抑癌基因DKK2过表达对人膀胱癌细胞增殖和迁移的抑制作用及其机制探讨

发布时间：2018-04-27 22:19

  本文选题：膀胱肿瘤 + Wnt信号通路　； 参考：《肿瘤》2017年11期

【摘要】：目的 :分析dickkopf 2(DKK2)基因在人膀胱癌细胞系及膀胱癌组织中的表达水平,以及DKK2过表达对人膀胱癌细胞增殖与迁移的影响,并初步探讨其作用机制。方法:通过RT-PCR、实时荧光定量PCR和蛋白质印迹法检测人膀胱癌细胞系及组织中DKK 2基因的相对表达水平。选用DKK2低表达的膀胱癌细胞系T24,进行DKK2过表达质粒转染后,采用RT-PCR、实时荧光定量PCR和蛋白质印迹法验证DKK2过表达效果;然后采用CCK-8法、平板克隆形成实验、划痕愈合实验、Transwell小室法和FCM法分别检测DKK2过表达对细胞增殖、克隆形成、迁移、侵袭及细胞周期的影响;进一步采用蛋白质印迹法和实时荧光定量PCR法检测DKK2过表达后膀胱癌细胞中增殖和迁移相关分子表达水平的变化。结果:人膀胱癌细胞系5637、T24、SW780、J82、HT-1197和HT-1376中DKK2 mRNA及蛋白表达水平均明显低于正常膀胱上皮细胞SVHUC-1(P值均0.01);同时与正常膀胱组织及配对的癌旁组织相比,膀胱癌组织中DKK2 mRNA及蛋白的表达均明显下调(P值均0.001)。DKK2过表达质粒转染后,膀胱癌T24细胞中DKK2 mRNA及蛋白的表达水平明显上调,而细胞的增殖、克隆形成、迁移和侵袭能力均受到明显抑制(P值均0.05),同时细胞周期被阻滞于G0/G1期(P0.001)。而且DKK2过表达的膀胱癌T24细胞中,p21和E-cadherin表达水平明显升高(P值均0.001),cyclin D1、vimentin和N-cadherin表达水平明显降低(P值均0.001)。结论:DKK2在人膀胱癌细胞及组织中低表达。DKK 2可能作为一个潜在的抑癌基因,参与膀胱癌进展。
[Abstract]:Aim: to investigate the expression of dickkopf 2 gene in human bladder cancer cell lines and bladder cancer tissues, and the effect of DKK2 overexpression on the proliferation and migration of human bladder cancer cells. Methods: RT PCR, real-time quantitative PCR and Western blot were used to detect the relative expression of DKK 2 gene in human bladder cancer cell lines and tissues. Bladder cancer cell line T24, which has low expression of DKK2, was used to transfect DKK2 overexpression plasmid, RT-PCR, real-time fluorescence quantitative PCR and Western blotting were used to verify the effect of DKK2 overexpression. The effects of DKK2 overexpression on cell proliferation, clone formation, migration, invasion and cell cycle were detected by FCM and Transwell chamber method. Western blotting and real-time fluorescence quantitative PCR were used to detect the expression of proliferative and migration-related molecules in bladder cancer cells after overexpression of DKK2. Results: the expression levels of DKK2 mRNA and protein in human bladder cancer cell line 5637, T24T24, SW780, J82, HT-1197 and HT-1376 were significantly lower than those in normal bladder epithelial cells, and were significantly lower than those in normal bladder tissues and matched adjacent tissues. The expression of DKK2 mRNA and protein in bladder cancer tissue was significantly down-regulated, and the expression of DKK2 mRNA and protein in T24 cells was upregulated, while the proliferation and clone formation of T24 cells were observed after transfection with both 0.001).DKK2 overexpression plasmids. Migration and invasion were significantly inhibited (P = 0.05) and cell cycle was blocked in G0/G1 phase (P 0.001). Furthermore, the expression of p21 and E-cadherin in T24 cells with overexpression of DKK2 was significantly increased (P = 0.001), and the expression of cyclin D _ (1) vimentin and N-cadherin decreased significantly (P = 0.001). Conclusion the low expression of DKK2 in human bladder cancer cells and tissues may be a potential tumor suppressor gene involved in the progression of bladder cancer.
【作者单位】： 重庆市中医院(重庆第一人民医院)肾内科;
【分类号】：R737.14

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