去甲基化药物5-氮-2'-脱氧胞苷抑制膀胱肿瘤细胞生长作用的实验研究

目的探讨去甲基化药物5-氮-2’-脱氧胞苷（5-aza-2’-deoxycytidine,DAC）对膀胱肿瘤细胞生长的抑制作用。方法采用WST-1法测定DAC对膀胱癌细胞株的生长影响作用,流式细胞仪检测其在诱导细胞凋亡与细胞周期捕获中的效果,APOPCYTO半胱天冬酶（Caspase）色度法分析DAC处理后Caspase 3及Caspase 9的活性,Western blot检测DAC处理后增殖细胞核抗原（PCNA）的表达情况。结果 DAC对膀胱肿瘤细胞的生长具有抑制作用,药物作用呈剂量依赖性,并且该作用不受p53表型影响。DAC不能诱导膀胱肿瘤细胞凋亡,细胞中Caspase 3、9的活性也未见激活,但DAC可以抑制膀胱癌细胞的增殖能力,PCNA蛋白的表达量呈剂量依存性降低,发生G2/M期捕获。用0、1和8μmol/LDAC处理RT112后G2/M期细胞分别为（36.3±3.4）%、（46.2±4.6）%和（56.5±6.2）%;处理TCCsup后为（37.5±3.8）%、（48.4±4.9）%和（60.1±6.7）%。结论 DAC对膀胱肿瘤细胞生长具有抑制作用,可能成为将来治疗膀胱恶性肿瘤的一种新型药物。

The effect of demethylation agent 5-aza-2'-deoxycytidine on the growth suppression of transitional cell carcinoma cells

Objectives To study the growth suppression effect of demethylation drug 5-aza-2’-deoxycytidine(DAC) on bladder tumor cells. Methods The growth suppression effect of DAC on four transitional cell carcinoma (TCC) cell lines was measured using the WST-1 assay. The effects of DAC on apoptosis induction and cell cycle arrest were analyzed by flow cytometry analysis. The activity of caspase 3,9 were analyzed by APOPCYTO Caspase Colorimetric Assay Kit and the PCNA expression was also investigated by western blot to clarify the mechanism of DAC against TCC. Results DAC inhibited the growth of all TCC cell lines tested in a dose-depend manner, however, growth suppression effect of DAC was independent of p53 status in TCC. DAC inhibited proliferation via inducing G2/M cell cycle arrest but not via inducing apoptosis. The expression of PCNA was decreased by DAC, but caspase 3, 9 activity was not activated. After treated with 0, 1 and 8 μmol/L DAC, cells of RT112 in G2/M phase was(36.3±3.4)%,(46.2±4.6)% and (56.5±6.2)%, TCCsup was(37.5±3.8)%,(48.4±4.9)% and (60.1±6.7)%, respectively. Conclusion DAC could suppress the growth of TCC cells and may be a new strategy to treat bladder malignancy in the future.