小干扰RNA沉默表皮生长因子受体对肾癌细胞增殖、迁移及侵袭能力的影响

Effects of small interfering RNA targeting epidermal growth factor receptor on proliferation, migration and invasion of renal cell carcinoma line renal cancer cell line

目的观察小干扰RNA（siRNA）沉默表皮生长因子受体（EGFR）对人肾癌细胞株（ACHN）细胞增殖、迁移及侵袭能力的影响。方法化学合成针对EGFR的siRNA，实验分3组，通过脂质体转染法转染进ACHN肾癌细胞中，利用Westernblot技术检测细胞中EGFR蛋白的敲除效率，通过锥虫蓝拒染法测定细胞生长曲线，用噻唑蓝（MTY）比色分析法、细胞划痕实验、Transwell分别检测细胞的增殖、迁移及侵袭能力。结果特异性siRNA可明显抑制EGFR的表达；生长曲线提示，RNA干扰（RNAi）组细胞在24、48、72、96h增殖抑制率分别为15．6％、38．4％、64．1％、68．0％，较空白对照组及阴性对照组生长明显减慢（P〈0．05）；24h后RNAi组划痕宽度为（109．97±7．50）μm，空白对照组及阴性对照组的划痕基本愈合（P〈0．05）；RNAi组穿膜细胞数[（153±13）个]，分别与空白对照组[（320±20）个]、阴性对照组[（300±16）个]比较，穿膜细胞明显减少（P〈0．05）。结论siRNA抑制EGFR表达后，肾癌细胞的增殖、迁移及侵袭能力显著降低。

Objective To investigate the effects of small interfering RNA （siRNA） silencing epidermal growth factor receptor （EGFR） gene on the ability of proliferation, migration and invasion of the renal cell carcinoma cell line renal cancer cell line （ACHN）. Methods ACHN cells were transfected with siRNA targeting human EGFR by liposome transfection reagent, and three experimental groups were set up. The expression level of EGFR was detected by using Western blotting. The cell growth curves were determined by trepan blue staining. The proliferation inhibition of ACHN cells was measured by methyl thiazolyl tetrazolium （MTF） assay. The migration of ACHN cells was detected by using wound healing assay, and the ability of invasion was examined by Transwell assay. Results The expression of EGFR gene could be suppressed obviously by sequence-specific shRNA targeting EGFR gene. The growth curve revealed the cell proliferation inhibition rate in RNAi group was 15.6%, 38.4%, 64. 1%, and 68.0% at 24, 48, 72, and 96 h respectively, which was significantly reduced as compared with the blank control group and negative group （P 〈0. 05）. In RNAi group cell scratch was （ 109. 97±7.5）μm after 24 h, and cell scratches in blank control group and negative group were basically healed （ P 〈 0. 05）. The number of migarating cells in RNAi group was （ 153±13 ）, which was significantly reduced as compared with the blank control group [ （320±20） ] and negative control group [ （300±16 ） ] （ P 〈 0. 05 ）. Conclusion siRNA targeting human EGFR could specially suppress the expression of EGFR gene in renal cell carcinoma cells, which could significantly reduce the capability of proliferation, migration and invasion of the renal cell carcinoma cells.