DIDS对十字孢碱诱导心肌细胞凋亡中磷脂酰肌醇3激酶/蛋白激酶B信号转导的影响

目的探讨氯离子通道阻断剂DIDS对十字孢碱诱导心肌细胞凋亡与磷脂酰肌醇3激酶/蛋白激酶B信号及其下游分子一氧化氮合酶/一氧化氮的关系。方法实验分为对照组、十字孢碱组、DIDS组、LY294002(特异性磷脂酰肌醇3激酶抑制剂)组和L-NAME(非特异性一氧化氮合酶抑制剂)组。在十字孢碱诱导心肌细胞凋亡模型上,观察DIDS对心肌细胞存活率、凋亡和磷脂酰肌醇3激酶/蛋白激酶B及其下游分子一氧化氮合酶/一氧化氮的影响。结果与十字孢碱组比,DIDS明显改善了细胞存活率,提高了细胞磷酸化蛋白激酶B活性2.1倍(P<0.01),增加了一氧化氮合酶和磷酸化一氧化氮合酶的水平和一氧化氮水平(P<0.01);LY294002预处理完全抑制了磷酸化蛋白激酶B、一氧化氮合酶和磷酸化一氧化氮合酶水平的升高及升高的一氧化氮,完全阻断了DIDS的抗细胞凋亡作用;L-NAME预处理也使升高的一氧化氮水平下降,但仅部分阻断了DIDS的细胞保护作用。结论DIDS通过激活磷脂酰肌醇3激酶/蛋白激酶B信号通路发挥其抑制十字孢碱诱导的心肌细胞凋亡作用。

The Effect of DIDS on Phosphatidylinositol 3'-Kinase/Proteinase B Signal Transduction of Staurosporine-Induced Cardiomyocyte Apoptosis

Aim To explore the effect of chloride channel blocker DIDS on cell signaling pathway phosphatidylinositol 3'-kinase/proteinase B (PI3K/Akt) and its downstream molecules endothelial nitric oxide synthase (eNOS) and nitric oxide (NO) in staurosporine (STS)-treated cardiomyocyte apoptosis. Methods Neonatal rat cardiomyocytes were exposed to STS in the presence or Absence of DIDS. Cell viability,apoptosis and expressions of Akt,phospho-Akt (p-Akt),eNOS,phospho-eNOS (p-eNOS) and NO production were determined. Results DIDS markedly improved cell viability on STS-exposed cardiomyocytes. DIDS resulted in a 2.1-fold increase of p-Akt over control levels,prevented the reduction in eNOS expression and phospho-eNOS levels induced by STS and significantly increased NO production (all P<0.01). Pretreatment with LY294002,a selective PI3K inhibitor,abolished DIDS-induced increases in p-Akt,eNOS,p-eNOS and NO production,and completely abrogated the DIDS-induced anti-apoptotic effect (P<0.01). Pretreatment with L-NAME,a non-selective NOS inhibitor similarly inhibited the increased NO but only partly abolished protective effects of DIDS (P<0.05). Conclusion DIDS inhibits STS-induced cardiomyocyte apoptosis via activating PI3K/Akt signaling pathway.