慢病毒介导的稳定表达LBH的前列腺癌细胞株的建立

目的:构建稳定表达LBH基因的人前列腺癌细胞株PC-3M-LBH,探讨LBH基因对PC-3M细胞增殖能力的影响。方法:构建表达LBH基因的重组慢病毒载体并制备出相应的慢病毒,感染低表达LBH基因的人前列腺癌PC-3M细胞后,经嘌呤霉素筛选获得细胞克隆;实时荧光定量PCR和蛋白印迹法(Western-Blot)分别检测细胞株中LBH的mRNA、蛋白表达水平;采用CCK-8法检测表达LBH基因后细胞增殖能力的改变。结果:成功构建了重组慢病毒表达质粒p Lenti-LBH并包装出了慢病毒,感染前列腺癌细胞后经嘌呤霉素筛选得到PC-3M-LBH细胞株;PC-3M-LBH细胞株中LBH基因的mRNA和蛋白表达显著上调;相对母细胞和NC对照组,PC-3M-LBH细胞在接种后第4天即出现明显的生长抑制,到第6天其生长抑制率达到19.7%。结论:构建的细胞株能稳定表达LBH基因,该基因的表达能显著抑制前列腺癌PC-3M细胞的体外增殖。

The Establishment of PC-3MCell Stably Expressing LBH Mediated by Lentiviral

Objective:To establish a PC-3M cell line stably expressing LBH gene and explore the effect of LBH gene on the proliferation of PC-3M cells.Methods:Recombinant lentiviral expression vector containing LBH gene was constructed and lentivirus was produced. After lentivirus infection of human prostatic carcinoma PC-3M cells, PC-3M-LBH cell line was obtained by screening with puromycin; Expression of LBH in different cells was examined by RT-PCR and Western blot; Cell proliferation was detected by CCK-8 assay.Results:Lentiviral expression plasmid plenti-LBH was constructed and lentivirus particle was successfully packed. After infected with lentivirus and screened by puromycin, a cell line PC-3M-LBH was obtained. LBH expression in PC-3M-LBH was significantly higher compared with the NC control group and parental cells. The proliferation of PC-3M-LBH cells decreased 19.7 % compared with NC control and PC-3Mat the 6th day after cells seeding.Conclusion:LBH gene expression can inhibit the proliferation of prostate cancer PC-3Mcells.