为探讨接受ONO-AE-248刺激后重要的表达蛋白-A1在中性粒细胞(polymorphonuelear neutrophils,PMN)非凋亡程序化细胞死亡中的的表达及意义,以进一步确定这种新型细胞死亡模式提供线索。采用Ficoll法新鲜分离健康志愿者外周静脉血PMN,与ONO-AE-248共同培养2 h后提取总RNA,并行反转录-PCR检测A1 mRNA的表达;Western blot检测A1蛋白的表达;透射电镜观察线粒体形态结果的变化。结果发现,ONO-AE-248刺激2 h时PMN A1 mRNA的表达明显上调(P〈0.01);蛋白印迹法显示A1蛋白的表达增高;透射电镜观察ONO-AE-248刺激下的早期PMN线粒体超微形态结构发生了剧烈改变。提示ONO-AE-248引起A1表达的上调可能是ONO-AE-248诱使中性粒细胞发生非凋亡性程序化死亡的根本原因之一。
To investigate the role of the expression protein A1 in non-apoptotic PCD of neutrophils induced by ONO-AE-248,human neutrophils were cultured in vitro with or without ONO-AE-248(5×10-5 mol/L)and the expression level of A1 was examined using RT-PCR and Western-blotting.The structure of mitochondria was observed by transmission electron microscope.It was found that PMN could express the mRNA and protein of A1.The expression of A1 and intensity of mitochondia in neutrophils treated with ONO-AE-248 was up-regulated or intensively changed.These results indicate that ONO-AE-248 can promote the non-apoptotic programmed cell death of PMNs in vitro.and the expression protein A1 produce lot of effects in the process of ONO-AE-248-induced neutrophil death that were different from those of control and TNF-α.