miR-27a-3p调控Ras/Raf/MEK/ERK信号通路抑制肾透明细胞癌细胞增殖的研究

Study on miR-27a-3p in inhibiting proliferation of clear cell renal cell carcinoma by regulating Ras/Raf/MEK/ERK signaling pathway

目的探讨miR-27a-3p对肾透明细胞癌细胞Ras/Raf/MEK/ERK信号通路的作用,观察其对肾透明细胞癌细胞增殖的影响。方法使用脂质体试剂lipofectamine 2000将miR-27a-3p模拟物或miR-NC转染至肾透明细胞癌细胞株Caki-1和A-498;qRT-PCR和Western blot检测miR-27a-3p的靶基因KRAS及下游通路蛋白的表达;流式细胞仪检测细胞周期;噻唑蓝(MTT)检测细胞活力;集落形成实验检测细胞增殖能力。结果 KRAS基因和下游通路蛋白的表达降低;转染miR-27a-3p后的肾透明细胞癌细胞周期明显被抑制,G0/G1期的细胞比例上升(P<0.01);肾透明细胞癌细胞活力和增殖能力降低(P<0.05)。结论过表达miR-27a-3p在体外可以显著降低肾透明细胞癌细胞Ras/Raf/MEK/ERK蛋白的表达,进而显著抑制其增殖能力。

Objective To investigate the effect of miR-27a-3p on Ras/Raf/MEK/ERK signaling pathway in clear cell renal cell carcinoma and to observe the effect of miR-27a-3p on the proliferation of clear cell renal cell carcinoma.MethodsThe miR-27a-3p mimics or miR NC was transfected into the clear cell renal cell carcinoma cell lines Caki1and A498using lipofectamine2000.The expression of KRAS and downstream pathway protein were detected by qRT PCR and Western blot.Cell cycle was detected by flow cytometry.Cell viability was detected by MTT assay.Cell proliferation was measured by colony formation assay.ResultsThe expression of KRAS gene and downstream pathway protein expression decreased.The cell cycle of clear cell renal cell carcinoma was significantly inhibited after transfection with miR-27a-3p,and the percentage of cells in G0/G1phase increased(P<0.01).The viability and proliferation abilities of clear cell renal cell carcinma decreased(P<0.05).ConclusionsOverexpression of miR-27a-3p significantly reduced the expression of Ras/Raf/MEK/ERK protein in clear cell renal cell carcinoma in vitro,and significantly inhibited its proliferation abilities.