miR-150和HMGA2在结直肠癌中的表达和临床意义

Expression of miR-150 and HMGA2 in colorectal cancer cells and their relation with cell proliferation

目的探讨miRNA-150(miR-150)和高迁移率蛋白A2(HMGA2)在结直肠癌细胞增殖周期中的作用。方法采用RT-PCR法检测人结直肠癌SW480细胞与正常结肠上皮细胞(FHC)的miR-150、HMGA2和周期蛋白A(Cyclin A)表达水平;并将SW480细胞分成miR-150对照组、miR-150模拟体组、小干扰对照组、小干扰HMGA2组和miR-150模拟体联合小干扰HMGA2组共5组,通过荧光素酶报告系统检测miR-150对照组和miR-150模拟体组人胚肾细胞的荧光活性;采用流式细胞术测定5组转染细胞G0/G1、G2/M、S期细胞比例;并采用细胞计数试剂盒8(CCK-8)法检测5组转染细胞细胞活力,用吸光度(OD)表示。结果SW480细胞组较FHC组miR-150表达水平下降,而HMGA2和Cyclin A表达水平升高(均P<0.05);与miR-150对照组比较,miR-150模拟体组pLUC-HMGA2-3′-UTR-野生型人胚肾细胞荧光活性明显为低(P<0.05);与miR-150对照组比较,miR-150模拟体组、小干扰HMGA2组和miR-150模拟体联合小干扰HMGA2组G0/G1期细胞比例均明显为高,而G2/M、S期细胞比例均明显为低(均P<0.05);与miR-150对照组及小干扰对照组比较,miR-150模拟体组、小干扰HMGA2组、miR-150模拟体组联合小干扰HMGA2组的OD值均明显下降(均P<0.05)。结论miR-150通过靶向抑制HMGA2进一步下调Cyclin A的表达从而阻断结直肠癌细胞增殖周期,抑制其增殖。

Objective To investigate the expression of micro RNA-150(miR-150)and high mobility group protein A2(HMGA2)in human colorectal cancer cells and their relation with cell proliferation.Methods The expression levels of miR-150,HMGA2,and Cyclin A in human colorectal cancer cell line SW480 and normal colorectal epithelial cell line(FHC)were detected with RT-PCR.SW480 cells were divided into five groups:miR-150 control,miR-150 mimic,small RNA interfere normal control(si-NC),si-HMGA2,and miR-150 mimic+si-HMGA2.The effect of miR-150 control and miR-150 mimic on HMGA2 expression was detected by luciferase reporting system.Flow cytometry was used to determine the cell ratios at G0/G1,G2/M and S stages.Cell proliferation was detected by using cell counting kit 8(CCK-8)test in terms of absorbance value(OD).Results HMGA2 and Cyclin A levels were higher significantly,whereas miR-150 expression was lower significantly in SW480 cells compared to those in FHC cells(P<0.05).Compared with the miR-150 control,miR-150 mimic transfection markedly reduced the relative luciferase activity in HEK293 T cells transfected with HMGA2-wt(P<0.05).On the contrary,it exhibited no significant impact on the relative luciferase activity in HEK293 T cells transfected with HMGA2-mut.Compared with the miR-150 control,the cell ratio of G0/G1 phase was higher in miR-150 mimic,si-HMGA2 and miR-150 mimic+si-HMGA2,and the cell ratio of G2/M/S phase was lower(P<0.05).Compared with the miR-150 control and si-NC,the cell proliferation was attenuated in miR-150 mimic,si-HMGA2 and miR-150 mimic+si-HMGA2 groups.Conclusion Mi R-150 can down-regulate cyclin A expression to block cell cycle and inhibit cell proliferation through targeted inhibiting HMGA2 in colorectal cancer cells.