稳定低表达DNA甲基转移酶3b基因对膀胱癌细胞生长和凋亡的影响

发布时间：2018-04-26 11:25

本文选题：膀胱癌 + DNMTb　；参考：《南方医科大学学报》2015年11期

【摘要】：目的探讨稳定低表达DNA甲基转移酶3b(DNMT3b)基因对人膀胱癌细胞生长和凋亡的影响。方法包装表达DNMT3b si RNA和阴性对照si RNA的慢病毒,通过慢病毒感染的方法获得稳定低表达DNMT3b的膀胱癌细胞BIU-87及对照细胞。MTT和流式细胞仪检测DNMT3b稳定低表达对细胞增殖和凋亡的影响;体内裸鼠成瘤实验观察DNMT3b稳定低表达对肿瘤的抑制效果;荧光定量PCR和Western blot检测DNMT3b稳定低表达对细胞生长和凋亡相关基因表达的影响;甲基化特异性PCR检测对细胞生长和凋亡相关基因甲基化的影响。结果荧光定量PCR和Western blot检测结果显示,DNMT3b m RNA和蛋白水平在BIU-87稳定细胞株中稳定低表达;DNMT3b稳定低表达可以抑制BIU-87细胞的生长,并且抑制裸鼠皮下瘤体的生长;DNMT3b稳定低表达可以促进BIU-87细胞的凋亡;DNMT3b稳定低表达可以增加凋亡及细胞生长相关基因DAPK,Bax和RASSF1A m RNA和蛋白水平的表达;DNMT3b稳定低表达可以减少凋亡及细胞生长相关基因DAPK,Bax和RASSF1A启动子区域的甲基化水平。结论 DNMT3b稳定低表达可能通过调控凋亡相关基因以及细胞生长相关基因启动子区域的甲基化水平来影响基因的表达,从而抑制BIU-87细胞的生长,诱导细胞凋亡。
[Abstract]:Objective to investigate the effects of stable and low expression of DNA methyltransferase (DNA) DNMT3b gene on the growth and apoptosis of human bladder cancer cells. Methods Lentivirus expressing DNMT3b si RNA and negative control si RNA were packaged. The stable and low expression of DNMT3b in bladder cancer cells BIU-87 and control cells were obtained by lentivirus infection and the effects of stable low expression of DNMT3b on cell proliferation and apoptosis were detected by flow cytometry. The inhibitory effects of stable and low expression of DNMT3b on tumor were observed in nude mice, and the effects of stable and low expression of DNMT3b on the expression of apoptosis-related genes were detected by fluorescence quantitative PCR and Western blot. Effect of methylation specific PCR on methylation of cell growth and apoptosis-related genes. Results the results of fluorescence quantitative PCR and Western blot showed that the stable and low expression of DNMT3b RNA and protein in BIU-87 stable cell lines could inhibit the growth of BIU-87 cells. Inhibiting the stable and low expression of DNMT3b in subcutaneous tumor of nude mice can promote the apoptosis of BIU-87 cells. The stable and low expression of DNMT3b can increase the expression of apoptosis and RASSF1A m RNA and protein levels. In order to reduce the methylation level in the promoter region of apoptosis and cell growth related gene DAPKG Bax and RASSF1A. Conclusion the stable and low expression of DNMT3b may affect the gene expression by regulating the methylation level of apoptosis-related genes and promoter regions of cell growth related genes, thus inhibiting the growth of BIU-87 cells and inducing apoptosis.
【作者单位】：南方医科大学珠江医院泌尿外科;
【基金】：广东省自然科学基金(S2012010009383,S2013040016823)
【分类号】：R737.14

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