PRDM5基因抑制前列腺癌细胞22Rv1生长

发布时间：2018-08-14 15:03

【摘要】：目的探讨PRDM5基因在前列腺癌细胞中的抑癌作用。方法采用聚合酶链反应(PCR)、限制性内切酶酶切、T4DNA连接酶连接等方法,克隆PRDM5基因并插入慢病毒载体中。将携带PRDM5基因的慢病毒质粒(或阴性对照质粒)与慢病毒包装质粒通过脂质体法共转染293T细胞,收集病毒上清,感染人前列腺癌细胞株22Rv1。用蛋白质印迹法检测细胞PRDM5的表达,细胞倍增实验和平板克隆实验检测细胞增殖和克隆形成能力,软琼脂克隆形成实验检测细胞非锚着依赖性生长能力。结果成功构建PRDM5重组慢病毒载体,并包装获得慢病毒上清。将PRDM5重组慢病毒载体感染22Rv1细胞后,筛选得到稳定表达细胞株,蛋白质印迹法结果显示该细胞株能稳定表达外源性PRDM5蛋白。过表达PRDM5的前列腺癌22Rv1细胞的增殖能力[倍增时间:(52.5±1.4)h vs(44.0±1.3)h]、克隆形成能力[克隆形成数:(1 114±98)/皿vs(1 361±123)/皿]和非锚着依赖性生长能力[克隆形成数:(94.6±8.7)/孔vs(154.0±3.5)/孔]均低于阴性对照组(P0.05)。结论前列腺癌22Rv1细胞中PRDM5的过表达具有抑制肿瘤细胞增殖、克隆形成和非锚着依赖性生长的能力。
[Abstract]:Objective to investigate the inhibitory effect of PRDM5 gene on prostate cancer cells. Methods PRDM5 gene was cloned and inserted into lentivirus vector by polymerase chain reaction (PCR),) restriction endonuclease digestion of T4 DNA ligase. The lentivirus plasmid carrying PRDM5 gene (or negative control plasmid) and lentivirus packaging plasmid were co-transfected into 293T cells by liposome method. The virus supernatant was collected and infected with human prostate cancer cell line 22Rv1. The expression of PRDM5 was detected by Western blotting, cell proliferation and clone formation were detected by cell multiplication assay and plate clone assay, and the ability of non-anchor dependent growth was detected by soft Agar colony forming assay. Results the recombinant lentivirus vector of PRDM5 was successfully constructed and the supernatant of lentivirus was obtained by packaging. After PRDM5 recombinant lentivirus vector was infected with 22Rv1 cell line, stable expression cell line was obtained. Western blot analysis showed that the cell line could stably express exogenous PRDM5 protein. The proliferative ability of 22Rv1 cells with overexpression of PRDM5 [(52.5 卤1.4) h vs (44.0 卤1.3) h], clone forming capacity [(1 114 卤98) / dish vs (1 361 卤123) / dish] and anchor-independent growth ability [(94.6 卤8.7) / pore vs (154.0 卤3.5) / well] were lower than those of negative control group (P0.05). Conclusion overexpression of PRDM5 in prostate cancer 22Rv1 cells can inhibit tumor cell proliferation, clone formation and anchor-independent growth.
【作者单位】：第二军医大学长海医院病理科;第二军医大学药学院生化药学教研室;解放军成都军区昆明总医院肿瘤科;第二军医大学长海医院泌尿外科;
【基金】：中国博士后科学基金面上项目(2013M532124) 上海市卫生和计划生育委员会科研课题面上项目(2013356)~~
【分类号】：R737.25

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