Role Of MitoK_ATP Channel In The Combination Of Rosuvastatin And Ischemic Postcondition On Myocardium Against T2DM Rats Ischemia-reperfusion Injury In Vivo

Background: Some clinical and animal experimental research showed that, statins preconditioning/ postconditioning reduce the ischemia-reperfusion injury significantly,but whether statins also reduce the ischemia-reperfusion injury in DM rats is not known. Recent reserch revealed that, rosuvastatin may restore the protection of ischemic postconditioning(IPO) to T2DM or hyperglycemia rats in cardic ischemia-reperfusion injury, futhermore, those reserch concluded that, this drug may get the pleasant effect throuth PI3K-AKT-eNOS pathway. However, the exact mechinasm of how PI3K-AKT-eNOS finally exert protection is unknown.Objective: The current study was conducted to observe the protection of RPO+IPO to IRI in T2DM rats ,and to investigate the following questions : 1.whether RPO+IPO could attenuate IRI in T2DM rats?2.If the treatment could produce the production,which one plays the major role in the production, mitoK_ATP channel or sarcK_ATP?3. To IRI T2DM rats in vivo, whether RPO+IPO+non-specific channel openers gets better production than RPO+IPO?4. Does there exist the relationship between the protection of RPO+IPO to IRI in T2DM rats and the expression of eNOS.Methods:100 healthy Wistar male rats were created as T2DM, weight 200±10g, and then 81 ones were randomly allocated into 9 groups (n=9,each group);(1)Sham group, threaded without occluding; (2)IRI group, the Left anterior descending artery(LAD) was occluded for 45mins followed by reperfusion (R) for 120mins without other interventions; (3)RPO +IPO group, LAD was occluded for 45mins, tnen pushed rosuvastatin 10mg/kg in external jugular vein slowly 3mins before R, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (4)RPO+IPO+5-HD group(5-HD group), LAD was occluded for 45mins, tnen pushed 5-HD10mg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (5)RPO+IPO+diazoxide group(diazoxide group), LAD was occluded for 45mins, tnen pushed diazoxide 12.5mg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (6)RPO +IPO+ HMR-1098 group(HMR-1098 group), LAD was occluded for 45mins, tnen pushed HMR-1098 5mg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (7)RPO+IPO+Cromakalim group(Cromakalim group), LAD was occluded for 45mins, tnen pushed Cromakalim 56μg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (8)RPO+IPO+glibenclamide group(glibenclamide group), LAD was occluded for 45mins, tnen pushed glibenclamide 5mg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last; (9)RPO+IPO+ nicorandil group(nicorandil group), LAD was occluded for 45mins, tnen pushed nicorandil0.3mg/kg,rosuvastatin 10mg/kg in external jugular vein slowly 10min and 3mins before R respectively, then treated three times of 10s R and 10s I before R,and followed by R for 120mins at last;Infarct size was evaluated by TTC,ultrastructure was observed by electron microscope,expression of phosphorylate eNOS,and total eNOS was evaluated by Western—blot,plasma cTnT were also measured.Results(1)As compared with IRI group, the myocardial infarct size was decreased significantly in RPO +IPO group and diazoxide group, HMR-1098 group, Cromakalim group, nicorandil group(P<0.05); The myocardial infarct size in 5-HD group,Glibenclamide group was similar to IRI group,and the difference was not significant (P>0.05);but The myocardial infarct size in 5-HD group,Glibenclamide group was increased significantly than RPO+IPO group and diazoxide group, HMR-1098 group, Cromakalim group, nicorandil group(P<0.05).(2)TEM revealed that myocardial cell have a large number of mitochondria in Sham group,and mitochondria structure was normal,the mitochondria had clear and regular cristae. while the mitochondria in IRI group,5-HD group and Glibenclamide group was swelling, cristae blurred , lost with a hardly visible or disrupted membrane for some mitochondrion, and vacuoles formation was observed. In RPO+IPO group , diazoxide group, HMR-1098 group, Cromakalim group and nicorandil group , the structure of most mitochondria maintained as originally, loss of matrix granule and swelling of mitochondria was found by chance, low electron density and dissolved mitochondrial crista was obseard only in small number of mitochondria, but mitochondrial membrane was complete(.3)As compared with IRI group, the level of cTnT in RPO+IPO group and diazoxide group, HMR-1098 group, Cromakalim group, nicorandil group was decreased significantly(P<0.05); The level of cTnT in 5-HD group,Glibenclamide group was similar to IRI group,and the difference was not significant (P>0.05);but the level of cTnT in 5-HD group,Glibenclamide group was increased significantly than RPO+IPO group and diazoxide group, HMR-1098 group, Cromakalim group, nicorandil group(P<0.05)(.4)As compared with Sham group, IRI group, the expression of phosphorylate eNOS in RPO+IPO group,5-HD group and diazoxide group was higher significantly(P<0.05); the expression of phosphorylate eNOS in IRI group was a little lower than Sham group,the difference was significan(tP<0.05);the expression of total eNOS was similar in all groups,there was no significant difference(P>0.05).Conclutions: RPO+IPO: 1.could attenuate IRI in vivo T2DM rats heart significantly. 2.mitoK_ATP channel but not sarcK_ATP channel plays the major role in the protection. 3. RPO +IPO+non-specific channel openers can't get more protection than RPO+IPO. 4. in the process of RPO+IPO, there exist no relationship between the intervention to mitoK_ATP and the expression of p-eNOS.