银杏内酯B对H_2O_2诱导人视网膜色素上皮细胞氧化损伤的保护作用

Protective effects of ginkgolide B on H_2O_2-induced oxidative damage in RPE cells

目的探讨银杏内酯B对体外培养的人视网膜色素上皮(retinal pigment epithelial,RPE)细胞氧化损伤的保护作用及可能机制。方法 RPE细胞传代培养24 h后,随机分为阴性对照组、氧化损伤组、银杏内酯B低浓度组(1 mol·L^(-1))和银杏内酯B高浓度组(10 mol·L^(-1)),银杏内酯B预处理24 h后,加入100μmol·L^(-1)H_2O_2继续孵育12 h,MTT比色法检测细胞活力,流式细胞仪检测细胞凋亡率,Hochest33258染色观察凋亡细胞形态,比色法检测凋亡相关因子Caspase-3及Caspase-9的表达。结果银杏内酯B能明显抑制H_2O_2诱导的RPE细胞活力的下降,MTT结果显示银杏内酯B低浓度组和银杏内酯B高浓度组细胞活性分别为(53.37±2.53)%和(69.57±3.17)%,与氧化损伤组的(31.33±2.41)%比较,差异均具有统计学意义(均为P<0.05);流式细胞计数结果显示银杏内酯B低浓度组和银杏内酯B高浓度组细胞凋亡率分别下降至(27.53±3.34)%和(13.30±2.25)%,与氧化损伤组的(48.13±2.68)%比较,差异均具有统计学意义(均为P<0.01)。此外,银杏内酯B还可以减少H_2O_2所致RPE细胞内Caspase-3及Caspase-9的表达。结论银杏内酯B通过抑制Caspase-3及Caspase-9的表达有效抑制了H_2O_2对RPE细胞的损伤,从而为其用于治疗RPE细胞损伤提供可靠的实验依据。

Objective To discuss the protective effects and possible mechanisms of ginkgolide B on oxidative damage of human retinal pigment epithelial （ RPE ） cells cultured in vitro. Methods RPE cells were cultured and passaged for 24 hours, and randomly divided into four groups： Negative control group, oxidative damage group, ginkgolide B low dose （1 mol·L^-1） group and ginkgolide B high dose （ 10 mol·L^-1） group,pre-treated with ginkgolide B for 24 hours, and then treated with 100 μmol·L^-1 HzO2 for 12 hours, cell viability were tested by MTT colorimetric detection, apoptotic rate was detected by flow cytometry, apoptotic cell morphology was observed by hochest33258 staining, expression of apoptosis-related factors Caspase-3 and Caspase-9 were tested by colorimetric detection. Results Ginkgolide B could effective inhibit H2O2-induced cells viability decrease in RPE cells, MTT assay results showed that cell viability in ginkgolide B low dose group and ginkgolide B high dose group were（ 53.37 ± 2.53 ） % and（ 69.57 ± 3. 17 ） % , there were statistically significant differences compared with oxidative damage group （ 31.33 ± 2.41 ） % （ all P 〈 0.05 ） ; Flow cell count results showed that cell apoptotic rates in ginkgolide B low dose group and ginkgolide B high dose group decreased to （ 27.53 ± 3.34 ） % and （ 13.30± 2.25 ） % , there were statistically significant differences compared with oxidative damage group （48.13 ± 2.68 ） % （all P 〈 0. 01 ）. Besides, ginkgolide B could decrease the expression of Caspase-3 and Caspase-9 in H2O2-induced oxidative damage of RPE cells. Conclusion Ginkgolide B can effectively inhibit H2O2-induced RPE cells damage by inhibiting the expression of Caspase-3 and Caspase-9, which provide reliable experimental basis for the treatment of injuries in RPE cells.