miR-34a-HK1通路在年龄相关性白内障发生过程中的分子机制

Cataract is the leading cause of blindness, and its pathogenesis is unclear. Energy metabolism is the basic metabolic activity of organisms, which is essential to maintain the normal function of the lens. The lens is in a relatively hypoxic environment, obtaining energy through glycolysis, and the lens mainly contains HK1 which is the rate limiting enzyme to glycolysis. When the function of HK1 is inhibited, it will cause the energy supply of cells to be insufficient, which will cause some pathological changes. miRNAs are involved in the regulation of transcription. Our preliminary studies have shown that the expression of miR-34a in lens epithelial cells of age-related cataracts is significantly higher than that of transparent lens. Besides, miR-34a could induce lens epithelial cell apoptosis. However, there is no research on the mechanism of miR-34a-HK1 in age-related cataractogenesis. In this study, it is speculated that miR-34a inhibits HK1 expression and induces apoptosis of lens epithelial cells. And it is also proposed that miR-34a could induce the occurrence of cataract in animal experiments. This study is expected to elucidate the mechanism of miR-34a-HK1 in age-related cataract and provide a new target for the treatment of cataract.

白内障是重要的致盲性眼病，其发病机制不清。能量代谢是生物体的基本代谢活动，是维持晶状体透明的必要条件。晶状体处于相对缺氧的环境，主要通过糖酵解获得能量，而参与糖酵解的限速酶主要是己糖激酶1（Hexokinase 1， HK1），当其功能受到抑制将导致细胞能量供应不足，引起细胞病理生理的改变。miRNA参与基因转录后的调控，预实验发现年龄相关性白内障晶状体上皮细胞中miR-34a的表达量较透明晶状体明显升高，同时miR-34a可以诱导晶状体上皮细胞发生凋亡。但目前尚无关于miR-34a-HK1在年龄相关性白内障发生机制的研究。本研究将miR-34a作为靶点，拟在细胞水平证明miR-34a通过下调HK1的表达，引起晶状体上皮细胞凋亡；同时拟在动物实验中证明miR-34a可以诱导白内障的发生。该研究旨在阐明miR-34a-HK1通路在年龄相关性白内障中发生的分子机制，为白内障的防治提供新的靶点。

目前白内障的发病机制不清，阻碍相关药物开发。能量代谢是生物体的基本代谢活动，是维持晶状体透明的必要条件。本研究从表观遗传和能量代谢角度，探究其在白内障发生过程中的作用。在前期研究的基础上提出miR-34a通过调控HK1，影响晶状体上皮细胞的能量代谢，造成细胞凋亡，从而参与白内障发生的假说。首先进行细胞实验，构建HK1 3’UTR并与miR-34a共转染，证明HK1是miR-34a的靶基因。将miR-34a转染至人晶状体上皮细胞，CCK8检测发现miR-34a可以抑制细胞增殖，流式、TUNEL和活化Caspase3检测提示miR-34a促进细胞凋亡。同样将siHK1转染至人晶状体上皮细胞，也得到同样的结果。此外，qPCR和western blot显示，miR-34a和siHK1均可下调HK1的表达。为了进一步验证假说，进行了动物实验。将小鼠晶状体取出，分别与miR-34a、siHK1和脱氧葡萄糖共培养，结果显示，与对照组相比，miR-34a、siHK1和脱氧葡萄糖均加速晶状体混浊；增加小鼠晶状体上皮细胞活化caspase3表达和TUNEL阳性细胞数量；下调HK1的蛋白表达。由此证明miR-34a-HK1通路调节晶状体上皮细胞的能量代谢,在白内障形成中发挥重要作用，为今后白内障的药物研发防治提供新的依据。