A549/DDP细胞的抗凋亡特性与其pH_i和[Ca^(2+)]_i变化相关

Changes of the pH_i and [Ca^(2+)]_i Are Responsible for the Anti-apoptotic Character of A549/DDP Cells

以临床用药剂量 (30 μmol/L)的顺铂处理敏感的人肺腺癌细胞A5 49和抗顺铂的A5 49/DDP细胞 ,在相同条件下培养 .对培养不同时间的两株细胞DNA琼脂糖凝胶电泳分析的结果表明 ,前者在培养 12h后即呈现DNA梯子 ,而后者在培养 48h后却未见凋亡特征 .用流式细胞计检测其凋亡峰也获得相似的结果 .当测定与凋亡相关的生物化学和生物物理变化时发现对顺铂敏感的A5 49细胞的线粒体膜电势显著降低 ,胞内更加酸化且胞内钙离子浓度升高 ;而抗顺铂药物的A5 49/DDP细胞的线粒体膜电势和pH值却维持在相对较高的水平 ,而其胞内钙离子浓度随培养时间的延长下降 .这些结果表明 ,抗顺铂的人肺腺癌A5 49/DDP细胞的抗凋亡特性与其胞内的相对碱化和较高的线粒体膜电势 ,以及胞内钙离子浓度的降低相关 .这些特性可能参与了A5

Human lung adenocarcinoma A549 cells and cisplatin-resistant A549/DDP cells were treated with clinically relevant doses of cisplatin (30 mu mol/L) and then further cultured under the same conditions. DNAs of both cell lines were extracted and subjected separately to agarose get electrophoresis. Results showed that DNA ladders could be seen in A549 cells cultured for 12 h, while no,apoptotic character appeared in cisplatin-resistant A549/DDP cells even after being cultured for 48 hour. This difference between two cell lines was further confirmed by apoptotic peaks measured with flow cytometry. Biochemical and biophysical experiments indicated that the mitochondrial membrane potential and pH(i) of cisplatin-sensitive A549 cells decreased significantly, whereas the intracellular free Ca2+ concentration increased greatly with the culture time. But the mitochondrial membrane potential and pH(i) remained at a relatively high level and the intracellular free Ca2+ concentration was reduced gradually with the culture time for cisplatin-resistant A549/DDP cells. It would be suggested that the anti-apoptotic character of A549/DDP cells is related with the relative intracellular basification, the persistence of mitochondrial membrane potential and decrease in intracellular free Ca2+ concentration which would be responsible for the resistance of A549/DDP cells to cisplatin.