缺氧、氧化应激对视网膜色素上皮细胞增殖及葡萄糖调节蛋白78表达的影响

发布时间：2019-03-08 14:12

【摘要】：目的观察缺氧、氧化应激对体外培养的视网膜色素上皮(retinal pigment epithelium,RPE)细胞增殖的影响及GRP78表达的变化。方法体外培养人RPE细胞,采用CoCl2诱发细胞缺氧模型、H2O2诱发氧化应激模型。实验分为量-效关系组:细胞分别经浓度为100μmol·L-1、200μmol·L-1、400μmol·L-1、800μmol·L-1的CoCl2、H2O2作用12h,用MTT法检测细胞增殖情况;时-效关系组:在CoCl2浓度为200μmol·L-1、H2O2浓度为400μmol·L-1的基础上,分别选取药物作用8h、12h、16h3个时间点收集细胞,采用免疫细胞化学染色检测GRP78蛋白的表达情况。结果量-效关系组:不同浓度CoCl2组RPE细胞活性与空白对照组相比均明显下降(均为P0.05);400μmol·L-1及800μmol·L-1 H2O2组细胞存活率较空白对照组均明显下降(均为P0.05)。时-效关系组:空白对照组GRP78表达阳性细胞率为(5.4±3.0)%;CoCl2作用8h、12h、16h GRP78阳性细胞率分别为(34.2±7.1)%、(54.4±9.1)%、(34.7±6.8)%;H2O2作用8h、12h、16hGRP78阳性细胞率分别为(37.8±7.1)%、(57.4±5.1)%、(42.5±3.9)%。经统计学分析,CoCl2、H2O2处理8h后,RPE细胞中GRP78表达均较空白对照组明显升高(均为P0.01),作用12h时GRP78的表达量最高,作用16h时GRP78的表达量又有下降。结论 GRP78可以作为RPE细胞缺氧、氧化应激的生化标志物,GRP78的表达变化与细胞应激状态有一定依赖关系。
[Abstract]:Aim to observe the effects of hypoxia and oxidative stress on the proliferation and expression of GRP78 in cultured retinal pigment epithelium (retinal pigment epithelium,RPE) cells. Methods Human RPE cells were cultured in vitro. CoCl2-induced hypoxia model and H2O2-induced oxidative stress model were used. The cells were treated with 100 渭 mol 路L-1 200 渭 mol 路L-1 400 渭 mol 路L-1 200 渭 mol 路L-1 800 渭 mol 路L-1 CoCl2,H2O2 for 12 h, and the cell proliferation was detected by MTT assay. Time-effect relationship group: on the basis of 200 渭 mol 路L-1 CoCl2 and 400 渭 mol 路L-1 H2O2, the cells were collected at 8 h, 12 h, 16 h, respectively, and the expression of GRP78 protein was detected by immunocytochemical staining. Fruit dose-response relationship group: the activity of RPE cells in different concentrations of CoCl2 group was significantly lower than that in blank control group (all P0.05). The survival rate of cells in 400 渭 mol 路L-1 and 800 渭 mol 路L-1 H2O2 groups was significantly lower than that in the control group (P 0.05). The positive rate of GRP78 expression was (5.4 卤3.0)% in the blank control group, and (34.2 卤7.1)%, (54.4 卤9.1)%, (34.7 卤6.8)% in the GRP78 positive cells at 8 h, 12 h and 16 h after CoCl2 treatment, respectively. The positive rate of GRP78 cells was (37.8 卤7.1)%, (57.4 卤5.1)%, (42.5 卤3.9)% at 8 h, 12 h and 16 h after H2O2 treatment, respectively. After treatment with CoCl2,H2O2 for 8 h, the expression of GRP78 in RPE cells was significantly higher than that in the control group (P0.01), the expression of GRP78 was the highest at 12 h and decreased at 16 h. Conclusion GRP78 can be used as a biochemical marker of hypoxia and oxidative stress in RPE cells. The change of GRP78 expression is dependent on the state of cell stress.
【作者单位】：河南省立眼科医院;河南省眼科研究所;吉林大学第二医院眼科;
【分类号】：R774.1

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