反义DLC-1基因对结直肠癌LoVo细胞增殖和细胞周期的影响  

Inhibition of DLC-1 gene expression by RNA interference (RNAi) in colon cancer LoVo cell line

作　　者：金月玲[1] 商延芳[1] 田小强[1] 方媛[1] 黄培林[1] 

机构地区：[1]东南大学医学院病理学与病理生理学系,南京210009

出　　处：《江苏医药》2008年第5期493-496,共4页Jiangsu Medical Journal

基　　金：国家自然科学基金资助(30471937)

摘　　要：目的应用核糖核酸干扰(RNAi)技术,通过对大肠癌细胞株LoVo中DLC-1基因mRNA表达的抑制,初步探讨DLC-1基因对大肠癌细胞株的生物学行为影响。方法构建DCL-1的RNAi重组体pGCsiDLC,转染大肠癌LoVo细胞株。采用RT-PCR观察转染前后细胞株中DLC-1 mRNA以及Bax、Bcl-2基因表达的改变,MTT比色法检测转染前后细胞增殖,流式细胞仪检测细胞周期。结果成功构建发卡siRNA真核表达载体pGCsiDLC;pGCsiDLC转染LoVo细胞后,电泳显示DLC-1 mRNA表达水平明显下降;LoVo细胞生长曲线增长幅度亦随时间的增加而逐渐增高,在48h最为明显,RNAi组细胞比脂质体组及空白对照组细胞增殖明显增高(P<0.05);干扰后的LoVo细胞株G0/G1期发生阻滞,而G2/M期细胞数量分布减少。而Bax和bcl-2基因表达在干扰前后未见明显改变。结论成功构建载体介导的DLC-1靶向RNAi重组体可有效抑制LoVo细胞DLC-1 mRNA表达;DLC-1基因可抑制结直肠癌细胞增殖并且对细胞周期重新分布,其可能与结直肠癌的发生发展有一定关系,可能是一个新的抑癌基因。Objective To investigate the affection of DLC-1 gene expression on biological behavior in colon cancer LoVo cells. Methods RNAi lipo-recombinent of DLC-1 gene（pGCsiDLC）was constructed and transduced into LoVo cell.RT-PCR weas used to detect DLC-1 mRNA level and Bax,Bcl-2 gene expression,MTT assay to measure cell proliferation and flow cytometry to observe cell cycle. Results DCL-1 transfection vector-pGCsiDLC was successfully constructed. After transfected with pGCsiDLC,the expression of DCL-1mRNA in LoVo cells was obviously decreased with an increased cell proliferation observably （P〈0.05）. Flow cytometry in cell cycle detection indicated that interfered LoVo cells in G2/M phase decreased obviously and the cells in G0/G1 phase increased markedly. There was no obvious change in Bax,Bcl-2 gene expression.Conclusion RNAi recombinent of DLC-1 gene could effectively inhibit the expression of DLC-1 mRNA in LoVo cells. DLC-1 gene may inhibit LoVo cell proliferation by changing cell cycle distribution. DLC-1 may be a new cancer-suppressing gene and play an important role in colon cancer mechanism.

关 键 词：核糖核酸干扰 DLC-1基因 LOVO细胞株 结直肠癌 

分 类 号：Q751[生物学—分子生物学]