MicroRNA靶向PI3K/Akt信号途径调控视网膜缺血再灌注损伤的研究

Retinal ischemia reperfusion (RIR) injury is a complex pathophysiologic process caused by many factors, closely related to many ophthalmological diseases and threatened visual acuity. PI3K/Akt singal pathway has been shown to paly a critical role in RIR, but sigle PI3K pathway couldn't explain the machenism very well. miRNAs are small noncoding RNAs, approximately 19~25 nucleotides in length, that may be pivotal regulators in physiological and pathologic states.Recent studies have shown there are specific miRNAs related to retinal diseases. But the function and machenism of miRNAs in RIR are still not clear. Previous study showed the expressions of miRNAs after RIR are abnormal, indicating miRNAs may be a causative factor for the development of RIR. We preliminarily screened a RIR-related miRNA named miR-182 whose targeted gene is Akt. This study aimed at determing the effect of up- or down-regulating miR-182 on the regulation of PI3K/Akt pathway in RGC and RIR retina. Our findings may provide new sights into understanding the signal pathway regulation in pathogenesis of RIR and offer new therapeutic targets for RIR.

视网膜缺血再灌注损伤(RIR)是由多种因素引起的复杂病理生理过程，与眼科许多疾病的发展密切相关。PI3K/Akt信号途径异常与其发生发展关系密切，但单一的PI3K途径不能很好解释RIR的分子机制。miRNA是近年来发现的一类19~25个核苷酸大小的内源性单链RNA，在生理和病理生理状态下发挥重要调控作用，已经发现视网膜相关疾病特异表达的miRNA，而在RIR中的作用及分子机制尚不清楚。我们前期研究表明：RIR后miRNA表达异常，提示miRNA在RIR发生/发展中可能起着重要作用，并利用芯片结合生物信息学技术初筛出一个与RIR相关的以Akt基因为靶点的miRNA- - miR-182。本课题通过对RGC及RIR视网膜上调或下调miR-182，探讨其如何影响PI3K/Akt通路来调控RIR，研究结果将利于阐明RIR发病的信号调控机制，并为治疗RIR选择新的治疗靶点提供理论依据。

视网膜缺血再灌注损伤(RIR)是由多种因素引起的复杂病理生理过程，与眼科许多疾病的发展密切相关，PI3K/Akt信号途径异常与其发生发展关系密切，我们的研究发现在视网膜缺血再灌注过程中，miR-182基因持续上调，导致其靶基因Akt表达减少，从而调控PI31k/Akt信号传导途径，诱导细胞凋亡，最终引发RIR的发生发展，因此本研究中我们构建miR-182 mimic及miR-182inhibitor 载体来上调、下调视网膜神经节细胞及不同时间点的RIR大鼠模型中miR-182的表达，通过离体及载体实验，结果证实AKT是miR-182的靶基因，miR-182可通过抑制AKT表达，调控PI3K/Akt信号传导途径，最终影响NF-kB及caspase 3的水平，引起RIR中炎症反应，诱导RGCs的凋亡，通过应用miR-182 mimic及miR-182 inhibitor转染RGCs及RIR大鼠模型，我们发现转染了miR-182的RGCs凋亡增加，RIR损伤加重，视网膜功能明显下降；而转染了miR-182 inhibitor的RGCs细胞凋亡减少，RIR损伤明显减轻，而在其中起到关键作用的信号传导途径就是AKT信号通路，通过对AKT因子的调控，可对其下游炎症基因NF-kB及凋亡基因caspase 3进行调控，最终可起到阻止RIR导致视网膜损伤的作用，从而为RIR新的治疗靶点和新药物的研制提供理论和实验依据。