The Study Of Protective Effect Of Edaravone On Myocardial Injury In Type 1 Diabetes

In recent years, the high speed of development of the society and economic in china made great changes in Chinese people’s dietary structure. Besides, the increasingly rapid pace of life, erratic hours and lack of exercise today have made the populations aging and the burden of social security more and more serious, followed by the increasing morbidity in diabetes mellitus year by year. The treatment of Diabetes mellitus and its complication is difficult and bring heavy economic burden to society. Hence it is urged to study the pathogenesis of diabetes mellitus and its complication. Diabetic cardiomyopathy(DCM) is one of the main complications in patients with diabetes mellitus. It is mainly manifested by the hypertrophy of myocardiium and cardic disfunction, all of which leads to the end stage of heart failure. Up to now, we still lack the effective therapy about DCM. Therefore, it is urged to clarify the pathogenesis of DCM and prob a more effective intervention target.Silent mating type information regulation 2 homolog 1, SIRT-1, an NAD+ dependent histone deacetylase, plays an important role in regulating generation of ROS in the heart. SIRT-1 can improve glucose metabolism, promote the decomposition of fat, improve insulin resistance, improve cardiac hypertrophy and cardiac function.SIRT-1 directly deacetylated PGC-1α and activated PGC-1α, overexpression of PGC-1α in vascular endothelial cells increased mitochondrial antioxidant enzyme expression, and decreased oxidative stress and cell death. SIRT-1 can regulate the expression of many antioxidant proteins, and correct the imbalance between oxidation and antioxidation in pathological state. In addition, SIRT-1 can regulate many transcription factors to inhibit cell apoptosis. Oxidative stress and apoptosis is an important pathological mechanism in the development of DCM. Thus, SIRT-1 may be one of the important targets for the prevention and treatment of DCM.Edaravone(EDA), a potent free radical scavenger,was approved in 2001 in Japan for the treatment of ischemia stroke. It has since been shown that edaravone can diffuse into many organs and, in addition to its neurovascular protective effects, edaravone exerts protective effects in a number of animal models of disease and tissue damage, including models of myocardial, muscle, lung, liver, pancreatic and renal injury. The preventive effects of edaravone on cardiomyocytes apoptosis, necrosis, as well as myocardial injury following ischemia reperfusion are well documented. As an anti-oxidant regent, it might provide a new method for the treatment of cardiovascular diseases. It is unknown that whether edaravone protects cardiac function loss during the development of diabetes. A variety of effects of edaravone is at least partially attributed to the elimination of ROS. Owing to activating SIRT-1, We speculate that edaravone has protective effect on diabetic cardiomyopathy.This experiment from the two aspects of whole body and cell levels discussed the protective effect of edaravone on diabetic cardiomyopathy in rats, so as to provide a theoretical basis for the prevention and treatment of diabetic cardiomyopathy.In this study, we examined the protective effect of edaravone on diabetic myocardial injury in type 1 diabetes rats. It provides theoretical basis for the prevention and treatment of diabetic cardiomyopathy.First, we investigate the protective effect of edaravone on myocardial injury of diabetes in vitro. Six-week-old male Wistar rats were housed were housed individually in a room(constant temperature, 12/12 h light/dark cycles) and received standard laboratory rat diet and water ad libitum.Type 1 diabetic was induced with a single intraperitoneal injection of streptozocin(STZ)(60 mg/kg) in our rats. 7 days after the injection a drop of blood was collected from the tail vein and blood glucose concentration was determined by using a digital blood glucose meter. Animals with a random blood glucose level >16.7 mmol/L were considered to be diabetic and were included into the study. Rats were divided randomly into three groups: vehicle-treated control(CON; n = 12), vehicle treated diabetic(Model; n = 12) and edaravone-treated diabetic(EDA; n = 10) groups. Animals were treated with saline or with edaravone(6 mg/kg/day) by intraperitoneal injection for 4 weeks. Body weight, Blood glucose(BG), Insulin(INS), malondialdehyde(MDA) and superoxide dismutase(SOD) in rat serum were determined, We measure cardiac function in rats with echocardiography. HE staining was used to observe the morphology of myocardium. TUNEL assay was performed to detect DNA strand breaks. For western blot detection of SIRT-1, PGC-1α, NRF-2 and Bcl-2, Bax, Caspase-3 protein expressions.Experimental results show that Edaravone treatment increases insulin and superoxide dismutase levels, E/A and EF% were significantly decreased in Model animals compared to CON,Treatment of Model animals with edaravone restored E/A and EF% to that of CON animals. In addition, IVS was significantly increased in Model animals compared to CON, treatment of Model animals with EDA restored IVS to that of control. Thus, EDA treatment improves cardiac function in type 1 diabetic rats. To evaluate cardiomyocytes hypertrophy, cardiomyocytes diameter were measured in H&E staining, when compared with controls, the cardiomyocytes diameters in Model animals were significantly increased, which indicative of cardiomyocytes hypertrophy, this increase was prevented by EDA treatment. Treatment with edaravone significantly decreased the number of TUNEL-positive nuclei in the myocardium. SIRT-1, PGC-1α and NRF-2 protein expressions were significantly decreased in diabetic animals compared to CON, treatment of diabetic animal with EDA increased SIRT-1, PGC-1α and NRF-2 protein expressions compared to diabetic animals. EDA treatment increases SIRT-1, PGC-1α, NRF-2 and Bcl-2 and reduces Bax and Caspase-3. The results showed that edaravone can significantly improve cardiac function in type 1 diabetic rats, decrease the apoptosis of myocardial cells, and its protective effect may be related to activation of SIRT-1.Next,we observe the effect of edaravone on the toxicity on high glucose treated H9c2 cells, reactive oxygen species, SIRT-1 protein expression in cell level, interpretation protective effect of edaravone on high glucose treat H9c2 cells and its mechanism. H9c2 cells were incubated for 24 h with various concentrations of EDA or high glucose with EDA, and cytotoxicity,ROS level, expression of SIRT-1,PGC-1α,NRF-2,Bcl-2,Bax and Caspase-3 examined. Our experimental results show that treatment of H9c2 cells with EDA significantly reduced ROS generation induced by high glucose. Edaravone treatment increases SIRT-1, PGC-1α, NRF-2 and Bcl-2 and reduces Bax and Caspase-3.Finally, we discuss the molecular mechanism of H9c2 myocardial cell damage protection of edaravone treated high glucose. As SIRT-1 regulates cell growth and survival in response to oxidative stresses, we next examined if knockdown SIRT-1 m RNA could abolish the protective effects of EDA. The results showed that knockdown SIRT-1 m RNA by small interfere RNA caused a reduction in SIRT-1 protein expression, accompanied by reduced levels of PGC-1α and NRF-2 expression in high glucose treated H9c2 cells. In addition, after knockdown SIRT-1, Bax and Caspase-3 was markedly increased and Bcl-2 was reduced in high glucose and EDA treated cells.In conclusion, edaravone has a protective effect on inhibiting myocardial injury in type 1 diabetes, SIRT-1 is one of the key targets of its protective role, edaravone could inhibit cardiac oxidative stress and apoptosis through the activation of SIRT-1, and the underlying mechanism may be related to regulate expression of PGC-1, NRF-2 and apoptosis related proteins through SIRT-1.