Study On The Mechanism Of Hypoxia Inducible Factor 1/Hypoxic Response Element Pathway In Rat Myocardial Ischemia/Diazoxide Postconditioning

Objective: To fathom whether the mito KATP channels were activated by the ischemic / diazoxide postconditioning,which may further activate the HIF-1/HRE pathway to mitigate myocardial ischemia reperfusion injury(MIRI).Methods: Male healthy SD rats(250-300 g)were anaesthetized by intraperitoneally injected with sodium pentobarbital and heparin,and the hearts were quickly harvested.By using Langendorff perfusion device and K-H solution,the rats' hearts were randomly divided into the following 10 groups(n=8,each group),i.e.group N,group I/R,group IPO,group IPO+2ME2,group IPO+5HD,group DPO,group DPO+2ME2,group DPO+5HD,group I/R+2ME2,group I/R +5HD.Group N,heart was perfused with K-H solution for 120 minute continually.Group I/R,after perfused heart with K-H solution for 20 minute,we stopped the perfusion for 40 minute and reperfused heart with K-H solution for 60 minute.Group IPO,after perfused heart with K-H solution for 20 minute,we stopped the perfusion for 40 minute,and then alternate ly perfused heart 10 s or stopped the perfusion 10 s for 6 rounds,at last reperfused heart with K-H solution for 58 minute.Group IPO+5HD,on the basis of group IPO,we perfused heart with 5-Hydroxydecanoate for 5 minute after stopped the perfusion for 35 minute.Group IPO+2ME2,on the basis of group IPO,we perfused heart with 2-Methoxyestradiol for 10 minute after stopped the perfusion for 30 minute.Group DPO,after perfused heart with K-H solution for 20 minute,we stopped the perfusion for 40 minute,and then perfused heart with diazoxide for 5 minute,at last reperfused heart with K-H solution for 55 minute.In the group DPO+5HD,group DPO+2ME2,group I/R+5HD and group I/R+2ME2,5-Hydroxydecanoate and 2-Methoxyestradiol were used on the basis of group DPO and group I/R respectively,and the use of two blocking agents were the same as that of group IPO+5HD and group IPO+2ME2.Except for the group N,the rest of the groups were given 4? ST.Thomas cardioplegia to attain cardiac arrest after stopping perfusion,and needed to be keep the ambient temperature at 32 ? during the whole cardiac ischemia.The following detection indexes in each group needed to be done.1.Cardiac hemodynamic indexes were recorded at the time of 20 minute perfusion and the end of reperfusion.2.At the end of perfusion,myocardial infarct size was detected by TTC staining.3.At the end of perfusion,the myocardial ultrastructure and mitochondria score of left ventricular tissue were observed under transmission electron microscope.4.At the end of perfusion,the expression of HIF-1?,HO-1,i NOS and VEGF of left ventricular tissue were detected by RT-PCR and Western blot.Results: Compared with group N,the infarct size and mitochondrial Flameng score of group I/R was significantly increased after myocardial ischemia reperfusion injury(P<0.05),and the myocardial structure was damaged by ischemia reperfusion injury,which was characterized by the destruction of the filament,formation of vacuoles,swelling of mitochondria,widening of mitochondrial cristae gap and rupture of mitochondria.The infarct size and mitochondrial Flameng score of group IPO / group DPO were significantly smaller than that of group I/R(P<0.05),at the same time,the myocardial structure of group IPO and group DPO were basically normal and the same as the group N,because there were few disruption of filament,swelling of mitochondria.In addition,both IPO and DPO could increase the m RN A expression of HIF-1? and the downstream of HIF-1/HRE pathway(HO-1,i NOS and VEGF),and also increase the protein expression of the downstream of HIF-1/HRE pathway(HO-1,i NOS and VEGF).But the myocardial protective effects and activation of HIF-1/HRE pathway related products of IPO/DPO,could be eliminated after the use of mito KATP channel specific blocker 5HD or HIF-1 alpha specific blocker 2ME2.The myocardial structure of group IPO+5HD,group IPO+2ME2,group DPO+5HD and group DPO+2ME2 were basically the same as the group I/R,because the destruction of the filament,formation of vacuoles,swelling of mitochondria,widening of mitochondrial cristae gap and the rupture of mitochondria were also observed.Conclusion: The activation of HIF-1/HRE pathway by opening mito KATP channels may get involved in the mechanism of IPO and DPO in red ucing myocardial ischemia reperfusion injury..