摘要
目的:观察miR-214对人结肠癌细胞株LoVo中轴突导向因子4D(semaphorin 4D,Sema4D)表达水平的影响,探讨miR-214对Sema4D的调控作用.方法:设计并合成miR-214模拟物(miR-214mimics),miR-214抑制剂(miR-214 inhibitor),二者对照序列(mi-control,in-control),分别与以脂质体lipofectamine 2000包裹后转染人结肠癌细胞株LoVo.RT-PCR检测转染24 h后各细胞株内miR-214的表达及Sema4D mRNA的表达变化,并用Western blot检测Sema4D蛋白的表达变化.结果:LoVo细胞株转染miR-214 mimics、mi-control,inhibitor、in-control 24 h后miR-214的相对表达量为8.003±0.651,3.464±0.332,0.740±0.088,2.620±0.166,mimics组miR-214表达较其对照组升高,inhibitor组表达较其对照组降低(P<0.05).Sema4D mRNA的相对表达量为:0.420±0.027,0.851±0.062,1.243±0.087,0.660±0.042;Sema4D蛋白/β-actin分别为:0.163±0.037,0.550±0.038,1.137±0.112,0.457±0.046.Sema4D mRNA及蛋白mimics组表达较其对照组降低,inhibitor组较其对照组升高(P<0.05).结论:人结肠癌细胞株LoVo中,Sema4D受miR-214的调控,miR-214可负向调控Sema4D mRNA的表达水平进而影响其蛋白的表达,miR-214可能作为结肠癌治疗的新靶点.
AIM:To observe the effect of miR-214 on Sema4 D expression in human colon cancer cell line LoVo.METHODS:miR-214 mimic,miR-214 inhibitor,and two control sequences(mi-control,in-control) were designed,synthesized and transfected into LoVo cells,respectively.RT-PCR was used to detect the expression changes of miR-214 and Sema4 D mRNA in transfected cells,and the expression change of Sema4 D protein was detected by Western blot.RESULTS:The relative expression levels of miR-214 in LoVo cells transfected with miR-214 mimic,mi-control,miR-214 inhibitor,and in-control were 8.003 ± 0.651,3.464 ± 0.332,0.740 ± 0.088,and 2.620 ± 0.166,respectively.The expression of miR-214 increased significantly in the mimic group compared with the control group,but was significantly lower in the inhibitor group than in the control group(P〈0.05).The relative expression levels of Sema4 D mRNA in LoVo cells transfected with miR-214 mimic,micontrol,miR-214 inhibitor,and in-control were 0.420 ± 0.027,0.851 ± 0.062,1.243 ±0.087,and 0.660 ± 0.042,respectively,and the relative expression levels of Sema4 D protein were 0.163 ± 0.037,0.550 ± 0.038,1.137 ±0.112,and 0.457 ± 0.046,respectively.Sema4 D mRNA and protein expression in the mimic group was decreased significantly compared with the control group,but was significantly increased in the inhibitor group compared with the control group(P〈0.05).CONCLUSION:Sema4D expression is regulated by miR-214 in human colon cancer cell line LoVo,and miR-214 can down-regulate the expression of Sema4 D mRNA and thereby influence the protein expression.miR-214 could be used as a new target for treatment of colon carcinoma.
出处
《世界华人消化杂志》
CAS
2015年第7期1115-1120,共6页
World Chinese Journal of Digestology
基金
2014云南省教育厅科研基金重点资助项目
No.2014Z075~~