α-硫辛酸对高糖条件下培养的人晶状体上皮细胞的保护作用

目的　探讨α-硫辛酸（alpha-lipoic acid，ALA）对高糖条件下培养的人晶状体上皮细胞系HLEC-B3的保护作用。方法　将传代培养的HLEC-B3细胞分为正常对照组、高糖组、ALA组。正常对照组不做任何处理，ALA组HLEC-B3经过不同浓度ALA（25 μmol·L^-1、50 μmol·L^-1、100 μmol·L^-1）预处理1 h后，与高糖组一同置于高糖条件下培养48 h。经上述处理后，即刻采用MTT检测各组晶状体上皮细胞的活性，流式细胞仪检测各组细胞内线粒体活性氧（reactive oxygen species，ROS）的变化，Western blot和RT-PCR检测各组细胞内锰超氧化物歧化酶（manganese superoxide dismutase，MnSOD）的表达。结果　高糖组HLEC-B3细胞活性为53.60%±4.10%，较正常对照组显著降低，差异有统计学意义（P<0.01），25 μmol·L^-1、50 μmol·L^-1、100 μmol·L^-1 ALA组细胞活性分别为65.30%±3.70%、72.70%±4.90%、83.40%±3.60%，均较高糖组显著增高（均为P<0.05）；流式细胞仪检测结果显示，高糖组细胞内ROS水平较正常对照组明显增高（P<0.01），各浓度ALA组ROS水平分别降低为14.70%±0.70%、8.70%±0.87%、5.20%±0.53%，差异均有统计学意义（均为P<0.01）。RT-PCR及Western blot检测结果显示，高糖培养后细胞内MnSOD mRNA和蛋白相对表达量均较正常对照组显著降低（均为P<0.01）；与高糖组相比，不同浓度ALA组细胞内MnSOD mRNA相对表达量分别增高为0.63±0.06、0.71±0.06、0.84±0.04；MnSOD蛋白相对表达量分别增高为0.25±0.03、0.31±0.02、0.45±0.04，差异均有统计学意义（均为P<0.05）。ALA对高糖诱导的HLEC-B3细胞损伤的保护作用呈一定的浓度依赖性。结论　ALA对高糖条件下培养的HLEC-B3具有一定的保护作用，且该保护作用可能是通过提高细胞内MnSOD的表达水平实现的。

Neuroprotective effects of alpha-lipoic acid on human lens epithelial cells cultured in the condition of high glucose in vitro

Objective　To investigate whether alpha-lipoic acid （ALA） possesses neuroprotective effects against high glucose-induced damage in vitro.Methods　The cultured human lens epithelial cells （HLEC-B3 cells） were divided into normal control group，high glucose group and ALA group.Normal control group left untreated，ALA group was pretreated with different concentrations of ALA （25 μmol·L^-1，50 μmol·L^-1，100 μmol·L^-1） for 1h，and then ALA group and high glucose group were cultured under high glucose conditions for 48h.After above treatment，the activity of lens epithelial cells in each group was detected immediately by MTT，and the changes in intracellular reactive oxygen species （ROS） were detected by flow cytometry.In addition，the expression of manganese superoxide dismutase （MnSOD） was detected by Western blot and RT-PCR in all groups.Results　The activity of HLEC-B3 cells in high glucose group was 53.60%±4.10%，which was significantly lower than that in normal control group （P<0.01）.The cell viability in 25 μmol·L^-1，50 μmol·L^-1，100 μmol·L^-1 ALA group was 65.30%±3.70%，72.70%±4.90% and 83.40%±3.60%，respectively，all which were higher than those in the high glucose group （all P<0.05）.Flow cytometry showed that intracellular ROS level in the high glucose group was significantly higher than that in the normal control group （P<0.01），but ROS levels were decreased to 14.70%±0.70%，8.70%±0.87%，5.20%±0.53% after 25 μmol·L^-1，50 μmol·L^-1，100 μmol·L^-1 ALA treatment，respectively，with significant difference （all P<0.01）.RT-PCR and Western blot results indicated that the relative expression of MnSOD mRNA and protein in the high glucose group was significantly lower than that in the normal control group （all P<0.01）.Compared with the high glucose group，MnSOD mRNA and protein relative expression levels were significantly increased to 0.63±0.06，0.71±0.06，0.84±0.04，and 0.25±0.03，0.31±0.02，0.45±0.04，respectively （all P<0.05）.In addition，the protective effects of ALA （25-100 μmol·L^-1） on HLEC-B3 cells induced by high glucose was in a dose-dependent manner.Conclusion　ALA has a protective effect on human lens epithelial cell line HLEC-B3 cultured in high glucose condition，and this protective effect may be achieved by increasing the expression level of intracellular MnSOD.