三氧化二砷对肾癌GRC-1细胞凋亡及p53、Survivin基因表达的影响

Effects of arsenic trioxide on expressions of p53 and survivin in human renal cell carcinoma GRC-1 cell line

目的观察三氧化二砷(As_2O_3)对人肾癌GRC-1细胞凋亡及p53、Survivin基因表达的影响,探讨其作用机制。方法将体外培养的人肾癌GRC-1细胞分为对照组(N)和实验组,实验组根据加入As_2O_3浓度的不同分为A、B、C、D、E(2、4、6、8、10μmol/L)组。48h后,四甲基偶氮唑蓝(MTT)法测定细胞增殖,流式细胞仪检测细胞凋亡和细胞周期变化．免疫细胞化学和RT-PCR方法测定p53和Survivin基因表达。结果As_2O_3可抑制GRC-1细胞的增殖．当As_2O_3由2μmol/L增加到10μmol/L时,细胞增殖率由88．3%降低到18．7%(F= 7546．587,P＜0．01)。各实验组p53、Survivin基因表达均较对照组减少,且随着As_2O_3的增加表达呈递减改变(F=1120．741、F=6 296．535,P＜0．01)。结论As_2O_3能够抑制GRC-1细胞增殖,并且具有浓度依赖性。能将细胞阻滞在G_2/M期,诱导细胞凋亡,其作用机制与p53和Survivin基因表达水平下降有关。

Objective To observe the effects of arsenic trioxide （As2O3） on p53 and survivin gene expression in human renal cell carcinoma（RCC） GRC-1 cell line in vitro and to elucidate its mechanisms. Methods RCC GRC-1 cells were divided into 2 groups： the experimental group and the control group in vitro. The experimental group was treated with As2O3 in different concentration from A to E, namely 2, 4, 6, 8 and 10 μmoL/L. 48 h later, the anti-proliferation effect of As2O3 on RCC GRC-1 cells were measured by MTY assay, Flow cytometry（FCM） was performed to evaluate the influences of As2O3 on cell cycle and cell apoptosis. The expressions of p53 and survivin were measured by immunocytochemistry staining and RT-PCR respectively. Results As203 inhibited the proliferation of GRC-1 cells, the proliferative rate was decreased from 88.3% to 18.7%, along with the dose of As2O3 increasing from 2 μmoL/L to 10 μmoL/L （F = 7 546.587,P〈0.01 ）. The expressions of p53 and survivin gene in treated group were declined compared with those of control group respectively, in a concentrationdependent manner（F = 1 120.741 ,F = 6 296.535,P 〈 0.01）. Conclusions As2O3 inhibits the proliferation of GRC-1 cells in a concentration-dependent manner, and induces the G2/M phase arrest and apoptosis of GRC-1 cells in vitro through a mechanisms of down-regulation of expressions of p53 and survivin gene.