辛伐他汀抑制缺氧条件下人视网膜色素上皮细胞HIF-1α和VEGF的表达水平

Simvastatin inhibits HIF-1α and VEGF expression in RPE cells under hypoxia conditions

摘要:

目的：探讨缺氧条件下辛伐他汀(Sim)对体外培养的人视网膜色素上皮细胞(RPE-19)的影响及其可能机制。

方法：将体外培养的人RPE-19细胞随机分为三组：对照组、缺氧组(培养基中CoCl₂的最终浓度为125μmol/L)和Sim处理组(在含有125μmol/L CoCl₂的RPE-19细胞培养基中加入3μmol/L Sim)。24h后，观察RPE-19细胞的形态，用MTT法检测细胞增殖，采用酶联免疫吸附试验(ELISA)和免疫印迹法检测缺氧诱导因子1-α(HIF-1α)和血管内皮生长因子(VEGF)的分泌水平和蛋白表达，采用免疫印迹法检测自噬蛋白的表达水平，TUNEL法检测细胞凋亡。

结果：缺氧条件下RPE-19细胞的形态发生了明显变化。缺氧组RPE-19细胞中HIF-1α和VEGF的蛋白表达明显增加，Sim治疗后显著降低。Beclin1和LC3B蛋白在CoCl₂+Sim组中的表达水平有所下降，且表达水平显著低于对照组和CoCl₂组。缺氧条件下，Sim抑制了RPE-19细胞增殖，促进了细胞的凋亡。

结论：Sim可抑制缺氧条件下RPE-19细胞HIF-1α和VEGF蛋白的表达，抑制细胞增殖，促进凋亡，Sim促进RPE-19细胞凋亡的机制可能与其抑制自噬有关。

Abstract:

AIM: To investigate the effects of simvastatin(Sim)on human retinal pigment epithelial cells(RPE-19)and the possible mechanisms in vitro under hypoxia.

METHODS: RPE-19 cells were divided into three group: control group, hypoxia group(the final concentration of CoCl₂ in the medium was 125 μmol/L), and Sim treatment group(3 μmol/L Sim was added in the RPE cells' medium which contain 125 μmol/L CoCl₂). After 24h, the morphology of RPE-19 cells were observed, the proliferation of cells were calculated by MTT, the secretion levels and protein expression of hypoxia-inducible factor 1-Alpha(HIF-1α)and vascular endothelial growth factor(VEGF)were detected by enzyme-linked immunosorbent assay(ELISA)and Western blotting. The expression level of autophagy protein was detected by Western blot and apoptosis was detected by TUNEL.

RESULTS: The morphology and activity of RPE-19 cells showed an apparent change under hypoxia. The expression of HIF-1α and VEGF protein were increased obviously in the hypoxia group and then significantly decreased after Sim treatment. Beclin1, and LC3B proteins were decreased in the CoCl₂+Sim group, and the expression levels were lower than the control and CoCl₂ group. Under hypoxia, Sim inhibited RPE cells' proliferation and promoted the apoptosis.

CONCLUSION:Sim inhibits RPE cells' proliferation, decreases HIF-1α and VEGF protein, and promotes apoptosis under hypoxia. Our results suggested that the mechanism by which Sim promoted apoptosis in RPE cells may be related to its inhibition of autophagy.