骨桥蛋白shRNA表达载体介导的RNA干扰对前列腺癌PC-3细胞生长和侵袭的影响  

Effect of short-hairpin RNA expression vector-mediated osteopontin RNA interference on proliferation and invasion of prostate cancer PC-3 cells

作　　者：柳昊[1] 陈安民[1] 郭风劲[1] 廖晖[1] 胡伟华[1] 解杰[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院骨科,湖北武汉430030

出　　处：《癌症》2010年第1期43-51,共9页Chinese Journal of Cancer

基　　金：国家"973"重点基础研究发展规划资助项目(No.2002CB513100)~~

摘　　要：背景与目的:骨桥蛋白(osteopontin,OPN)诱导多种恶性肿瘤细胞的浸润和转移与基质金属蛋白酶(matrix metalloproteinase,MMP)-2和MMP-9的活化密切相关。本研究旨在探讨OPN在人前列腺癌PC-3细胞的增殖和侵袭过程中的作用以及IκB激酶(Ikappa B kinase,IKK)在NF-κB介导的信号通道中可能的功能。方法:将OPN短发夹RNA(short-hairpin RNA,shRNA)重组质粒转染PC-3细胞并应用不同浓度的IKK抑制剂分别抑制IKKα和IKKβ的活性,然后采用实时PCR和Western blot分别检测OPN、MMP-2和MMP-9三种mRNA和蛋白的表达水平,流式细胞术、MTT法和Transwell实验分别检测细胞生长周期变化、细胞增殖和和侵袭能力。结果:与对照组相比,转染重组质粒的细胞中OPN、MMP-2和MMP-9三种蛋白的表达量分别下降了55.22%、51.71%和28.35%(P<0.05),凋亡二倍体数目显著增加,细胞的迁移能力和侵袭能力分别下降了45.48%和51.96%(P<0.05)。此外,特异性地抑制IKKβ酶的活性能显著降低MMP-2和MMP-9的表达水平。结论:shRNA表达载体介导的OPN基因沉默能明显抑制PC-3细胞的恶性生物学行为;在OPN诱导MMP-2和MMP-9活化的过程中,IKKβ酶的活性可能发挥了重要的作用。Background and Objective：Studies showed that osteopontin （OPN） regulates cell migration and invasion in a variety of cancers,which associates with the activities of matrix metalloproteinase （MMP）-2 and MMP9.This study was to investigate the role of OPN in the proliferation and invasion of human prostate cancer PC-3 cells and the possible functions of I-κB kinase （IKK） in nuclear factor kappa B （NF-κB）-mediated signaling pathways.Methods：OPN short-hairpin RNA （shRNA） recombinant plasmids （PCs） were transfected into PC-3 cells and the different concentrations of IKK inhibitors were used to inhibit the activities of IKKα and IKKβ.The mRNA and protein expression levels of OPN,MMP-2,and MMP-9 were detected by real-time PCR and Western blot.The different cell cycles,cell proliferation,and invasion abilities of PC-3 cells were detected by flow cytometry,MTT,and Transwell chamber assays,respectively.Results：Compared with untreated cells,the protein expression levels of OPN,MMP-2,and MMP-9 in P...更多C-3 cells transfected with PCs were decreased by 55.22%,51.71%,and 28.35%,respectively,and the abilities of migration and invasion of PCs were decreased by 45.48% and 51.96%,respectively （P〈0.05）.Moreover,the inhibition of IKKβ activity reduced the expressions of MMP-2 and MMP-9.Conclusions：A shRNA expression vector-mediated OPN gene silencing can inhibit the malignant physiological behaviors of PC-3 cells.IKKα may play a crucial role in the OPN-induced activation of MMP-2 and MMP-9 via NF-κBmediated I-κB /IKKβ pathway.

关 键 词：前列腺肿瘤 PC-3细胞 短发夹RNA 骨桥蛋白 基质金属蛋白酶 RNA干扰 核因子κB 

分 类 号：R737.25[医药卫生—肿瘤] R730.5[医药卫生—临床医学]