吲哚美辛诱导的HL-60白血病细胞凋亡与JNK信号转导途径活化

目的：观察吲哚美辛对HL-60白血病细胞增殖的抑制作用及细胞凋亡的诱导作用，了解c-Jun N-末端激酶(JNK)信号转导途径在介导吲哚美辛诱导的HL-60细胞凋亡中的活化状态，揭示吲哚美辛诱导HL-60细胞凋亡的分子机制。方法：以台盼蓝染色检测药物干预后的HL-60细胞活力；分析HL-60细胞的增殖能力；DNA梯状胶电泳及吖啶橙／溴化乙锭染色形态学分析鉴定细胞凋亡；Western印迹检测凋亡信号蛋白caspase-9，-3和PARP的裂解激活以及JNK信号途径蛋白质MEK4，JNK，P-JNK，P-C-Jun的表达及活化状态。结果：200～400txmol的吲哚美辛能够显著抑制HL-60细胞增殖和诱导HL-60白血病细胞凋亡，并呈浓度和时间依赖性；HL-60细胞凋亡伴有easpase-9，3和PARP的表达上调及裂解激活；MEK4蛋白表达呈浓度依赖性上调；磷酸化JNK和磷酸化C-Jun呈浓度依赖性上调。结论：吲哚美辛可抑制HL-60细胞增殖和诱导细胞凋亡；JNK信号途径活化介导了吲哚美辛诱导的凋亡。JNK途径的活化导致了下游caspase途径的活化。

Indomethacin-induced HL-60 leukemic cell apoptosis and the activation of C-jun NH2- terminal kinase signal transduction pathway]

OBJECTIVE: To observe the effect of indomethacin on inhibiting the proliferation of HL-60 cells and inducing HL-60 cells apoptosis and to explore the activation of C-Jun NH2-terminal kinase (JNK) signal transduction pathway in mediating indomethacin-induced HL-60 cell apoptosis. METHODS: The cell viability was determined by Trypan blue staining and the state of cell proliferation was analyzed; DNA ladder pattern and AO/EB staining were applied to identify the cell apoptosis; the apoptotic signal proteins including caspase-9, caspase-3, and PARP and the proteins of JNK signal pathway such as MEK4, JNK, P-JNK, and P-C-Jun, were detected by Western blotting. RESULTS: Indomethacin at 200 to approximately 400 micromol significantly inhibited the proliferation of HL-60 cells and induced the cell apoptosis in a time- or concentration-dependent manner; caspase-9, caspase-3, and PARP were cleaved and activated in undergoing apoptotic cells; and the expressions of MEK4, P-JNK, and P-C-Jun were upregulated with the increase of indomethacin concentration. CONCLUSION: Indomethacin can inhibit the proliferation of HL-60 cell and induce leukemic cell apoptosis. The activation of JNK signal transduction pathway mediates the event of indomethacin-induced HL-60 cell apoptosis. JNK signal pathway is located in the upstream of caspase signal pathway.