RNA干扰敲减PIWIL1表达对人结肠癌细胞SW620增殖,粘附及侵袭能力影响  

Knocking-down of PIWIL1 by RNA Interference Impacts Proliferation,Adhesion and Invasion of Human Colon Cancer Cell Line SW620

作　　者：任超逸[1] 吴海东[1] 浦永[1] 汤石明[1] 李欣[1] 刘民[1] 汤华[1] 

机构地区：[1]天津医科大学基础医学院天津市生命科学中心实验室,天津300070

出　　处：《中国生物化学与分子生物学报》2010年第1期55-61,共7页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金项目(No.30873017);天津市自然科学基金重点项目(No.08JCZDJC23300)~~

摘　　要：PIWIL1为AGO蛋白(Argonaute proteins)PIWI亚家族的成员之一,在睾丸中特异表达.其在干细胞自我更新、RNA干扰和翻译调节中起着重要的作用.本文采用实时PCR方法检测PIWI基因家族中PIWIL1、PIWIL2、PIWIL3、PIWIL4在人结肠癌SW620细胞中mRNA表达水平,首次证实了在SW620细胞中,PIWIL1相对PIWIL2、PIWIL3、PIWIL4,其表达水平最高(P(0.05).构建表达针对PIWIL1的小发卡结构干扰RNA的重组质粒,并用Western印迹验证其达到较高的干扰效率.用脂质体将重组质粒转染入SW620细胞中,通过MTT实验、集落形成实验、细胞聚集实验及细胞侵袭转移实验,分别观察到其可抑制SW620细胞的生长、增殖、侵袭转移能力,增强了SW620细胞的粘附能力.提示PIWIL1可能在结肠癌发生、发展过程中发挥作用.PIWIL1 encodes a member of the PIWI subfamily of Argonaute proteins,which is specifically expressed in testis and plays important roles in stem cell self-renewal,RNA interference,and translational regulation in diverse organisms.In order to study the differential expression patterns of PIWI gene family,we used real-time PCR to detect the mRNA level of PIWIL1,PIWIL2,PIWIL3 and PIWIL4 in human colon cancer cell line SW620.The results showed that the PIWIL1 mRNA level was higher in SW620 cells compared to PIWIL2,PIWIL3 and PIWIL4（P﹤0.05）.Then,the plasmid which expresses hairpin siRNA for PIWIL1 gene was constructed and transfected into SW620 cells via lipofectamine 2000.The RNA interference plasmid can effectively down-regulate PIWIL1 gene expression.MTT assay,colony formation assay,aggregation experiment and transwell assay were used to study the effect of PIWL1-shRNA on cell viability,proliferation capacity,adhesion,movement and invasion in SW620 cells.In PIWIL1-depleted SW620 cells,the cell viability and proliferation were inhibited and the invasive ability was also significantly reduced.However,the experiment showed a significant increase of cell aggregation in PIWIL1-shRNA transfected cells compared with un-transfecterd cells or control vector transfected cells.These results suggest that PIWIL1 may play an important role in the pathogenesis of human colon cancer.

关 键 词：结肠癌 SW620细胞 PIWIL1 RNA干扰 

分 类 号：Q527[生物学—生物化学] R735.35[医药卫生—肿瘤]