三氧化二砷联合西罗莫司体外抑制人肾癌细胞786-O增殖及对PI3K/Akt/mTOR通路的影响

Arsenic trioxide combined with sirolimus inhibit human reanl cell carcinoma 786-O cell line proliferation and the effect of PI3K/Akt/mTOR pathway

目的观察三氧化二砷(As_2O_3)联合西罗莫司(sirolimus)对786-O人肾透明细胞癌细胞增殖的影响,并通过其对PI3K/Akt/mTOR蛋白表达的影响探讨促凋亡机制。方法体外培养786-O细胞,电镜观察药物对细胞形态的影响;MTT法检测单药作用及联合用药的细胞增殖抑制率;流式细胞法分析药物诱导786-O的细胞凋亡率;Western blot分析药物对PI3K、Akt、mTOR蛋白表达的影响。结果电镜结果可见786-O细胞发生细胞凋亡的形态学改变,MTT法显示As_2O_3和sirolimus均有抑制增殖作用,联合用药效果好于单独用药;PI/Annexin V双染分析表明sirolimus 3μmol/L、A_2O_3 3μmol/L、A_2O_3 3μmol/L+sirolimus 3μmol/L的凋亡率分别为7.17%、14.49%、20.34%;Western blot法显示A_2O_3联合sirolimus可同时下调PI3K、Akt、mTOR蛋白表达,效果显著优于单独用药。结论 A2O3联合sirolimus可明显抑制人肾癌786-O细胞增殖,并促进其凋亡,其机制与同时调控PI3K、Akt、mTOR蛋白表达有关。

Objective To investigate the effect of arsenic trioxide （As2o3） combined with sirolimus on the proliferation of 786-O human renal clear cell carcinoma cells and to explore the mechanism of pro-apoptotie mechanism through its effect on P13K/ AkffmTOR protein expression. Methods 786-O cells were cultured in vitro and the effects of the drugs on the morphology of the cells were observed by electron microscopy; MTT assay was used to detect the cell proliferation inhibition rate of monotherapy and combination therapy; flow eytometry was used to analyze the apoptotie rate of 786-O cells; Western blot analysis of PI3K, Akt, mTOR protein expression. Results The morphological changes of apoptosis in 786-Ocells were observed by electron microscopy. As203 and sirolimus were inhibited by MTT, and the combined effect was better than that of single drug; PI/Annexin V double staining analysis showed that the apoptotic rates of sirolimus 3 p.mol/L, A2O3 3 p.mol/L, A2O3 3 μmol/L＋sirolimus 3 μmol/L were 7.17%, 14.49% and 20.34%. Conclusions A2O3 combined with sirolimus can significantly inhibit the proliferation of human renal cell carcinoma 786-0 cells and promote its apoptosis, its mechanism and regulation of PI3K, Akt, mTOR protein expression.