组蛋白去乙酰化酶抑制剂介导的Akt/Gsk3β通路激活在糖尿病大鼠心肌缺血再灌注损伤中的保护作用

Protection Effect of Histones Deacetylases Inhibitor on Ischemia/Reperfusion Induced Myocardium Injury in Diabetic Rats

目的:探讨组蛋白去乙酰化酶(HDAC)抑制剂(TSA)对糖尿病大鼠心肌缺血再灌注(MI/R)损伤是否具有保护作用及其机制。方法:SD大鼠腹腔注射链脲佐菌素(STZ)建立糖尿病大鼠模型,饲养8周。大鼠麻醉后行机械通气,结扎冠状动脉左前降支(LAD)30 min随后开放120 min以建立心肌缺血再灌注模型。TTC和Evens blue双重染色法测定梗死面积,TUNEL染色检测心肌凋亡程度,免疫印迹法检测蛋白激酶B(Akt)和糖原合酶激酶-3β(Gsk3β)的磷酸化水平以及凋亡相关蛋白Bcl-2,Bax的表达量。结果:HDAC抑制剂能够降低糖尿病大鼠的心肌梗死面积,减轻MI/R损伤所致心肌凋亡;合并给予Akt抑制剂后,TSA的保护作用被抑制;进一步研究表明HDAC抑制剂提高了高糖状态下MI/R损伤后的心肌Akt磷酸化水平,同时也提高了Akt下游蛋白p-Gsk3β的表达量。结论:Akt的激活是HDAC抑制剂发挥糖尿病MI/R损伤保护作用的关键机制,HDAC抑制剂可能是通过Akt/Gsk3β信号通路发挥保护作用。

Objective：To investigate whether histones deacetylases（HDAC）inhibitor Trichostatin A（TSA）could confer protection against ischemia/reperfasion injury in diabetic rats by using myocardium ischemia/reperfusion（MI/R）model,and probe the underlying mechanisms.Methods：Diabetes was induced by intraperitoneal injection of STZ in male Sprague-Dawley rats.8weeks later,rats were anesthetized by pentobarbital sodium,and followed tracheotomy and artificial ventilation.Silk suture was placed around the left anterior descending artery（LAD）,the ligature was tightened to occlude the LAD,after 30 min of ischemia,then 120 min reperfusion was performed by releasing slipknot.Infarct size was assessed by TTC/Evans blue dye.TUNEL positive cells were detected to measure the apoptosis degree.The proteins levels of Akt,p-Akt,Gsk3β,pGsk3β,Bax and Bcl-2were assessed by immunoblotting of myocardium lysates.Results：HDAC inhibitorcould relieve MI/R injury by decreasing infarct size of diabetic rats,reduce cell apoptosis ratio（MI/R group vs MI/R＋TSA group,P〈0.05）.It＇s notable that co-treatment with Akt inhibitor MK-2206 partly inhibited TSA-elicited protective effect（MI/R＋TSA＋MK group vs MI/R＋TSA group,P〈0.05）.Further research found TSA administration significantly elevated the expression of p-Akt and phosphorylated its downstream targets GSK-3β（MI/R group vs MI/R＋TSA group,P〈0.05）.Conclusion：Histones deacetylases inhibition triggers protective effects against MI/R injury through Akt/Gsk3βmodulated mitochondrial apoptotic pathways in diabetic rats.