雷帕霉素对高糖诱导人晶状体上皮细胞氧化应激的保护作用

摘要

目的 探讨雷帕霉素对高糖诱导的人晶状体上皮细胞(human lens epithelial cells,HLECs)氧化应激的影响.方法 建立高糖诱导的HLECs氧化损伤模型,用不同浓度(10 nmol·L-1、100 nmol·L-1)雷帕霉素干预,CCK-8比色法检测细胞存活率,倒置显微镜观察细胞形态并拍照,H2DCFDA检测细胞内活性氧水平,Western Blotting检测各组凋亡相关蛋白Bcl-2及Bax的表达情况.结果 CCK-8检测结果显示,用10 nmol·L-1、100 nmol·L-1雷帕霉素处理后,HLECs存活率分别提高到62.86％±3.45％和77.47％±4.21％,与高糖组(42.32％±3.10％)比较差异均有统计学意义(均为P＜0.05);H2 DCFDA荧光探针染色结果显示,雷帕霉素处理后HLECs内活性氧生成量下降,氧化损伤细胞减少;此外,雷帕霉素可以抑制高糖诱导的HLECs内凋亡蛋白Bax、Bcl-2的表达,抑制效应与剂量正相关.结论 雷帕霉素可以抑制高糖诱导的HLECs氧化损伤及凋亡的发生,对糖尿病性白内障具有预防作用.%Objective To investigate the effects of rapamycin on human lens epithelial cells (HLECs) against high glucose-induced oxidative stress.Methods A oxidative damage model of HLECs induced by high glucose was established,and intervention with different concentrations of rapamycin for the cells was performed for detecting the survival rate by CCK-8 assay,observing cell morphology by inverted microscope,and measuring the level of intracellular reactive oxygen species (ROS) by H2DCFDA staining,as well as determining the expression of apoptosis related Bcl-2 and Bax proteins by Western blot in all groups.Results CCK-8 results showed that the survival rate of HLECs was increased to 62.00％ ± 3.74％ and 79.57％ ± 5.26％ after treated with 10 nmol · L-1 and 100 nmol · L 1 rapamycin,and the difference was statistically significant when compared with the high glucose group (42.32％ ± 3.10％) (all P ＜ 0.05);H2DCFDA fluorescent probe staining demonstrated that rapamycin could suppress ROS generation in HLECs and alleviate oxidative damage to cells;besides,rapamycin could downregulate the expression of Bcl-2 and Bax protein in HLECs and the inhibitory effects were positively correlated with the concentration of rapamycin.Conclusion Rapamycin can inhibit high glucose-induced oxidative damage and apoptosis m human HLECs,which provides a prevention effect for diabetic cataract.