地西他滨体外对小鼠急性粒-单核白血病细胞WEHI-3的抗肿瘤作用及机制探讨

Anti-tumor effect and mechanism of decitabine on acute granulocyte-monocytic leukemia cell line WEHI-3 in mice in vitro

目的探讨去甲基化药物地西他滨对于小鼠急性粒-单核白血病细胞WEHI-3的凋亡、细胞周期的影响及其作用机制。方法急性粒-单核白血病细胞(WEHI-3)采用小剂量地西他滨0.25μmol/L连续体外处理3 d。采用瑞氏吉姆萨染色法观察细胞形态变化,AnnexinV-PI法检测WEHI-3不同时间的凋亡情况,流式细胞术检测细胞周期变化情况,Q-PCR检测处理前后mRNA的表达情况。结果地西他滨处理后,WEHI-3细胞体积明显增大,胞质增多,形态不规则,染色质浓缩、边缘化;PBS处理的WEHI-3细胞G1期占39.64%,S期占49.43%,G2期占8.74%,地西他滨处理后WEHI-3细胞G1期增加至78.02%,S期减少至15.86%,G2期减少至2.91%,可见地西他滨处理后WEHI-3被阻滞于G1期;经地西他滨处理后WEHI-3细胞中MMD、TSG101、TAF7L、CITED2 mRNA表达水平显著提高(P均<0.01)。结论地西他滨可上调MMD、CITED2、TAF7L、TSG101基因表达,诱导小鼠急性粒-单核白血病WEHI-3细胞凋亡、细胞周期阻滞。

Objective To investigate the effects of demethylation drug decitabine on the apoptosis and cell cycle in acute myelomonocytic leukemia cells in mice and its mechanism. Methods Acute myelomonocytic leukemia cells（WEHI-3） were treated with decitabine（0.25μmol/L） for 3 days. In this study, changes in cell morphology were observed using Wright＇s Giemsa staining, apoptosis at different time points of WEHI-3 was measured by Annexin V-PI method, cell cycle change was detected by flow cytometry, mRNA change was detected by Q-PCR. Results After treating WEHI-3 cells with decitabine, the volume of cells increased significantly, with increase of cytoplasm and irregular shape, and the chromatin was concentrated and marginalized. The cell cycle characteristics of PBS treated cells were quantified as follows： G1 phase 39.64%, S phase 49.43%, G2 phase 8.74%. After treatment with decitabine, G1 phase of WEHI-3 cells increased to 78.02%, S phase decreased to 15.86%, and G2 phase decreased to 2.91%, which suggested that WEHI-3 was arrested in G1 phase. The expression levels of MMD, TSG101, TAF7L and CITED2 mRNA in WEHI-3 cells increased significantly after treated by decitabine（all P〈0.01）. Conclusion Decitabine promotes apoptosis of WEHI-3 cells, and may affect the proliferation of WEHI-3 cells through MMD, CITED2, TAF7L and TSG101. Decitabine can up-regulate the expression of MMD, CITED2, TAF7L and TSG101 genes, induce apoptosis and arrest cell cycle of acute-monocytic leukemia WEHI-3 cells in mice.