The Role Of DIDS On Rats Myocardial Ischemia-reperfusion-induced Apoptosis And The Regulation Of PI3K/Akt In Vivo

Acute myocardial infarction (AMI) is the serious stage of the coronary heartdisease (CHD). The decline of effective systolic function and the loss ofmyocardial cells may lead to the occurrence of sudden cardiac death anddecreased quality of life, thus, it is harmful to human health and life. Therefore,the methods of reducing the acute myocardial infarction size, improving heartfunction effectively in order to rescue the patient's life are particularly important.The most effective treatment of acute myocardial infarction is re-establishingcoronary blood flow, including thrombolysis, percutaneous coronary angioplasty(PTCA) or coronary artery bypass grafting. However, reperfusion of ischemicmyocardium can itself result in exacerbating the cellular injury and dysfunctionsustained during the ischemia period. It has been reported that apoptosis is animportant mechanism in I/RI. Ischemic preconditioning and postischemiapreconditioning can inhibit myocardial apoptosis, reduce the myocardial infarctsize and decrease the incidence of the cardiac dysrhythmia. Therefore, it is of great research importance to look for a new anti-ischemic / reperfusion injury ofmyocardial apoptosis drugs and methods to explore its possible mechanism4,4'-Diisothiocyanostilbene-2, 2'-disulfonic acid (DIDS) is a broadspectrum of the chloride channel blocker. Our previous studies have demonstratedthat it plays an important role in the protection of myocardial ischemiareperfusion-induced cardiac dysrhythmia, the myocardial necrosis and thedisorder of cardiac function in vivo.However, what is its mechanism? It was notclear yet. Our research is to investigate the role of DIDS on rats myocardialischemia-reperfusion-induced apoptosis and the regulation of PI3K/Akt in vivo.Aim1. To establish rat I/R infarction model, to observe the effect of DIDS on themyocardial ischemia-reperfusion-induced infarct size and myocardialapoptosis.2. To observe the expression of PI3K/Akt signaling molecule and the regulationof DIDSMethods1. The preparation of rat heart ischemia-reperfusion infarction model andexperiment group: Routinly established rat heart I/R infarction model. Thehealthy male SD rats were divided into 3 groups randomly,6 rats in erverygroup. I/R groups: model group, ischemic 30min/reperfusion 4h, duringreperfusion the saline was injected via the femoral vein; I/R+DIDS groups: therat model was the same as I/R groups, during reperfusion DIDS (14mg/kg)was injected via the femoral vein; LY294002 pretreatment 30min+I/R+DIDSgroup: pre-ischemia 30min, LY294002 (0.3mg/kg)was injected via thefemoral vein, the rat model was the same as I/R+DIDS groups. Saline, DIDSand LY294002 are given by the program-controlled micro-pump (4ml/kg/h). 2. Measurement of myocardial infarct size: Evan`s blue-TTC-methodDetection of cardiomyocyte apoptosis: TUNEL-method3. Detection PI3K/Akt signaling molecule: Western blottingResultResults1. Compare to I/R group, in DIDS group MI and AI were significantly attenuated,and were 38.8%±7.7% vs 54.2%±10.8% (P<0.01, n = 6) and 8.9%±1.8%vs 17.6%±3.5% (P<0.01, n = 6) respectively. DIDS can inhibit thecardiomyocyte apoptosis and reduce the infarct size.2. The expression of p-Akt was increased in DIDS group, 2.5 times higher thanin I/R group (P<0.01,n=6). Pretreatment with LY294002 abolished DIDSinducedincreases in p-Akt (P>0.05, n=6), and completely abrogated theDIDS-induced IS/AAR [52.4%±10.4 % vs 54.2%±10.8 %, P>0.05] and antiapoptotic(18.7%±3.7 % vs 17.6%±3.5%, P>0.05,n=6) effect. But, there is nosignificant change in the level of expression of Akt.Conclusion1. myocardial M/I lead to Myocardial I/R increased, apoptosis increased and thelevels of Akt phosphorylation reduced.2. DIDS can inhibit I/R-induced myocardial apoptosis and reduce myocardialinfarct size.3. DIDS can reduce ischemia-reperfusion myocardial infarct size and inhibitmyocardial apoptosis by activating PI3K/Akt signaling pathway.