miR-21下调程序性死亡细胞4对抑制裸鼠肾细胞癌肿瘤转化和增殖的实验研究  

Inhibitory effect of down-regulation of programmed death cell 4 by microRNA-21 on tumorigenesis and proliferation of renal cell carcinoma in nude mice

作　　者：吴佳成 陆雅君[2] 姜力 WU Jiacheng;LU Yanjun;JIANG Li(Department of Urology, Heping Branch Hospital of Northern Theater Command, Liaoning Province,Shenyang 110000, China)

机构地区：[1]北部战区总医院和平分院泌尿外科,沈阳110000 [2]中国医科大学研究生院,沈阳110000

出　　处：《疑难病杂志》2019年第1期61-66,共6页Chinese Journal of Difficult and Complicated Cases

摘　　要：目的研究微小核糖核酸-21(miR-21)下调程序性死亡细胞4(PDCD4)对裸鼠肾细胞癌的影响和关系。方法将24只BALB/c雄性裸鼠按随机数字表法分为以下3组:阴性对照(NC组,n=8)、miR-21模拟物组(n=8)和miR-21抑制剂组(n=8)。将肾细胞癌786-0细胞皮下移植到小鼠的腋下,然后每天注射NC小干扰RNA(siRNA),前体-miR-21(模拟物)或抗-miR-21(抑制剂)。786-0细胞移植后16 d,从小鼠中取出肿瘤并称重。使用逆转录一定量PCR分析癌组织中miR-21和PDCD4 mRNA的表达。采用免疫组织化学和Western blot检测PDCD4蛋白在癌组织中的表达,用PDCD4 siRNA或NC siRNA转染786-0细胞,观察沉默PDCD4对肿瘤细胞生长、增殖和侵袭的影响。雌性软琼脂克隆形成、EdU细胞增殖测定和Transwell迁移和侵袭测定。另取16只BALB/c雄性裸鼠随机分为2组:NC(n=8)和PDCD4 siRNA(n=8)。将786-0细胞皮下移植到小鼠的腋下,每天注射NC siRNA或PDCD4siRNA。移植后16 d取出肿瘤并称重。结果与NC组相比,miR-21模拟组的肿瘤重量显著增加,miR-21抑制剂组中的肿瘤重量显著降低(P<0.01)。裸鼠肾癌模型中的miR-21表达在miR-21模拟组中显著上调,与NC组相比显著降低(P<0.01)。在miR-21抑制剂组中,miR-21模拟组PDCD4蛋白水平显著降低,miR-21抑制剂组显著增加(P<0. 01),而PDCD4 mRNA表达没有显著改变(P>0.05)。在EdU增殖测定中,NC siRNA组中的新细胞合并EdU的平均百分比为28.6%,并且在PDCD4 siRNA转染的细胞中显著增加至44.7%(P <0. 05)。在软琼脂克隆形成实验中,Transwell和迁移和侵袭实验显示,PDCD4 siRNA转染细胞的集落形成、迁移和侵袭能力显著增加,与NC组相比,肿瘤重量在PDCD4 siRNA组中显著增加(P<0.01)。结论 miR-21可促进裸鼠肾癌模型癌细胞增生和增殖,转录后下调PDCD4蛋白表达可抑制癌细胞增生和增殖,提示增加PDCD4表达或抑制miR-21表达可能成为治疗肾癌的有效新治疗策略。Objective To study the effect of microRNA-21 (miR-21) down-regulation of programmed death cell 4 (PDCD4) on renal cell carcinoma in nude mice and its relationship. Methods Twenty-four BALB/c male nude mice were randomly divided into three groups: negative control group (NC group, n =8), mimic group ( n =8) and inhibitor group ( n = 8). Renal cell carcinoma 786-O cells were subcutaneously transplanted into the axilla of mice and then injected with NC small interfering RNA (si RNA), precursor microRNA-21 (mimic) or anti-microRNA-21 (inhibitor) every day. Sixteen days after transplantation of 786-O cells, the tumors were removed from the mice and weighed. Reverse transcription quantitative PCR was used to analyze the expression of microRNA-21 and PDCD4 in cancer tissues. The expression of PDCD4 protein in cancer tissues was detected by immunohistochemistry and Western blot. 786-O cells were transfected with PDCD4 siRNA or NC siRNA. The effects of silencing PDCD4 on the growth, proliferation and invasion of cancer cells were observed. Female soft agar cloning, EdU cell proliferation assay and Transwell migration and invasion assay. Another 16 BALB/c male nude mice were randomly divided into two groups: NC ( n =8) and PDCD4 siRNA ( n =8). 786-O cells were subcutaneously transplanted into the axilla of mice and injected with NC siRNA or PDCD4 siRNA every day. Tumors were removed and weighed 16 days after transplantation. Results Compared with NC group, the weight of tumors in mimic group increased significantly and that in inhibitor group decreased significantly. The expression of microRNA in nude mice kidney cancer model was significantly up-regulated in the mimic group of microRNA, and significantly lower than that in the NC group. In the miR-21 inhibitor group, the PDCD4 protein level was significantly decreased in the mimic group, and significantly increased in the miR-21 inhibitor group, while the PDCD4 gene expression was not significantly changed. In EdU proliferation assay, the average percentage of new cells i

关 键 词：程序性死亡细胞4 微小核糖核酸-21 裸鼠 肾癌 肿瘤转化 增殖 

分 类 号：R737.11[医药卫生—肿瘤]