miR-204-5p在氧糖剥夺/复糖复氧致SH-SY5Y细胞损伤中的作用研究

目的观察miR-204-5p在氧糖剥夺/复糖复氧(oxygen-glucose deprivation/reoxygenation, OGD/R)致人神经母细胞瘤细胞(SH-SY5Y)损伤中的作用并从炎症/凋亡途径探讨其机制。方法 SH-SY5Y细胞分为control组、OGD/R组、OGD/R+miR-204-5p mimic组、OGD/R+miR-204-5p inhibitor组、OGD/R+miR-204-5p mimic negative control组、OGD/R+miR-204-5p inhibitor negative control组。采用MTT法测定细胞增殖、流式细胞术、TUNEL染色法检测细胞凋亡、酶联免疫法检测炎症因子(IL-10、IL-1β、TNF-α、PGE2)含量、qRT-PCR检测miR-204-5p的表达、western blot检测炎症及凋亡相关蛋白(COX-2、Bcl-2、Bax)的表达。结果与对照组相比,OGD/R组细胞miR-204-5p表达显著降低,IL-1β、TNF-α、PGE2含量增多,IL-10含量减少,COX-2、Bax蛋白水平上升,Bcl-2蛋白水平下降,细胞损伤明显加重;与OGD/R组比较,miR-204-5p mimic下调COX-2、Bax蛋白表达,上调Bcl-2蛋白表达,IL-10含量增加,IL-1β、TNF-α、PGE2含量减少,细胞活力明显增加、凋亡率显著降低,细胞损伤减轻。结论 miR-204-5p对OGD/R致SH-SY5Y细胞损伤具有明显保护作用,其机制可能与减轻细胞炎症和凋亡有关。

Effect of miR-204-5p on SH-SY5Y cells Injury Induced by Oxygen-glucose Deprivation and Reoxygenation

OBJECTIVE To observe the effect of miR-204-5 p on oxygen-glucose deprivation and reoxygenation(OGD/R) injury in SH-SY5 Y cells and to explore its possible mechanism from inflammatory/apoptotic pathways. METHODS SH-SY5 Y cells were divided into control group, OGD/R group, OGD/R+miR-204-5 p mimic group, OGD/R+miR-204-5 p inhibitor group, OGD/R+miR-204-5 p mimic negative control group, OGD/R+miR-204-5 p inhibitor negative control group. Cell viability was measured by MTT assay, cell apoptosis was detected by flow cytometry and TUNEL staining. The changes of IL-10, IL-1β, TNF-α, PGE2 were accessed by enzyme-linked immunosorbent assay. RT-PCR was used to detect the expression of miR-204-5 p and Western blot was used to measure the expressions of inflammatory and apoptosis-related proteins(COX-2, Bcl-2, Bax). RESULTS Compared with control group, the expression of miR-204-5 p was decreased obviously, COX-2, Bax, IL-1β, TNF-α, PGE2 were up-regulated significantly and Bcl-2, IL-10 were down-regulated, and the cell injury was significantly aggravated in OGD/R group. Compared with the OGD/R group, miR-204-5 p mimic had enhanced cell viability and IL-10 content significantly, declined apoptosis rate and IL-1β, TNF-α, and PGE2 content. Simultaneously, reduced COX-2 and Bax protein expression, and increased Bcl-2 protein expression were detected in miR-204-5 p mimic group. CONCLUSION miR-204-5 p has a protective effect on OGD/R-induced SH-SY5 Y cell injury, its mechanism may be associated with the reduction of cellular inflammation and apoptosis.