促红细胞生成素对乳鼠心肌细胞肥大及P13K/Akt-eNOS信号转导通路的影响

目的 探讨促红细胞生成素(EPO)对血管紧张素Ⅱ(AngⅡ)诱导的肥大心肌细胞的影响,以及磷脂酰肌醇3激酶(PI3K)/丝氨酸苏氨酸激酶(Akt)-内皮型一氧化氮合酶(eNOS)信号转导通路在其中的作用.方法 分离乳鼠心肌细胞,利用AngⅡ诱导建立心肌细胞肥大模型,以心肌细胞表面积和心钠素(ANF)mRNA表达作为心肌细胞肥大观察指标.观察不同浓度EPO对肥大心肌细胞的影响,并利用PI3K抑制剂LY294002和一氧化氮合酶抑制剂L-NAME对其相关机制进行探讨,I司时对细胞培养液中一氧化氮(NO)浓度进行检测,蛋白免疫印迹法检测磷酸化Akt(p-Akt)、Akt、磷酸化eNOS(p-eNOS)和eNOS蛋白表达情况.结果 20 U/ml EPO能抑制由AngⅡ诱导的心肌细胞肥大,表现为心肌细胞表面积和ANF mRNA表达均减少(P＜0.05).EPO能激活Akt,促进eNOS及p-eNOS表达增加(均P＜0.05),并使NO合成增加(P＜0.01).LY294002和L-NAME能逆转EPO的抗心肌细胞肥大作用,减少NO产最(P＜0.05).蛋白免疫印迹法榆测显示,LY294002能够抑制EPO对p-Akt、p-eNOS和eNOS蛋白表达的促进作用,而L-NAME能抑制eNOS的磷酸化(均P＜0.05).结论 EPO能够抑制AngⅡ诱导的心肌细胞肥大,该作用可能是通过激活P13K/Akt信号转导通路,促进eNOS表达与活化,从而促进NO的合成来实现的.

Erythropoietin inhibits angiotensin Ⅱ induced cardiomyocyte hypertrophy in vitro via activating PI3K/Akt-eNOS pathway

Objective To explore the effect of erythropoietin (EPO) on angiotensin Ⅱ (Ang Ⅱ) induced neonatal rat cardiomyocyte hypertrophy and the association with PI3K/Akt-eNOS signaling pathway. Methods Cardiomyocytes were isolated from new-born Sprague-Dawley rats and stimulated by Ang Ⅱ in vitro. The cell surface area and mRNA expression of atrial natriuretic factor (ANF) of cardiomyocytes were determined in the presence and absence of various concentrations of EPO, phosphatidylinositol 3'-kinase (PI3K) inhibitor LY294002 and nitric oxide synthase (NOS) inhibitor L-NAME. Intracellular signal molecules, such as Akt, phosphorylated Akt, eNOS and phosphorylated eNOS protein expressions were detected by western blot. Nitric oxide (NO) level in the supernatant of cultured cardiomyocytes was assayed by NO assay kit. Results EPO (20 U/ml) significantly inhibited Ang Ⅱ induced cardiomyocyte hypertrophy as shown by decreased cell surface area and ANF mRNA expression (all P ＜0.05). EPO also activated Akt and enhanced the expression of eNOS and its phosphorylation (all P ＜ 0.05), increased the NO production (P ＜0.01). These effects could be partially abolished by cotreatment with LY294002 or L-NAME (all P ＜ 0.05). Conclusion EPO attenuates Ang Ⅱ induced cardiomyocytes hypertrophy via activating PI3K-Akt-eNOS pathway and promoting NO production.