阿糖胞苷对肺腺癌细胞凋亡的诱导作用及其机制

STUDY ON MECHANISM OF APOPTOSIS OF LUNG ADENOCARCINOMA CELL INDUCED BY CYTARABINE

目的探讨阿糖胞苷(Ara-C)对人肺腺癌细胞株A549的凋亡诱导作用及其机制。方法Ara-C体外作用于A549细胞,噻唑蓝还原法(MTT法)检测Ara-C对A549细胞增殖的抑制作用;Hoechst33258荧光染色观察细胞核形态学的变化;单细胞凝胶电泳技术(comet assay)测定A549细胞DNA的损伤程度;以Western blotting进一步证明A549细胞发生凋亡。结果Ara-C对A549细胞的增殖有明显抑制作用;观察到特异性的凋亡小体;Ara-C导致A549细胞发生DNA链断裂,并呈明显剂量依赖性增强:Western blotting显示caspase 8,9,3都不同程度被激活,多聚ADP核糖聚合酶(PARP)被剪切降解。结论揭示了Ara-C明显诱导A549细胞凋亡;细胞凋亡通路不仅通过线粒体途径,还通过膜受体途径,两条通路协同作用使凋亡信号增强;提示Ara-C对A549细胞有明显的化疗作用。

Objective To study the mechanism of apoptosis of human lung adenocarcinoma cell line A549 induced by cytarabine （Ara-C） in vitro. Methods Ara-C induced dose-dependent inhibition of cell proliferation by MTr assay. Morphological changes of the A549 cells stained with Hoechst33258 were observed by microscopy; The DNA fragments were detected by single-cell gel electrophoresis （ comet assay） ; The expressions of caspase3,8,9, and poly （ADP-ribose） polymerase （PARP） apoptosis-related protein were investigated by Western blotting. Results After a 72 hours exposure to Ara-C, A549 cells were detected by classic apoptotic morphology, DNA fragmentation, activation of caspase 3, and PARP degradation. Ara-C induced DNA damage was associated with activation of caspase 9, a mitochondrial-activated caspase. In contrast, a lower level of caspase 8, the death receptor-activated caspase, was detected in Ara-C-exposed A549.Conelusion Ara-C can effectively induce apoptosis of human lung adenocarcinoma cell llne A549 and simultaneously Ara-C-induced apoptosis is mediated mainly through a mitochondrion-dependent pathway and partly through a receptor-dependent pathway.