| Objective: To observe whether gliclazide can improve myocardial injury in diabetic rats and to explore whether this protective effect is regulated by Rho proteins as small molecule guanosine-binding proteins(RhoA),downstream effector proteins Rho kinases(ROCK1)and endothelial nitric oxide synthase(eNOS).Methods: 1.Animal experiment: 60 healthy SD(Sprague Dawley)rats with a body mass(200±20)g,free drinking water and diet.The rats were randomly divided into two groups: normal group(NC,n=10)and model making group(n=50).Diabetic rats were established by high sugar and high fat diet combined with intraperitoneal injection of streptozotocin.38 diabetic rats were randomly divided into model group(MC),gliclazide group(Glic),glibenclamide group(Glib)and fasudil group(Fas).The rats in gliclazide group(80mg/kg)and glibenclamide group(2.5mg/kg)were administrated by gavage;the rats in normal control group and model group were administrated by distilled water of corresponding dose daily;and the rats in fasudil group(10mg/kg)were administrated by intraperitoneal injection.The body mass and fasting blood glucose of each group were recorded every week for 8 weeks.At the end of the experiment,the heart weight was measured,and the heart weight index(HWI)was calculated;the contents of glycosylated hemoglobin(HbA1c),total cholesterol(TC),triglyceride(TG),high density lipoprotein(HDL-C),low density lipoprotein(LDL-C),the level of serum malondialdehyde(MDA)and the activity of superoxide dismutase(SOD)were measured;the shape of myocardial tissue was observed by HE and Masson staining.The expression of RhoA,ROCK1,eNOS,Bcl-2 and Bax protein was detected by Western blot.2.H9c2 cell experiment: H9c2 cells were inoculated into culture medium and randomly divided into normal group(NC),model group(MC),osmotic control group(OS),gliclazide low dose group(Glic+L),gliclazide high dose group(Glic+H),glibenclamide group(Glib)and fasudil group(Fas).NC and OS groups were treated with 5.5 mmol/L of glucose and the other groups were cultured in medium with 35mmol/L of glucose concentration for 72 h.The activity of H9c2 cells in each group was detected by MTT method,the activity of SOD,the level of MDA and ROS in each group were measured,and the level of apoptosis in each group was detected by flow cytometry.Results: 1.Animal experiment: Compared with the NC group,the body mass,FBG,HWI,HbA1 c,LDL-C,TG,TC,MDA and CVF of the rats in the MC group were significantly increased(P < 0.01),SOD and HDL-C were significantly reduced(P < 0.01),The level of myocardial fibrosis and apoptosis was significantly increased(P < 0.01).RhoA,ROCK1 and Bax protein expressions were significantly increased(P < 0.01).The eNOS and Bcl-2 protein were significantly decreased(P < 0.01).Drug treatments ameliorated myocardial fibrosis and inhibited myocardial apoptosis.Glic treatment decreased FBG,HWI,HbA1 c,LDL-C,TG,TC and MDA,increased SOD and HDL-C(P < 0.01 or P < 0.05);decreased collagen deposition,inhibited cardiomyocyte apoptosis(P < 0.01);decreased RhoA,ROCK1 and Bax protein expression;increased eNOS,Bcl-2 protein level(P < 0.01 or P <0.05).2.H9c2 cell experiment:(1)compared with the normal group,the proliferation ability of cells in the model group decreased significantly(P < 0.01),the content of ROS and MDA increased,the activity of SOD decreased,and the apoptosis rate increased significantly(P < 0.01).(2)Gliclazide and fasudil could improve the activity and apoptosis of H9c2 cells(P < 0.01 or P < 0.05).Gliclazide could increase the activity of SOD,reduce the level of ROS and MDA,and down regulation the oxidative stress of H9c2 cells.Conclusion: 1.Gliclazide significantly alleviates myocardial injury and reduces myocardial apoptosis in diabetic rats,and itsmechanism may be related to lowering blood glucose,down-regulation oxidative stress and regulating RhoA / ROCK1 / eNOS signaling pathway.2.High glucose can significantly induced increased apoptosis in H9c2 cells,gliclazide and fasudil could down regulation oxidative stress and alleviate apoptosis. |
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