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目的:本研究旨在阐明硫氧还蛋白(Trx)心肌保护作用的机制。方法:采用成年雄性小鼠心肌30min缺血/3h再灌注模型,再灌注前10min随机给予磷酸缓冲液(PBS,对照)或重组人Trx(hTrx,2mg/kg)腹腔注射。在再灌注终末,摘取实验动物的心脏,分别检测心肌细胞凋亡和心肌组织中硝基酪氨酸的含量。结果:再灌注前给予Trx可以显著减少缺血/再灌注导致的心肌细胞凋亡(P<0.01);与对照组相比,Trx组心肌组织中硝基酪氨酸的含量显著减少(4.8±0.53比9.5±0.86pmol/mg蛋白质,P<0.01),提示Trx可能通过阻断蛋白硝基化和随后的细胞凋亡而发挥心脏保护作用。为了寻找Trx可以阻断蛋白硝基化的直接证据,在培养的成年心肌细胞给予SIN1(过氧亚硝酸阴离子供体)50μmol处理、或SIN1和hTrx(500nmol)共同处理后,观察Trx在过氧亚硝酸阴离子诱导的细胞凋亡中的作用。结果表明SIN1处理导致心肌细胞凋亡显著增加(P<0.01),Trx可以显著减少SIN1导致的心肌细胞凋亡(P<0.01)。结论:硫氧还蛋白,一种新的抗凋亡和心肌保护分子,通过阻断过氧亚硝酸阴离子导致的蛋白硝基化和随后的细胞凋亡而发挥心脏保护作用。
Aims: This study aimed to elucidate the mechanism of cardioprotection of thioredoxin (Trx). Methods: The myocardium of adult male mice were reperfused with 30 min ischemia / 3 h reperfusion. Phosphate buffer (PBS, control) or recombinant human Trx (hTrx, 2 mg / kg) were injected intraperitoneally 10 min before reperfusion. At the end of reperfusion, the hearts of experimental animals were harvested for the detection of myocardial apoptosis and the content of nitrotyrosine in myocardium. Results: Trx before reperfusion could significantly decrease the apoptosis of myocardial cells induced by ischemia / reperfusion (P <0.01). Compared with the control group, the content of nitrotyrosine in myocardial tissue of Trx group was significantly decreased (4.8 ± 0.53 vs 9.5 ± 0.86 pmol / mg protein, P <0.01), suggesting that Trx may play a cardioprotective role by blocking protein nitration and subsequent apoptosis. In order to find direct evidence that Trx can block protein nitration, Trx was observed in peroxisomes after treatment with 50 micromol of SIN1 (peroxynitrite anion donor) or co-treatment of SIN1 and hTrx (500 nmol) in cultured adult cardiomyocytes Nitrite anion-induced apoptosis in the role. The results showed that the apoptosis of cardiomyocytes induced by SIN1 was significantly increased (P <0.01), Trx significantly reduced the apoptosis of cardiomyocytes induced by SIN1 (P <0.01). Conclusion: Thioredoxin, a novel anti-apoptotic and cardioprotective molecule, exerts cardioprotective effects by blocking protein nitration and subsequent apoptosis induced by peroxynitrite.