| Objectives: The study aimed to(1)explore the effect of uncoupling proteins 2(UCP2)overexpression on myocardial ischemia/reperfusion(I/R)injury in diabetics and the specific regulatory mechanism for mitochondrial function;(2)To clarify the specific mechanism of dexmedetomidine(Dex)in alleviating myocardial I/R injury in diabetes by regulating UCP2,so as to provide reference for clinical prevention and treatment of myocardial I/R injury in diabetic patients;(3)To verify the effect of Dex on myocardial I/R injury in patients with diabetes from the clinical level.Methods: Part I: The SPF grade C57BL/6 male mice were selected to establish the type II diabetic model.Myocardial I/R injury model was established by ligating the left anterior descending coronary artery of diabetic mice for 60 minutes of ischemia followed by 120 minutes of reperfusion.Diabetic mice were divided into five groups: Sham group,I/R group,I/R+UCP2 inhibitor(Genipin)group,I/R+AAV9 group and I/R+AAV9-UCP2 group.The expression of UCP2 protein in myocardial tissue was detected by Western blot.The expression of UCP2 m RNA in myocardial tissue was detected by q PCR.Cardiac function was assessed by echocardiography.The concentration of c Tn I in serum and the content of MDA and the activity of SOD in myocardial tissue were detected by ELISA.The myocardial infarct size was determined by Evans Blue/TTC.The morphological structure of myocardial tissue was observed by HE staining.The ultrastructure of myocardial mitochondria was observed by transmission electron microscope.The myocardial mitochondrial membrane potential(ΔΨm)and ROS production were detected by flow cytometry.The activity of F1F0-ATP synthase and ATP content were detected by microplate reader.Part II: Diabetic mice were divided into five groups: Sham group,I/R group,I/R+Genipin group,I/R+Dex group and I/R+Dex+Genipin group.Cardiac function was assessed.The concentrations of c Tn I,TNF-α,IL-6 and IL-10 in serum were detected.The content of MDA and the activity of SOD in myocardial tissue were detected.The myocardial infarct size was measured.The morphological structure of myocardial tissue and the ultrastructure of myocardial mitochondria were observed.The ΔΨm,the opening of m PTP,ROS production and ATP content in cardiomyocytes were detected.The apoptosis of cardiomyocytes were detected by TUNEL kit.The expression of IκBα,p-IκBα,NF-κB p65,p-NF-κB p65,BNIP3,Cyt-C,Cleaved caspase-3 and UCP2 in myocardial tissue were detected.The expression of UCP2 m RNA was detected.Part III: A total of 75 patients with type II diabetes mellitus scheduled for elective cardiac valve replacement under cardiopulmonary bypass(CPB)were randomly divided into Control group,Dex-post group and Dex group.HR and MAP were recorded at 5 minutes after entering the operating room(T0),before aortic cross-clamping(T1),at the end of CPB(T2),immediately after the end of surgery(T3),6hours after aortic unclamping(T4)and 24 hours after aortic unclamping(T5).The concentrations of c Tn I,MDA,TNF-α and IL-6 and the activity of SOD in serum were measured at T0,T2,T4 and T5.The type of surgery,CPB time,ascending aortic occlusion time,surgery time,average intraoperative blood glucose and the incidence of spontaneous recovery of heart beat,hypertension,hypotension,and bradycardia during the surgery were recorded.The postoperative duration of mechanical ventilation,length of ICU stay and incidence of cardiovascular complications were also recorded.Results: Part I:Compared with Sham group,the expression of UCP2 protein and m RNA were increased,SV,EF and FS were significantly decreased,the concentrations of c Tn I and MDA were significantly increased,SOD activity was decreased,myocardial infarction area was increased,myocardial structure was severely damaged,mitochondria were significantly swollen,ROS production was increased,the ΔΨm,F1F0-ATP synthase activity and ATP synthesis were significantly decreased in I/R group(P<0.05).Compared with I/R group,the expression of UCP2 protein and m RNA were significantly decreased,myocardial infarction area was increased,the structure and function damage of myocardial tissue and mitochondrial were more serious,oxidative stress and ROS production was increased,theΔΨm,F1F0-ATP synthase activities and ATP synthesis were further decreased in I/R+Genipin group(P<0.05).Compared with I/R group,the expression of UCP2 protein and m RNA were significantly increased,myocardial infarct size was significantly reduced,the structure and function of myocardial tissue and mitochondrial were significantly improved,oxidative stress was decreased,ROS production was inhibited,ΔΨm was stable,F1F0-ATP synthase activity and ATP synthesis were significantly improved in I/R+AAV9-UCP2 group(P<0.05).Part II: Compared with Sham group,SV,EF and FS were significantly decreased,the concentrations of c Tn I,TNF-α,IL-6,IL-10 and MDA were significantly increased,SOD activity was decreased,myocardial infarction area was increased,myocardial structure was severely damaged,mitochondria were significantly swollen,ROS production was increased,ΔΨm was decreased,the opening of m PTP was increased,the apoptosis rate of cardiomyocytes was increased,the expressions of p-IκBα,p-NF-κB p65,BNIP3,Cyt-C and Cleaved caspase-3 were increased,and the expression of UCP2 protein and m RNA were increased in I/R group(P<0.05).Compared with the I/R group,myocardial infarct size was significantly reduced,the structure and function of myocardial tissue and mitochondrial were significantly improved,ROS production was inhibited,oxidative stress,inflammatory response and apoptosis were significantly reduced,the synthesis of ATP was significantly increased and the expression of UCP2 protein and m RNA were significantly increased in I/R+Dex group(P < 0.05).The up-regulation of UCP2 by Dex was inhibited and the myocardial protective effect of Dex was weakened after the addition of Genipin(P < 0.05).Part III: Compared with the Control group,Dex could stabilize the perioperative hemodynamics,significantly reduce the concentrations of postoperative c Tn I,MDA,TNF-α and IL-6,significantly reduce the incidence of postoperative arrhythmia,significantly shorten the postoperative mechanical ventilation time and ICU length of stay(P<0.05).The Dex group had a better effect than the Dex-Post group(P < 0.05).Conclusions:(1)Overexpression of UCP2 can protect diabetic myocardium against I/R injury by inhibiting ROS production and mitochondrial oxidative stress and improving ATP synthesis;(2)Dex can attenuate myocardial I/R injury in diabetics by inhibiting the production of ROS,lowering myocardial oxidative stress,inflammatory response mediated by ROS-dependent NF-κB activation and apoptosis mediated by m PTP sustained opening,effectively improving mitochondrial function and energy metabolism,which is at least partially dependent on up-regulating UCP2;(3)Dex can improve oxidative stress and inflammatory reaction in patients with diabetes after cardiac valve replacement under CPB,reduce myocardial I/R injury,shorten postoperative mechanical ventilation time and ICU hospitalization time,and effectively improve the clinical prognosis of diabetic patients undergoing cardiac surgery. |
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