临床肿瘤学杂志

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RNA干扰卷曲螺旋结构域蛋白34表达对膀胱癌细胞生物学行为的影响

巩艳青,宁向辉,丁义,郭应禄   

  1. 100034 北京 北京大学第一医院泌尿外科 北京大学泌尿外科研究所 国家泌尿男生殖系肿瘤研究中心
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2016-01-31 发布日期:2016-01-31
  • 通讯作者: 郭应禄

The impact of RNA interfering CCDC34 on the biological functions of bladder cancer cells

GONG Yanqing, NING Xianghui, DING Yi, GUO Yinglu   

  1. Department of Urology,Peking University First Hospital,Institute of Urology,Peking University,National Urological Cancer Center,Beijing 100034,China
  • Received:1900-01-01 Revised:1900-01-01 Online:2016-01-31 Published:2016-01-31
  • Contact: GUO Yinglu

摘要: 目的 探讨卷曲螺旋结构域蛋白34(CCDC34)的表达对膀胱癌细胞生物学行为包括增殖、凋亡、克隆形成和侵袭能力的影响。方法 选取膀胱癌5637细胞分别转染阴性对照siRNA(阴性对照组)和CCDC34siRNA(CCDC34干扰组)慢病毒载体,荧光显微镜观察慢病毒感染效率;实时荧光定量PCR和Western blotting验证CCDC34的干扰效率;BrdU掺入实验、流式细胞术、克隆形成实验和Transwell小室侵袭实验检测干扰CCDC34表达对5637细胞增殖、凋亡、克隆形成及侵袭能力的影响。结果 阴性对照组与CCDC34干扰组的慢病毒感染效率均在85%以上。与阴性对照组相比,感染CCDC34siRNA的细胞中CCDC34 mRNA表达水平受到显著抑制(1.00±0.06 vs. 0.16±0.01,P<0.01),且CCDC34蛋白表达下降。BrdU实验显示,CCDC34干扰组的细胞增殖能力较阴性对照组受到显著抑制(P<0.05),且细胞凋亡显著增加[(28.09±0.39)% vs.(9.50±0.01)%,P<0.01)]。克隆形成实验显示,CCDC34干扰组的克隆形成能力显著低于阴性对照组(22±4 vs.310±5,P<0.01)。Transwell小室侵袭实验显示,CCDC34干扰组膀胱癌细胞的侵袭转移率显著低于阴性对照组(0.53±0.03 vs.2.58±0.10,P<0.01)。结论 慢病毒介导的RNA干扰技术可下调CCDC34蛋白在膀胱癌5637细胞中的表达,并显著抑制肿瘤细胞的增殖、克隆形成和侵袭能力;CCDC34有望成为膀胱癌的生物标记物和治疗的新靶点。

Abstract: Objective To evaluate the impact of RNA interfering coiled-coil domain-containing 34(CCDC34)on the proliferation, apoptosis, colony formation and invasion of human bladder cancer cell lines. Methods Bladder cancer 5637 cells were infected with lentivirus-mediated CCDC34 negative control siRNA(negative control group)and CCDC34 siRNA(CCDC34siRNA group)respectively. Fluorescence microscopy was used to observe the infection efficiency. qPCR and Western blotting were used to confirm the interfering efficiency of CCDC34. The cell proliferation,apoptosis,colony formation and invasion ability were analyzed by BrdU incorporation assay,flow cytometry,lowdensity cell colony formation and transwell experiments. Results The lentivirus infection efficiency was above 85% for both CCDC34-siRNA group and negative control group. The mRNA expression of CCDC34siRNA group was greatly reduced compared with negative control group(1.00±0.06 vs. 0.16±0.01,P<0.01),as well as the expression of CCDC-34 protein. Compared with negative control group,cell proliferation was significantly inhibited in the CCDC34-siRNA group (P<0.05);the percentage of 5637 cells in apoptosis phase was also significantly increased in the CCDC34-siRNA group[(28.09±0.39)% vs.(9.50±0.01)%,P<0.01)]. The number of colonies in CCDC34-siRNA group was statistically less than that in negative control group(22±4 vs. 310±5,P<0.01). The cell invasion rate of CCDC34-siRNA group was significantly lower than that of negative control group(0.53±0.03 vs.2.58±0.10,P<0.01). Conclusion Down-regulation of CCDC34 in bladder cancer cell line 5637 via lentivirusmediated siRNA can inhibit the cell proliferation,colony formation and invasion. CCDC34 may serve as a biomarker or even a therapeutic target for bladder cancer.

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