三氧化二砷对膀胱癌T24细胞Apaf-1基因表达的影响

目的：观察三氧化二砷 (arsenic trioxide，As2O3)对人膀胱癌T24细胞增殖及凋亡蛋白酶活化因子-1 (apoptotic protease activating factor-1，Apaf-1)基因表达的影响。方法：取处于对数生长期的T24细胞，用0.25%胰酶消化成细胞密度为1×105/mL的单细胞悬液，再用含有不同浓度 (0、1、2、4、8 μmol/L)As2O3的培养基作为药物组，并设置空白对照组，置于培养箱中分别培养24、48和72 h后，用二甲基四氮唑蓝 (MTT)法检测As2O3对T24细胞的增殖抑制率；取T24细胞在不同浓度 (0、1、2、4、8 μmol/L)的As2O3培养液中培养72 h后用脱氧核糖核酸原位末端转移酶标记技术 (TUNEL)法检测细胞凋亡，用RT-PCR及Western blotting检测As2O3 对Apaf-1 mRNA和蛋白表达的影响。结果：与对照组相比，2、4、8 μmol/L As2O3剂量组能有效抑制T24细胞增殖，且具有剂量-反应关系 (24 h时r=-0.962，P=0.038；48 h时r =-0.959，P=0.041；72 h时r=-0.973，P=0.027)。As2O3于1～8 μmol/L作用72 h时细胞出现典型的凋亡变化，具有剂量-反应关系 (r=0.993，P =0.007)；同时Apaf-1 mRNA和蛋白的表达均明显增强，且具有剂量-反应关系 (mRNA：r=0.986，P=0.014；蛋白：r=0.989，P=0.000)。结论：As2O3能够有效抑制膀胱癌T24细胞生长、增殖并诱导其凋亡，机制可能与其上调Apaf-1基因的表达有关。

Effect of arsenic trioxide on Apaf-1 gene expression in bladder cancer T24 cells

OBJECTIVE: To observe the effects of arsenic trioxide (As2O3) on human bladder cancer T24 cell proliferation and apoptosis protease activating factor-1 (Apaf-1) gene expression. METHODS：T24 cells in the logarithmic growth phase，were digested with 0.25% trypsininto a cell density of 1×105/mL single cell suspension.Then cells were inculated with different concentrations (0，1，2，4，8 μmol/L) of As2O3，as well as blank control group for 24，48 and 72 h. Cell proliferation inhibition rate was evaluated by MTT. After cell treatment with As2O3(0，1，2，4，8 μmol/L) for 72 h，deoxyribose situ terminal transferase labeling(TUNEL) method was used to measure apoptosis；RT-PCR and Western blotting to assess the impact of As2O3 on Apaf-1 mRNA and protein expression. RESULTS：Compared with the control group， the 2，4，8 μmol/L As2O3dose groups could effectively inhibit the proliferation of T24 cells in a concentration-dependent manner (24 h，r =0.962，P=0.038；48 h，r =0.959，P=0.041；72 h，r =0.973，P=0.027). Cells treated with As2O3 at 1-8 μmol/L for 72 h， showed typical apoptotic changes in a concentration-dependent manner (r= 0.993，P=0.007); whilst Apaf-1 mRNA and protein expressions were also significantly increased in a concentration-dependent (mRNA，r =0.986，P=0.014；protein，r =0.989， P=0.000). CONCLUSION：As2O3 could effectively inhibit bladder cancer T24 cell growth，proliferation and induce apoptosis. The possible mechanism may be related to increased Apaf-1 gene expression.