Jak激酶抑制剂AG490对Lewis肺癌细胞增殖及Jak-Stat信号通路的影响

Influence of Jak kinase inhibitor AG490 on Lewis lung cancer cell proliferation and Jak-Stat signaling pathways

目的探讨Jak激酶抑制剂AG490对Lewis肺癌细胞增殖与凋亡的作用及其机制。方法 AG490作用于体外培养的Lewis肺癌细胞。MTT法检测细胞增殖,吖啶橙荧光染色观察细胞凋亡形态、流式细胞术检测细胞周期与凋亡,逆转录聚合酶链反应(reverse transcription polymerase chain reaction,RT-PCR)检测Stat3、Stat5和Bcl-2、Bax mRNA表达。结果 100μmol.L-1AG490作用24和48h后,对Lewis肺癌细胞细胞增殖抑制率分别为37.95%和52.99%。流式细胞术显示,经50和100μmol.L-1AG490作用24h后,细胞凋亡率由处理前的0.76%分别提高到12.56%和16.76%(P<0.01)。荧光显微镜下可见到典型的细胞凋亡形态学改变。细胞Stat3、Stat5活化程度降低,促进Bcl-2基因表达下调,提升细胞中Bax mRNA表达。结论阻断Jak-Stat3信号通路可抑制Lewis肺癌细胞增殖,并可能通过下调凋亡抑制基因Bcl-2表达诱导细胞凋亡。

Objective To explore the effect of AG490 on Lewis lung cancer cells line.Methods Lewis lung cancer cells in log phase were collected and devided into negative control group and experimental groups.Experimental groups were treated with different concentrations of AG490（25,50 and 100 μmol·L-1）.Changes of cell proliferation were analyzed by MTT assay,and the alteration of cell cycles was determined by flow cytometry after propidium iodide staining.Acridine orange fluorescence labeling was applied to measure the apotosis ratios of Lewis lung cancer cells.Expression levels of Stat3,Stat5,Bax and Bcl-2 mRNA were examined by a real time PCR method.Results The proliferation inhibition rates of AG490（100 μmol·L-1） on Lewis lung cancer cells after 24 and 48 h treatment were 37.95% and 52.99%.After incubation with 50 and 100 μmol·L-1 AG490 for 24 h,the apoptosis rates of Lewis lung cancer cells increased from 0.76% to 12.56% and 16.76%,respectively（P0.01）;typical changes in apoptotic morphology were observed by fluorescence microscopy;moreover,activation of Stat3 was inhibited,mRNA and protein levels of Stat3 and Bcl-2 were reduced.Conclusion Blocking of Jak-Stat3 signaling pathway in Lewis lung cancer cells could remarkably inhibit cell proliferation and inactivate Stat3,as well as induce cell apoptosis by the reducing of Stat3 and Bcl-2.