逆转录病毒介导重组CXCR4对前列腺癌细胞侵袭力的影响

Effects of retroviral-mediated recombinant CXCR4 gene on invasion of prostate cancer cell line in vitro

目的:构建CXCR4基因重组逆转录病毒表达载体pLEGFP-CXCR4,并转染PC-3细胞,探讨趋化因子受体CXCR4对人前列腺癌细胞体外侵袭能力的影响。方法:分离健康人外周血单个核细胞,提取总RNA,以其为模板,RT-PCR法扩增CXCR4,获取CXCR4基因全长序列,装入带有绿色荧光蛋白(GFP)的逆转录病毒质粒pLEGFP-N1,用脂质体法转染PC-3细胞,采用实时定量PCR和Western blotting观察CXCR4表达情况,并通过体外细胞-基质粘附试验和Transwells小室检测肿瘤细胞体外侵袭能力的变化。结果:成功构建pLEGFP-CXCR4载体,转染人前列腺癌细胞72h后,CXCR4 mRNA及蛋白质水平明显上调,细胞的体外侵袭力明显增强,增殖活性增强。结论:CXCR4转染能够明显提高CXCR4基因mRNA及蛋白水平,提高人前列腺癌细胞体外侵袭能力。

Objective：To constructing CXCR4 gene recombinant retrovirus vector PLEGFP - N1, then transfect PC -3 cells, and study the effect of chemokine receptor CXCR4 - targeted on invasion capability of human prostate Cancer cell line in vitro. Methods：Total RNA was extracted from monocyte separated from peripheral blood of health human. And CXCR4 gene was amplified by RT- PCR,then inserted it into retroviral vector which contain Green Flu Protein, named pLEGFP- N1. Then transfected with lipofectamine mediation to PC -3 cells. The mRNA level of CX- CR4 was detected by real - time PCR, expression of CXCR4 protein was detected by Western blotting at 72h after transfection. The abilities of invasion were detected by Matrigel invasion assay. Results ：The retrovirus vectors pLEG- FP - CXCR4 were constructed successfully, the expression of CXCR4 mRNA and protein was up - regulated significantly after transfection of CXCR4. The invasion and proliferation capabilities of PC - 3 cells were enhanced. Conclusion： CXCR4 effectively up - regulates the expression of CXCR4 gene and improves invasion capability of PC - 3 cells in vitro.