慢病毒介导RNA干扰HPIP蛋白抑制肿瘤细胞增殖  

Lentiviral-Mediated Knock-Down of Human HPIP Inhibits Cell Growth and Proliferation

作　　者：徐小洁[1] 范忠义[2] 韩永健[1] 张浩[1] 丁丽华[1] 韩聚强[3] 王凌雪[1] 杨智洪[1] 杜楠[2] 叶棋浓[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100850 [2]解放军总医院第一附属医院,北京100048 [3]北京军区总医院,北京100700

出　　处：《生物技术通讯》2011年第3期321-324,共4页Letters in Biotechnology

基　　金：国家自然科学基金(30800205和31071174);国家重大科学研究计划(2007CB914603);全军医药卫生科研基金(06J021)

摘　　要：目的:建立高效稳定的造血相关的PBX相互作用蛋白质(HPIP)小干扰RNA(siRNA)细胞导入方法,检测HPIP的表达对肿瘤细胞生长增殖的影响。方法:构建人HPIP慢病毒siRNA干扰载体,将重组质粒转染人胚肾293T细胞,通过实时定量RT-PCR及Western印迹分析检测Lenti-H1 HPIP siRNA的干扰效果;将Lenti-H1 HPIPsiRNA与4个包装质粒共同转染293T细胞,包装成慢病毒后,感染宫颈癌HeLa和肝癌HepG2细胞,经嘌呤霉素筛选2周后,收集细胞进行Western印迹检测;用结晶紫实验检测其对肿瘤细胞生长增殖的影响。结果:构建的Lenti-H1 HPIP siRNA能有效抑制HPIP的表达;结晶紫实验显示,慢病毒介导的HPIP siRNA导致细胞增殖的显著抑制。结论:慢病毒介导的HPIP敲减能明显抑制肿瘤细胞的增殖,HPIP可能是一个潜在的肿瘤治疗新靶点。Objective：To establish an efficient and stable way to deliver hematopoietic PBX-interacting protein（HPIP） small interfering RNA（siRNA） and examine the effect of HPIP siRNA on cell proliferation.Methods：pLenti-H1/HPIP siRNA was constructed and transfected into 293T cells.Western blot and real-time RT-PCR were carried out to detect the inhibitory effect of Lenti-H1 HPIP siRNA.The four viral packaging vectors together with pLenti-H1/HPIP siRNA were used to transfect 293T cells to produce lentivirus particles.After HeLa and HepG2 cells were infected with the obtained viruses and screened for 2 weeks with puromicin,the expression of HPIP was examined by Western blot analysis.Crystal violet assay was performed to investigate the effect of lentivirus on tumor growth and proliferation.Results：Western blot and real-time RT-PCR showed that pLenti-H1/HPIP siRNA could suppress the expression level of HPIP.Crystal violet analysis showed that down-regulation of HPIP could obviously suppress cell growth and proliferation.Conclusion：pLenti-H1/HPIP siRNA can significantly inhibit the expression of HPIP and suppress cell growth and proliferation.HPIP might serve as a novel target for cancer therapy.

关 键 词：造血相关的PBX相互作用蛋白质 RNA干扰 慢病毒 细胞增殖 

分 类 号：Q786[生物学—分子生物学] R735.7[医药卫生—肿瘤]