Pan Cuicui, Wu Xiaohou, Luo Chunli, Yang Shuzhe, Pu Jun, Wang Chunyuan, Shen Shali
Department of Laboratory Medical Diagnostics, Ministry of Education, Chongqing Medical University, Chongqing, China.
Urol Int. 2010;85(3):347-54. doi: 10.1159/000318889. Epub 2010 Jul 13.
We aimed to investigate the mechanisms of hepaCAM inactivation in transitional cell carcinoma of the bladder through the analysis of hepaCAM exon 2 methylation.
The methylation of hepaCAM exon 2 and the expression of hepaCAM were determined by methylation-specific restriction PCR assay and RT-PCR in bladder cancer cells (T24, BIU-87) as well as in 55 paired bladder cancer specimens. The methylated bladder cancer cells were treated with 5-Aza- 2'-deoxycytidine (5-Aza-CdR), a demethylating agent. MTT was used to detect the proliferation of T24 and BIU-87 cells.
The proliferation of T24 and BIU-87 cells was suppressed by treatment with different concentrations of 5-Aza-CdR; the expression of hepaCAM was absent in T24 and BIU-87 cells, and we found that exon 2 of hepaCAM was methylated in the 2 cells. hepaCAM mRNA was re-expressed and the methylation status of hepaCAM exon 2 was reversed after treatment with 5-Aza-CdR. The expression of hepaCAM mRNA in bladder cancer tissues was significantly lower than that in adjacent tissues. The methylation rate of hepaCAM exon 2 was significantly higher in bladder cancer tissues than in adjacent tissues. The methylation of hepaCAM exon 2 was related to hepaCAM expression in bladder cancer tissues.
Downregulation of hepaCAM expression plays an important role in the tumorigenesis and development of bladder cancer. DNA methylation may be important for downregulation of hepaCAM expression in bladder cancer.
通过分析hepaCAM外显子2甲基化情况，探讨其在膀胱移行细胞癌中失活的机制。
采用甲基化特异性限制性PCR法和RT-PCR法检测膀胱癌细胞（T24、BIU-87）及55对膀胱肿瘤组织标本中hepaCAM外显子2的甲基化情况及hepaCAM的表达。用去甲基化剂5-氮杂-2'-脱氧胞苷（5-Aza-CdR）处理甲基化的膀胱癌细胞。采用MTT法检测T24和BIU-87细胞的增殖情况。
不同浓度的5-Aza-CdR处理可抑制T24和BIU-87细胞的增殖；T24和BIU-87细胞中未检测到hepaCAM的表达，且发现这两种细胞中hepaCAM外显子2发生甲基化。经5-Aza-CdR处理后，hepaCAM mRNA重新表达，hepaCAM外显子2的甲基化状态逆转。膀胱癌组织中hepaCAM mRNA的表达明显低于癌旁组织。膀胱癌组织中hepaCAM外显子2的甲基化率明显高于癌旁组织。hepaCAM外显子2的甲基化与膀胱癌组织中hepaCAM的表达相关。
hepaCAM表达下调在膀胱癌的发生发展中起重要作用。DNA甲基化可能是膀胱癌中hepaCAM表达下调的重要原因。
