Preconditioning Of Shexiang Tongxin Dripping Pills Reduces Myocardial Ischemia-reperfusion Injury In Rats By PI3K-Akt-eNOS Pathway

Objectives Shexiang Tongxin Dripping Pills can upregulate the level of nitric oxide in the blood vessel and inhibit inflammatory reaction,thus reduce myocardial ischemia injury.The aim of the present study was to test whether Shexiang Tongxin Dripping Pills preconditioning before myocardial ischemia would reduce the myocardial ischemia/reperfusion injury in rats.The probable molecular mechanism was also investigated.Methods We performed left main coronary artery ligation to establish myocardial ischemia/reperfusion model in male Sprague Dawley(SD)rats.Seventy-eight rats were randomly divided into 7 groups.Except for the sham group(n = 6),all rats in other groups(each n = 12)underwent 30 min of myocardial ischemia,followed by reperfusion of 5 min or 120 min:(1)Sham group,threading on coronary artery without ligation was done after thoracotomy;(2)Ischemia/reperfusion(Control)group,no additional intervention was given;(3)Ischemic postconditioning(Post)group,rats were subjected to 3 cycles of 10 s reperfusion/10 s ischemia before persistent reperfusion;(4)–(6)Low-,medium-and high-dose Shexiang Tongxin Dripping Pills(SX-L,SX-M,SX-H)group,1.5,3.0 or 6.0 mg/kg·d of Shexiang Tongxin Dripping Pills solution were given by gastric lavage for 2 weeks before ischemia/reperfusion;(7)High-dose Shexiang Tongxin Dripping Pills preconditioning with LY294002(SX-H + LY294002)group,0.3mg/kg of PI3K-specific inhibitor LY294002 was slowly injected into the femoral vein 15 min before reperfusion;the remaining steps were the same as in the SX-H group.The size of myocardial ischemia and infarction was evaluated by double staining using triphenyl-tetrazolium chloride and Evan's Blue.The expression and phosphorylation level of protein kinase B(Akt)and its downstream endothelial nitric oxide synthase(e NOS)in the ischemic site were measured by Western blot.The expression of Akt m RNA was measured by RT-PCR.Results 1.There was no significant myocardial ischemia or infarct in the Sham group.2.Compared with the Control group,myocardial infarct sizes in the Post group,SX-M group,SX-H group were smaller[(48.6±4.0)% vs(24.6±4.4)%,(27.2±4.5)%,(25.8±3.6)%,P<0.01],the size of myocardial infarction in the SX-L group and SX-H + LY294002 group were not decreased.There was no significant difference of the above-mentioned cardioprotective effect between the Post group,SX-M group and SX-H group.3.Compared with the Control group,Akt phosphorylation levels,e NOS phosphorylation levels and Aktm RNA expression levels in the Post group,SX-M group,SX-H group increased[p-Akt/t-Akt :(1.00±0.00)vs(2.26±0.09),(2.16±0.06),(2.23±0.11),P<0.01;p-e NOS/t-e NOS :(1.00±0.00)vs(2.09±0.07),(2.01±0.13),(2.08±0.12),P<0.01;Aktm RNA:(1.00±0.00)vs(1.41±0.02),(1.39±0.03),(1.41±0.03),P<0.01].4.Compared with the Control group,the cardioprotective effect,increased Akt and e NOS phosphorylation levels and Aktm RNA expression level in the SX-H group could be completely blocked by PI3 K specific inhibitor LY294002(P<0.01).Conclusion Preconditioning of Shexiang Tongxin Dripping Pills can reduce myocardial ischemia-reperfusion injury in rats by activating PI3K-Akt-e NOS pathway.