目的:探讨微小RNA-338.3p(miR-338-3p)对人结直肠癌细胞侵袭迁移能力及Smoothened(SMO)蛋白表达的影响。方法:应用实时荧光定量PCR(Real—timePCR)检测四种人结肠癌细胞系Lovo、HT-29、HCT116、SW620中miR-338—3p表达状况,并以人脐静脉内皮细胞(HUVEC)作为对照。采取脂质体转染miR-338-3p前体(pre—miR-338.3p)至人结直肠癌细胞系SW620,观察细胞转染前后miR-338—3p的表达变化.Transwell侵袭实验及划痕实验检测转染前后细胞侵袭及迁移能力的改变;以Western—blotting和RT-PCR分析sMO蛋白及mRNA表达状况。结果:检测四种结直肠癌细胞系中miR-338—3p表达水平均较正常对照细胞显著降低,差异具有统计学意义(P〈0.01);SW620细胞转染pre—miR-338—3p后,miR-338—3p表达水平升高,sMO蛋白表达下降,同时肿瘤细胞侵袭、迁移能力明显增强,与对照组相比差异具有统计学意义(P〈0.01);但SMOmRNA表达水平在转染前后差异无统计学意义(P〉0.05)。结论:MiR-338—3p可通过抑制结直肠癌细胞系中SMO蛋白表达从而抑制肿瘤细胞侵袭转移。
Objective: To investigate the effects of microRNA-338-3p(miR-338-3p) on cell invasion and migration and Smoothened protein(SMO) expression in human colorectal cancer(CRC) cells. Methods: Real-time polymerase chain reaction( RT-PCR)was employed to detect the miR-338-3p expression in Lovo, HT-29, HCT116 and SW620, the four types of human CRC cell lines, and in human umbilical vein endothelial cells (HUVEC) , the controls. Human CRC cell line,SW620,was transfected with a miR-338-3p precursor(pre-miR-338-3p) by using liposomes. This allowed determination of the changes in miR-338-3p expression, and CRC cell invasion and migration by Transwell and scratch assays following transfection. Western-blotting and semi-quantitative RT-PCR were adopted to analyze Smoothened(SMO) protein and mRNA expression. Results:Compared with control cells, all human CRC cell lines yielded a significantly attenuated expression of miR-338-3p( all P 〈 0.01 ). Transfection of 5W620 cells with pre-miR-338-3p, when compared with control cells ,was associated with augmented miR-338- 3p expression,attenuated SMO protein expression and improved capacity of invasion and migration( all P 〈 0. 01 ) , but not the expression of SMO mRNA (P 〉 0.05 ). Conclusion: MiR-338-3p suppresses CRC invasion and migration via inhibiting SMO protein expression.