MicroRNA-29通过自噬影响RPE细胞功能和脉络膜血管新生的研究

Recent evidences indicated that activation of autophagy in RPE cells protects against oxidative stress and possibly decreases the formation of choroidal neovascularization(CNV). Further evidences demonstrated that impaired autophagy in RPE contributes to the progression of age-related macular degeneration(AMD). Our previous studies indicated that (1) activation of autophagy in RPE protects RPE against oxidative stress; (2) The three members of microRNA-29 family are significantly decreased in choroidal-RPE tissue at the early stage of CNV. Our preliminary results indicated that miR-29 were possibly autophagy inducers in RPE cells. To date, there are limited reports about the regulation of autophagy by miR-29, through which miR-29 may repress CNV. In this project, we will further investigate whether miR-29 are autophagy inducers,the targeted autophagy related genes, and the pathways regulated by miR-29. We will further study the anti-oxidation mechanism in RPE mediated by miR-29. Finally, we will examine if miR-29 could decrease CNV in a CNV animal model. In summary, this project will aim to clarify the potential roles of miR-29 on the function of RPE cells and CNV through autophagy. The accomplishment will provide invaluable data in the roles of autophagy in RPE cells and CNV, and also a potential target for related ocular deseases including AMD.

RPE细胞可通过激活自噬，减少在氧化胁迫下的损伤和减少脉络膜血管新生，这两点可能在预防和延缓年龄相关性黄斑变性进展中发挥重要的作用。申请人前期研究发现，自噬的激活可以减少RPE细胞在氧化胁迫下的损伤；miR-29家族的三个成员的表达在小鼠脉络膜血管新生过程中显著下调。申请人最近的研究还发现miR-29可能正调控自噬。目前还没有关于miR-29调控自噬及其在CNV中的研究报道。本研究将在前期工作基础上，研究确定miR-29是否调控自噬及其调控的自噬相关的靶基因；检测miR-29调控自噬相关的信号通路；检测miR-29通过调控自噬对RPE细胞对抗氧化损伤和脉络膜-视网膜(内皮)细胞的血管形成能力的影响；最后动物体内检测miR-29是否可以抑制CNV的形成。本研究有助于增强我们对RPE细胞自噬功能的了解，为相关疾病包括AMD的治疗提供新的思路和作用靶点。

视网膜色素上皮细胞（Retinal pigment epithelium cell，RPE）位于视网膜和脉络膜中间，为视网膜传输营养和代谢产物，吞噬光感受器的外节膜盘，是血视网膜重要组成部分。研究表明RPE细胞可通过激活自噬保护RPE细胞减少氧化损伤，减少脉络膜血管新生，因此激活RPE细胞自噬可能在预防、延缓和治疗年龄相关性黄斑变性（Age-related Macular Degeneration，AMD）中具有重要的作用。申请人前期研究发现，自噬的激活可以减少RPE细胞在氧化胁迫下的损伤；并且miR-29家族的三个成员的表达在小鼠脉络膜血管新生过程中显著下调，前期研究还发现miR-29过表达后RPE细胞内自噬相关蛋白LC3-II表达上调，p62表达下调，表明miR-29可能调控自噬。本研究在前期工作基础上，主要研究了miR-29是否正向调控自噬及其机制，及miR-29通过调控自噬是否调控了细胞功能。我们发现miR-29可以正向调控自噬，并且研究确定了miR-29通过两个途径调控自噬：1，直接结合到靶基因LAMTOR1 (late endosomal/lysosomal adaptor, p18) 的3’-UTR，从而抑制mTOR锚定至溶酶体，从而抑制mTOR的磷酸化而激活自噬；2，miR-29抑制p85α的表达，从而抑制AKT/PKB介导的mTOR的磷酸化，从而激活自噬。我们还发现miR-29过表达激活自噬增加了RPE细胞清除αB-crystallin (R120G)突变体形成的蛋白沉积的能力，及减少脉络膜-视网膜(内皮)细胞血管形成的影响。这些研究结果有助于增强我们对RPE细胞自噬功能的了解，为相关疾病包括AMD的治疗提供新的思路和作用靶点。