摘要
目的通过研究促红细胞生成素(EPO)联合应用粒细胞集落刺激因子(G-CSF)对缺氧心肌细胞的影响,探讨其对缺氧心肌细胞发挥保护作用可能的分子机制。方法采用95%N2+5%CO2气体充分置换的1%胎牛血清DMEM培养液培养24 h,建立缺氧心肌细胞模型。将心肌细胞分为5组,对照组、缺氧组、缺氧+EPO组、缺氧+G-CSF组和缺氧+EPO+G-CSF联合给药组。建立缺氧模型24 h后收集5组心肌细胞标本。采用Northern-blot方法检测各组心肌细胞中的Bcl-2、Bax、Caspase-3 mRNA的表达,采用Western-blot方法检测各组心肌细胞中的Bcl-2、Bax、细胞色素c(Cyt C)、磷酸化信号转导与转录活化因子3(p-STAT-3)、Caspase-3蛋白的表达。结果成功建立了心肌细胞缺氧模型。与缺氧组相比:EPO、G-CSF和联合给药组明显提高缺氧心肌细胞的存活率,减少凋亡率和总死亡率(P<0.01)。与缺氧组相比:EPO组和G-CSF组Bcl-2表达增加(P<0.01),Bax、Caspase-3表达减少(P<0.01),联合治疗组作用最明显,强于单独药物治疗组(P<0.01),EPO和G-CSF组之间无明显差异(P>0.05);Cyt C表达在EPO组、G-CSF组和联合给药组明显降低(P<0.01);三组之间无明显差异(P>0.05);P-STAT-3蛋白表达在联合给药组明显增高(P<0.01)。结论 EPO联合G-CSF治疗,可能通过激活线粒体途径和Janus激酶/信号转导和转录激活子(JAK/STAT)途径联合发挥更强的抗凋亡作用。
Objective To evaluate the mechanism of erythropoietin （EPO） in combination with granulocyte- colony stimula- ting factor （ G - CSF） on hypoxia cardiomyocytes protection. Methods The air in the cultural medium was changed by 95% N2 and 5% CO2 for hypoxia cardiomyocytes. Cardiomyocytes were divided into five groups： normal cardiomyocytes, hypoxia cardiomyocytes, hypoxia cardiomyocytes treated with EPO, hypoxia cardiomyocytes treated with G - CSF, and hypoxia cardiomyocytes treated with EPO in combination with G -CSF. Twenty four hours after hypoxia, cardiomyocytes were collected. Bcl -2, Bax, Caspase -3 mRNA level were determined by Northern - blot analysis and Bcl - 2, Bax, cytochrome C, P - STAT - 3, Caspase - 3 protein level were deter- mined by Western - blot analysis in each group. Results Model of hypoxia cardiomyocytes was established successfully. Compared with that in the hypoxia group, the survival rate of hypoxia cardialmyocytes was higher and the apoptotic rate and total necrotic rate were lower significantly in EPO, G -CSF and combination groups （ P 〈 0.01 ）, meanwhile, the expression of Bcl -2 was upregulated, Bax and Caspase - 3 were downregulated significantly in group EPO, G - CSF and combination （ P 〈 0.01 ）, and these changes were more remarkable in the combination group than that in the EPO or G - CSF group. Cytochrome C was much downregulate in EPO, G - CSF and combination group （ P 〈 0.05 ）, P - STAT - 3 expression was upregulated in combination group （ P 〈 0.05 ）. Conclusion For hypoxia cardiomyocyte, combination treatment of EPO and G - CSF shows better protective effects by activating both cbondriosome and JAK - STAT signaling pathways.
出处
《中国体外循环杂志》
2012年第4期227-231,共5页
Chinese Journal of Extracorporeal Circulation
基金
国家科技支撑计划(2009BAI86B04)
