Effect Of Long-term Insulin Treatment On Plasma And Cardiac Adiponectin Levels And The Cardioprotection Against Myocardial Ischemia/Reperfusion In Streptozotocin-induced Diabetic Mice

【BackgrounBackground】Now, diabetes has become global epidemic. It is estimated that by 2025,the number of diabetic patients is expected to rise to 380 millionworld-wide.Type 1 diabetes mellitus is mainly caused by a selective destructionof pancreaticβcells；with the loss of endogenous insulin, diabetic complicationand metabolic disorders happen. Cardiovascular disease is one of the majorcomplications of diabetes, resulting in a high percentage of morbidity andmortality.Diabetes and its cardiovascular complications are both related tomultiple pathogenic factors, including hyperglycemia and inflammatoryresponses. Under this condition, exogenous insulin is required to compensate theresulting metabolic disorders.Several epidemiologic studies demonstrated that atight glycaemic control as well as an intensive insulin treatment reduces the risk of late diabetic complication, and improves the quality of life.Adiponectin is a protein secreted predominantly by differentiatedadipocytes, with levels ranging from 5 to 30μg/ml, and accounts for 0.01% oftotal plasma protein. Experimental studies show that adiponectin can modulatelipid and glucose metabolism via activating AMP-activated protein kinase(AMPK) activities, besides a protective role in the development of insulinresistance and inflammation.There is a local cardiac-specific adiponectin systemwhich play important roles in maintaining energy homeostasis incardiomyocytes.However, the relationship between adiponectin and type 1 diabetes is stillunclear.The influence of exogenous insulin on the local cardiac-specificadiponectin system has been unknown,and the effect of the above influenceagainst Myocardial Ischemia/reperfusion injury should be further studied.【Objectives】1. To investigate the time-course change of plasma,adipo-derived and cardiacadiponectin levels in STZ-induced diabetic mice;2. To determine the effect of insulin treatment on plasma, adipo-derived andcardiac adiponectin levels and to discuss the possible mechanism of cardiacadiponectin in the protection against myocardial ischemia/reperfusion injury.【MethodMethods】Part 1:1. 72 male mice were randomly assigned to control(D1,D7,D14),Diabetic(D1,D7,D14)and insulin treatment (D1,D7,D14)groups.The micein diabetic and insulin treatment groups were consecutively given STZperitoneal injection (50mg/kg·d) for 5 days,and then observed for another5 days.Diabetic model was established if random fast blood glucose >10 mmol/L.Then,the mice in insulin treatment group were given insulin(5IU/kg·d)through subcutaneous injection;2. Collect the adipose tissues,blood,and hearts from all groups at the time of1,7,14 days after the establishment of STZ-induced diabetic model;3. With Insulin Radioimmunoassay Kit,plasma insulin levels were detected;4. Plasma adiponectin levels were measured through ELISA;5. Adipo-derived adiponectin levels were confirmed by Western blot;6. The expression levels of adiponectin in cardiac tissues between groups werecompared by RT-PCR.Part 2:1. Same with step 1 of part 1;2. Extracorporeal perfusion and Myocardial Ischemia/reperfusion wereperformed with Langendorff;3. Hearts of all mice were randomly divided to 6 groups:A. Extracorporeal perfusion with non Ischemia/reperfusion, including controlgroup and diabetic group;B. Extracorporeal perfusion with Ischemia/reperfusion, including control group,diabetic group, insulin treatment group, and AMPK inhibitor group;C. The size of myocardial infarction was determined through TTC staining.D. The levels of cardiomyocytes apoptosis between groups were compared byTUNEL.【ResultResults】Part 1:1. 46 diabetic models were established and 2 died,the achievement ratio washigh;Compared with normal group, 2. Plasma insulin levels in type 1 diabetic mice decreased(P<0.05);3. The levels of adiponectin from plasma and adipose tissues in type 1 diabeticmice firstly increased(P<0.01),and then decreased; The expression levels ofadiponectin in cardiac tissues decreased(P<0.05).Compared with diabetic group,4. The insulin levels of mice in insulin treatment group were elevated(P<0.05)；5. The levels of adiponectin from plasma and adipose tissues in mice treatedwith insulin were elevated(P<0.01); in cardiac tissues, the adiponectinexpression levels were elevated(P<0.05).Part 2:1. 30 diabetic models were successfully established and 2 were lost；Under the condition of extracorporeal perfusion with nonIschemia/reperfusion,2. Considering the size of myocardial infarction and the levels ofcardiomyocytes apoptosis, there was no difference between control groupand diabetic group;Under the condition of extracorporeal perfusion with Ischemia/reperfusion,3. When compared with control group, the size of myocardial infarction andthe levels of cardiomyocytes apoptosis significantly increased(P<0.01);4. Compared with diabetic group, the size of myocardial infarction as well asthe levels of cardiomyocytes apoptosis decreased in insulin treatment group(P<0.01);5. Compared with insulin treatment group, AMPK inhabitor could partlyincrease the size of myocardial infarction and the levels of cardiomyocytesapoptosis(P<0.01). 【ConclusionConclusions】1. The levels of adiponectin from plasma and adipose tissues in type 1 diabeticmice show a bi-directional change along the course of the disease.2. In cardiac tissues, local adiponectin level gradually decreases.3. Long-term insulin treatment increases plasma,adipo-derived and local cardiacadiponectin levels .4. The rise of cardiac adiponectin induced by insulin treatment may increase thetolerance of cardiomyocytes against ischemia/reperfusion injury.