目的:研究缺氧大鼠肺动脉平滑肌细胞(PASMCs) PKCα/c-fos、Bax/Bcl-2表达的变化及对其增殖和凋亡的影响.方法:将原代培养的大鼠PASMCs分为常氧组(5％CO2、21％O2、74％N2)和低氧组(5％CO2,2％ O2,93％N2)24h,48 h.通过氮蓝四唑盐(MTT)比色法测定PASMCs的增殖水平,以流式细胞仪Annexin V-FITC/PI染色法检测细胞凋亡率,并通过用RT-PCR和Western blot法检测PKCα/c-fos、Bcl/Bcl-2基因和蛋白表达的变化.实验至少重复3次,每组设置至少3个复孔.结果:低氧24 h和48 h组PASMCs增殖明显,与常氧组相比差异具有统计学意义(P＜o.01),而细胞凋亡率无显著变化.相对于常氧组,低氧24h和48 h组PASMCs PKCα、c-fos表达增强,Bax表达变化不明显,而Bcl-2表达增加.结论:低氧培养的PASMCs增殖明显,凋亡无明显变化.同时PKCα、c-fos表达增强,而抗凋亡基因Bcl-2表达增加.%Objective:To explore the effects of PKCα/c-fos,Bax/Bcl-2 on the proliferation and apoptosis of rat pulmonary arterial smooth musclecells(PASMCs) in hypoxia.Methods:The PASMCs of rats had been isolated and cultured,and then were cultured under normoxia (5％CO2、21％O2、74％N2)and hypoxia(5 ％ CO2,2％ O2,93 ％ N2) condition for 24 h and 48 h,respectively.The proliferation of PASMCs was tested by methylthiazolyltetrazolium (MTT).The changes of PASMCs apoptosis were detected by Annexin V-FITC/PI staining combined flow cytometry.RT-PCR and Western blot analysis were performed to detect the gene and protein levels of PKCα/c-fos,Bax/Bcl-2,respectively.All experiments were repeated three times with at least triplicate samples.Results:The proliferation of PASMCs in hypoxia group were stronger than that of normoxia group after 24 h and 48 h (P ＜ 0.01),while the apoptosis rate did not change significantly.Meanwhile,the higher levels of PKCα,c-fos,Bcl-2 mRNA and proteins after hypoxia for 24 h and 48 h were detected by RT-PCR and Western blot analysis,while the expression levels of Bax had no significant difference under normoxic and hypoxic conditions.Conclusion:Elevated proliferation and expression of PKCα,c-fos,Bcl-2 was observed in rat PASMCs in hypoxia,and while the apoptosis rate had no significant change.
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