多柔比星诱导肺腺癌细胞凋亡机制的探讨

Mechanism of apoptosis of lung adenocarcinoma cells induced by doxorubicin

目的:探讨多柔比星(ADM)诱导肺腺癌细胞系A549细胞凋亡机制。方法:应用噻唑蓝(MTT)比色法检测ADM对A549细胞增殖抑制作用,透射电镜观察细胞形态学改变;细胞电泳仪测定A549细胞表面电荷的变化;流式细胞仪(FCM)测定A549细胞Fas、Bcl-2蛋白水平变化;比色法检测Caspase-3活性的变化。结果:ADM可抑制细胞增殖,作用24、48及72h的IC50值分别为5.2、4.8和4.2mg/L,电镜下见到明显的凋亡细胞;细胞表面电位在4h开始下降,4～24h期间下降最明显,>24h下降趋势降低;Fas蛋白表达明显上调,Bcl-2蛋白表达下降,Caspase-3活性明显增强。结论:ADM能抑制并诱导A549细胞凋亡,细胞表面电荷下降可能是ADM诱导细胞凋亡的早期事件,其作用可能与上调Fas蛋白、下调Bcl-2蛋白表达有关。

OBJECTIVE： To study the mechanism of apoptosis of A549 ceils induced by doxorubiein and explore its possible mechanisms. METHODS： MTT assay and Wright-Giemsa staining were used to examine apoptosis in A549 cells. The changes of cell surface charge were observed by the elctrophoresis mobility. The expression levels of Fas and Bcl 2 antigens were measure with FCM. Colorimetric assay were employed to detect the activation of Casepase-3. RESULTS： ADM inhibited the proliferation of A549 cells and IC50 （s） at 24, 48 and 72 h were 5.2, 4.8 and 4.2 mg/L respectively, and the typical apoptosis morphology of A549 cells was observed through transmission electron microscopy after the treatment by ADM. The electrophoresis mobility decreased with the passing of the time. The decrease appeared at about 4 h and greatly enhanced in 4--24 h, which occurred before the typical changes of cells morphology of apoptosis. After 24 hours, there was a reducation in electrophoresis mobility. The expression of Fas antigens increased and Bcl-2 antigens decreased. Caspase-3 was also activated by ADM. CONCLUSIONS： Doxorubicin can inhibit the proliferation and induce the apoptosis of A549 cells. The changes of cell surface charge can reflect the apoptosis, especially in early time. It is an early event of apoptotic A549. The expression levels of Fas and Bcl-2 antigens may be relative to the changes of cell surface charge.