组蛋白H3乙酰化对脑胶质瘤中Nanog基因的调控作用

摘要

Objective To investigate the modulatory effects of acetylation of histone H3 on the proliferation-related factor Nanog in glioma. Methods U87 glioma cells were cultured in vitro and then exposed to apicidin at different concentrations and different time points as experimental group, while U87 cells exposed to the dimethylsulfoxide (DMSO) as DMSO group and not receiving any solvent as blank control group. MTT assay was used to detect the effect of apicidin on cell proliferation and select suitable intervention concentration. Real-time PCR was employed to measure the Nanog mRNA expression. Western blot was applied to detect the acetylation of histone H3. Chromatin immunoprecipitation real-time PCR was used to measure the level of the acetylation of histone H3 in promoter region of Nanog. Results Apicidin significantly inhibited the proliferation of U87 cells by MTT assay, and the half maximal inhibitory concentration (IC50) was 1.74±0.13 μmol/L in the experimental group. Compared with the blank control group, the Nanog mRNA expression of U87 glioma cells decreased by 46.52%±0.53% in the experimental group (P<0.05), the expressions of acetylation of histone H3 and Nanog protein in U87 glioma cells also decreased obviously (P<0.05), and the acetylation of histone H3 expression in the promoter region of Nanog decreased by 46.52%±0.82%(P<0.05). Conclusions Low level of the acetylation of histone H3 can inhibit Nanog expression, and Nanog expression is regulated by the acetylation of histone H3 in glioma cell line.%目的：探讨组蛋白H3乙酰化修饰对脑胶质瘤增殖相关标记因子Nanog的调控作用。方法体外培养胶质瘤U87细胞,用不同浓度的Apicidin在不同时间内干预U87细胞作为实验组,另设加入DMSO溶剂的DMSO组,及不加药的空白对照组。MTT增殖实验观察Apicidin对细胞增殖的影响,并选出合适的干预浓度。Real-time PCR检测Nanog mRNA表达水平变化,Western blot检测组蛋白H3乙酰化及Nanog蛋白水平,染色质免疫共沉淀(ChIP) Real-time PCR技术检测Nanog启动子区域组蛋白H3乙酰化水平。结果 MTT结果显示：实验组细胞生长出现显著抑制,48 h内细胞增殖的半数抑制浓度IC50为(1.74±0.13)μmol/L。与空白对照组比较,实验组脑胶质瘤U87细胞中Nanog mRNA表达降低46.52%±0.53%(P＜0.05),H3乙酰化水平和Nanog蛋白也均明显降低(P＜0.05),实验组Nanog启动子区组蛋白H3乙酰化水平降低46.52%±0.82%(P＜0.05)。结论在脑胶质瘤细胞系中,组蛋白H3低乙酰化水平抑制Nanog表达,Nanog受组蛋白H3乙酰化修饰的调控。