-PILR协同调节DCM mitophagy。这是DCM进展过程中一个重要的、先前未表征的调节回路，lnc-PILR/p53/parkin 可作为治疗 DCM相关mitophagy的潜在治疗靶点。