促红细胞生成素对新生大鼠心脏成纤维细胞氧化应激时一氧化氮合酶系统的影响  

Effects of erythropoietin on nitric oxide synthase system in neonate rat cardiac fibroblasts after oxidative stress

作　　者：张新金[1] 马业新[1] 文渊[1] 徐雪晶[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院心内科,武汉430030

出　　处：《临床心血管病杂志》2008年第11期863-866,共4页Journal of Clinical Cardiology

摘　　要：目的:探讨促红细胞生成素(EPO)对心脏成纤维细胞(CFs)氧化应激时一氧化氮合酶(NOS)系统的影响,以及PI3-K/Akt信号途径在其中的作用。方法:应用胰酶和胶原酶双酶法分离培养新生大鼠CFs,EPO、过氧化氢(H2O2)和PI3-K抑制剂LY294002不同因素干预。化学酶法检测CFs培养液中的NO浓度以及总NOS和其亚型的活性。Western blot检测Akt、p-Akt、eNOS、iNOS蛋白的表达。结果:氧化应激使CFs中iNOS的表达显著上调,活性增强;eNOS蛋白的表达明显下降。同时CFs培养液中NO的合成增加显著,达到(25.94±2.57)μmol/L,是对照组的4倍多(P<0.01)。EPO显著抑制CFs氧化应激时iNOS蛋白的表达,同时上调eNOS蛋白,总的NO浓度也显著下降。Akt磷酸化形式p-Akt的表达水平也明显提高。PI3-K抑制剂LY294002能阻断EPO促进eNOS和p-Akt蛋白表达的作用,但EPO抑制iNOS的作用不能完全被阻断。结论:EPO能促进新生大鼠CFs氧化应激时eNOS蛋白的合成,抑制iNOS蛋白的表达。其促进eNOS的表达是通过激活PI3-K/Akt细胞信号途径来实现,抑制iNOS的机制有待进一步探讨。Objective：The aim is to investigate the effects of erythropoietin on nitric oxide synthase system expression and activity in neonate rat cardiac fibroblasts after oxidative stress and the role of P13-K/Akt singaling pathway. Method：Neonatal rat cardiac fibroblasts （CFs） were isolated by collgenase and trypsinase and differential attachment technique. EPO, hydrogen peroxide （H2O2） and LY294002, an inhibitor of P13-K, were added in re lated group respectively. The levels of nitric oxide （NO） and the activites of NOS and its isoforms were measured by chemical enzymic method. The expressions of Akt, p-Akt, eNOS and iNOS were detected by Western blot. Resuit：Oxidative stress markedly increased the expression of iNOS and decreas the expression of eNOS. Meanwhile, the production of NO in CFs culture fluid increased obviously, reaching to （25.94±2.57）umol/L, which was four times more as great as that of control group （P〈0. 01）. EPO predominantly suppressed the expression of iNOS and the level of NO after oxidative stress. Meanwhile, the expressions of phosphated Akt, p-Akt, and eNOS were all been up regulated by EPO. The effects of up-regulating eNOS and p-Akt by EPO were blocked by inhibitor of P13-K LY294002, but the suppression effect on expression of iNOS in CFs induced by EPO was not blocked completely. Conclusion： EPO down-regulates the expression of iNOS and up-regulates the expression of eNOS in neonatal rat CFs induced by oxidative stress. The up-regulating effect on the expression of eNOS is implemented by activating the P13-K/Akt signaling pathway and the mechanism of suppressing the expression of iNOS is still unknown.

关 键 词：氧化应激 促红细胞生成素 一氧化氮合酶 心脏成纤维细胞 

分 类 号：R542.2[医药卫生—心血管疾病]