硝基化修饰导致硫氧还蛋白不可逆的丧失了对心脏的保护作用(英文)

Thioredoxin nitration causes irreversible loss of its cardioprotective property

目的:本研究旨在明确过氧亚硝酸阴离子是否可以导致硫氧还蛋白(thioredoxin,TRX,一种新的抗氧化和抗凋亡蛋白质)硝基化修饰失活,从而促进缺血/再灌注心肌细胞凋亡。方法和结果:硫氧还蛋白中加入100μmol/L SIN-1(过氧亚硝酸阴离子供体)离体孵育可导致TRX发生硝基化,提示心肌组织内TRX也对硝基修饰敏感。为进一步确定TRX硝化是否改变其心肌保护作用,采用缺血30 min,再灌注3 h(凋亡检测)或24 h(心肌梗死范围)小鼠缺血/再灌注模型,分别于再灌前10 min给予磷酸盐缓冲液(PBS,对照组),TRX或硝化TRX。TRX治疗可以显著减少缺血/再灌注导致的心肌细胞凋亡和心肌梗死范围(P<0.01);而在体外用SIN-1孵育TRX,则完全阻断了其心肌保护作用;此外,用SIN-1和血红蛋白(清除NO)或SIN-1和超氧化物岐化酶(SOD,清除过氧化物)同时孵育TRX,其心肌保护作用则完全保留。结论:上述结果表明,SIN-1通过产生过氧亚硝酸阴离子使TRX失活(丧失对心脏的保护作用),提示过氧亚硝酸阴离子可能通过使TRX硝基化失活而发挥促凋亡作用。

AIM： The present study determined whether peroxynitrite may cause nitrative inactivation of thioredoxin （Trx） , a novel anti-oxidant and anti-apoptotic protein, thus facilitating myocardial apoptosis. METHODS and RESULTS： Exposure of recombinant Trx to 100 M of SIN-1, a peroxynitrite donor, resuited in significant Trx nitration as determined by Western blotting, indicating that cardiac Trx is sensitive to nitrative modification. To determine whether nitration of Trx may alter its cardioprotective effect, adult mice were subjected to 30 minutes myocardial ischemia and 3 hours （apoptosis） or 24 hours （ infarct） of reperfusion and treated with vehicle, native or nitrated human Trxl （10 rain before reperfusion）. Administration of native Trxl significantly reduced post-ischemic myocardial apoptosis and decreased infarct size （ P 〈0.01 ）. However, in vitro incubation of thioredoxin with SIN-1 （ to nitrate Trx1 ） prior to its administration completely abolished thioredoxing cardioprotective effect. Moreover, when Trxl was incubated with SIN-1 in the presence of hemoglobin （ to scavenge NO） or SOD （ to scavenge superoxide）, the cardioprotective effect of Trxl was fully preserved. CONCLUSION： These results indicate that SIN-1 inactivates thioredoxin in a peroxynitrite-dependent fashion and suggest that peroxynitrite may exert its pro-apoptotic effect by nitrative inactivation of thioredoxin.