| BackgroudPrevelance of diabetes is high and is increasing in the world. It has been an important disease that threats to human health. Prevelance of diabetes is also increasing in China which has been one of the most patients. And China has to be confronted with the health burden. Diabetes mellitus which is characterized by glucose metabolism disorder can cause a variety of complications, including cardiovascular disease. Ischemia heart disease is the principal cause of death in patients with type2diabetes mellitus. Several recent studies implicated high blood glucose have a damage of myocardial cells directly, resulting in more frequent arrhythmia and more lager infarction area after myocardial/ischemia, which indicates that diabetic heart increased sensitivity to ischemic injury. And the specific mechanism is not clear.Recent studies have shown, own to the high level of plasma glucose, protein will be modified by glucose or derivatives. Protein nonenzymatic glycation reaction begins with attachment of a reducing sugar to a biological amine of a protein, and then can undergo further rearrangement to form advanced glycation end products without enzyme. Proteins structure is closely related to its biological functions. Their biological functions have been changed following their structure changed. Protein nonenzymatic glycation is thought the cause of DM. Evidence has shown that ROS could be actived by advanced glycation end-products(AGEs), enhancing the myocardial apoptosis and increasing myocardial ischemic/infarction area. AGEs are a complex group of compounds formed and the the molecular pathway for increasing diabetic heart sensitivity to myocardial ischemia/reperfusion injury is still not clear.Thioredoxin (Trx) is a12Kd protein ubiquitously expressed in all living cells with redox function.Recently, more and more evidence has shown Trx plays an important role in reducing death. Their biological functions have been changed following their structure changed. As our previous study shown, Trx was incubated by methylglyoxal and there was AGEs detected by immunopercipitation. It indicated that Trx was modified by glucose.Based on these fact, we speculate that the nonenzymatic glycation of thioredoxin inhibits its activity resulting in decreasing anti-apoptosis function, leading to inccresing cardiomyocyte injury after ischemia/reperfusion. Therefore, this research will build type2diabetes mellitus rat treated with ALT-711to confirm that:(1) whether nonenzymatic glycation of Trx will happen and whether affect its activity;(2) whether nonenzymatic glycation of Trx will increase susceptibility to myocardial ischemia/reperfusion injury.Material and Methods1.Six-week old male Wistar rats were randomly divided into three groups:Control group, DM group, DM+ALT-711group. Control group were fed basic diet, DM group and DM+ALT-711group were fed high sucrose and high fat diet for four weeks and then injected with STZ (40mg/kg.i.p.) and Control group were injected with sodiumcitrate solution.48h after the injection, the level of fasting blood glucose was determined.The fasting blood glucose value above11.1mmol/L was considered to be DM. DM+ALT-711group were trated with ALT-711(40mg/kg) by gavage at12th week for8weeks. The levels of fasting blood glucose were monitored every week. The body weights of rats were also measured every week. Take the myocardial tissue of rat at the0,4th,6th,8th,12th,16th,20th week for deteminnation. Measure the Trx activity.Detect the expression of Trx by Western blot.Detect the expression of AGEs by immunohistochemistry.Detect the glycation of Trx by immunoprecipitation.2. Six-week old male Wistar rats were made to be DM and then were randomly divided into two groups:sham group, MI/Rgroup at the20th week. MI/R model was ischemia for3h and reperfusion for30min.The myocardial infarct size in diabetic MI/R group was measured by Evan’s Blue and TTC staining.The caspase-3activity in diabetic MI/R group and the apoptosis index were determined.Results1.The levels of body weight and fasting blood glucoseCompared with control group, the bodyweight of DM group and DM+ALT-711group significantly increased before the STZ-injected. DM group and DM+ALT-711group were significantly lower than control group at14th week. There is no obvious chang in their fasting blood glucose before the STZ injection. Compared with control group, the fasting blood tglucose of DM group and DM+ALT-711group significantly increased after injection and maintained high level.2.The activity of Trx in diabetic rats decreased but expression of Trx in diabetic rats has no obvious changeCompared with control group, the relative activity of Trx in DM group and DM+ALT-711group significantly decreased and their value was lowest at20th week.But the treatment of ALT-711could restore Trx relative activity.The expression of Trx dramatically increased in diabetic rats for4,6,8,12,20weeks. However, there is no obvious change of Trx expression in control group. 3.The expression of AGEs in diabetic rats increased and the glycation of Trx in diabetic groupCompared with control group, the expression of AGEs began to express at6th week. AGE expression significantly increased at12th week. The glycation of Trx could be detected at8th,12th,16th,20th week.4.Treatment withALT-711significantly decrease the expression of AGEs.It also could decrease the glycation of Trx.5.The myocardial infarct size in diabetic MI/R group increasedCompared with control MI/R group, the infarct size in diabetic MI/R group significantly increased. Treatment with ALT-711reduced infarct size. Treatment with ALT-711could restore the cardial function.The caspase-3activity in diabetic MI/R group increased. Treatment with ALT-711, the caspase-3activity reduced. The apoptosis index in diabetic MI/R group increased. Treatment with ALT-711, the c apoptosis index reduced.ConclusionThe nonenzymatic glycation of Trx will inhibit its activity especially anti-apoptosis function in diabetic heart which leading to increasing susceptibility to myocardial ischemia/reperfusion injury; ALT-711could restore Trx activity in DM myocardial and reduced diabetic heart sensitivity to myocardial ischemia/reperfusion injury. |
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