阿霉素诱导肺癌细胞凋亡的实验研究

Induction of apoptosis of lung cancer cells by doxorubicin

目的：建立培养肺癌细胞凋亡模型。方法：人肺癌细胞Ａ５４９在含２０μｍｏｌ／Ｌ阿霉素培养液中培养４ｈ，换用普通培养液培养，在不同时间点收集漂浮细胞，检测漂浮细胞计数，锥虫蓝排斥实验，吖啶橙染色，电镜观察，ＤＮＡ电泳，流式细胞仪分析。结果：处理的Ａ５４９细胞形态学表现为锥虫蓝拒染，细胞皱缩变形，细胞膜完整，部分呈泡状突起，细胞核染色质浓聚、边集或碎裂，凋亡小体形成。细胞ＤＮＡ电泳见梯状电泳带，流式细胞仪检测发现凋亡峰出现，符合凋亡细胞特征。凋亡细胞主要存在于漂浮细胞中，最多出现于改用普通培养液培养１０～１２ｈ。结论：阿霉素可以诱导Ａ５４９肺癌细胞发生凋亡。

Objective: To establish apoptosis model of cultured lung cancer cells. Methods: A549 human lung adenocarcinoma cells were treated in cultured fluid containing 20 μmol/L doxorubicin for 4 h. At different time points of post incubation the detached cells were harvested for analysis by counting, trypan blue exclusion, acridine orange dying, electromicroscopy, DNA agarose gel electrophoresis and flow cytometry. Results: The morphology of the treated cells was characterized by positive trypan blue exclusion, membrane budding, cytoplasmic condensation, chromatin condensation, fragmentation, margination and apoptotic body formation. Agarose gel electrophoresis analysis revealed DNA cleavage (called DNA ladder). An apoptotic peak appeared in the analysis of flow cytometry. All those results suggested that apoptosis of A549 cells was successfully induced by doxorubicin. Most of the apoptotic cells appeared in the detached cell group after 10～12 h of post incubation. Conclusion: Apoptosis of A549 lung cancer cells can be induced by doxorubicin.