Effects Of Puerarin Combined With Alogliptin On Insulin Resistance And Myocaidial Fibrosis In Diabetic Rats

Objective: To investigate the effects of kudzu root combination with alogliptin on blood glucose,blood lipid,insulin resistance and to investigate the effect on myocardial protection and it's possible mechanism of diabetes rats which induced by high sugar and high fat diet with intraperitoneal injection of STZ.Methods: SPF clean grade SD male rats,of which 8 rats were randomly chosed as normal group(NC),while the remaining rats were made for type 2 diabetic model.The 42 rats which had been made successfully as type 2 diabetic models were randomly divided into several groups: Diabetes model group(distilled water,n=8),alogliptin group(5mg·kg-1·d-1,n=9),pue group(80mg·kg-1·d-1,n=8),pue with alogliptin group(40mg·kg-1·d-1+2.5mg·kg-1·d-1,n=9),alogliptin with metformin group(2.5mg·kg-1·d-1+50mg·kg-1·d-1,n=8),The rats were treated respectively,the normal group and diabetes model group were injected with normal saline.Body weight and fasting blood glucose were measured every two weeks.After six weeks of continuous administration,take abdominal aortic blood,isolate the hearts and measure the left ventricular mass(LVWW)and the heart wet weight(HWW).The left ventricular mass index(LVWI)and the heart weight index(HWI)were calculated.ELISA method was used to determine of total cholesterol(TC),triglyceride(TG),high density lipoprotein cholesterol(HDL-C)and low density lipoprotein cholesterol(LDL-C)levels in serum.Colorimetric assay was used for glycosylated hemoglobin(Hb A1c).ELISA method was used to measure fasting insulin(FINS)in serum,and insulin resistance index(HOMA-IR)and insulin sensitivity index(HOMA-ISI)were calculated(HOMA-ISI=ln1/(FBG×FINS),HOMA-IR= FBG×FINS÷22.5).ELISA method was used to measure the content of Ang?and TNF-?in serum.HE staining was used to observe the pathological changes of myocardium.MASSON staining was used to observe the changes of myocardial collagen fibers and calculate the CVF.Western Blot was used to detect the expression of TGF-?1,Smad3,Smad7 and CTGF protein in myocardium of rats in each group.The average optical density(IOD/Area)of protein TGF-?1 was measured by immunohistochemical method.Results:(1)Compared with the normal control group,body weight decreased significantly(P<0.01)in DM model group and the treatment groups,while FBG,Hb Alc,FINS,TC,TG,LDLC increased significantly(P<0.01 or P<0.05),showed obvious insulin resistance,and HDLC decreased significantly(P<0.01).Ang II in serum the content of TNF-increased(P<0.01 or P<0.05);HE staining showed myocardial hyperchromatic nuclei,and the gap was increased,myocardial hypertrophy,disorganized,produced obvious pathological injury.Masson staining showed large number of collagen deposition,and CVF increased significantly(P<0.01).Viscera index HWI and LVWI increased significantly(P<0.01)showed myocardial hypertrophy.The expression of protein TGF-?1?Smad3?CTGF increase in myocardial tissue(P<0.05 or P<0.01),while the expression of protein Smad7 decrease(P<0.05 or P<0.01).Immunohistochemistry results showed that the expression of protein TGF-?1 increased in diabetic rats(p<0.01).(2)Compared with DM model group,the weight of treatment groups increased(P<0.01 or 0.05),while FBG,Hb Alc,FINS,TC,TG,LDLC decreased(P<0.01 or P<0.05)and showed insulin sensitivity index improved(P<0.01 or 0.05).HDLC of the treatment groups increased(P<0.01)in addition to the alogliptin group.The combination group is better than the two single drug treatment group(P<0.01 or 0.05).The content of Ang II and TNF-? in serum decreased(P<0.01 or P<0.05),and the combination group decreased more significantly(P<0.01 or P<0.05).HE staining showed that the disorder level of myocardial cells of treatment groups has improved in different degrees,fibers were arranged orderly compared with model group.The alogliptin combined with puerarin group improved better than the single drug groups.Masson staining showed reduced blue collagen deposition,the disordered arrangement of myocardial cells were improved,and CVF increased significantly(P<0.01).The alogliptin combined with puerarin group improved more obvious.The organ index HWI and LVWI decreased(P<0.01)and the degree of myocardial hypertrophy have different degrees of improvement.The alogliptin combined with puerarin group is better than single drug therapy groups(P < 0.05),and has a better effect than the alogliptin combined with metformin group in the improvement of LVWI index(P<0.05).The expression of protein TGF-?1?Smad3 decrease in myocardial tissue(P<0.05 or P<0.01).The expression of protein CTGF decrease in myocardial tissue in treatment groups beside puerarin group(P<0.01).The expression of protein Smad7 increase in myocardial tissue(P<0.05 or P<0.01).Immunohistochemistry results showed that the expression of protein TGF-?1 decreased in treatment groups(p<0.01).(3)Compared with single drug treatment,puerarin combined with alogliptin significantly decrease the expression of protein TGF-?1,Smad3 and increase the expression of Smad7 protein(P<0.05 ? P<0.01),which significantly improve the myocardial fibrosis.And it's better than the group of alogliptin combined with metformin(P<0.05).Conclusion:(1)Puerarin combined with alogliptin decreases fasting blood glucose,glycated hemoglobin levels,reduce insulin levels in serum to improve insulin resistance,improve the abnormal metabolism of serum lipids on diabetic rats which induced by high sugar and high fat diet with intraperitoneal injection of STZ.The effect of combination therapy is superior to single drug treatment group.(2)Puerarin combined with alogliptin lowers the content of Ang?,TNF-? on diabetic rats which induced by high sugar and high fat diet with intraperitoneal injection of STZ.The effect of combination therapy is superior to single drug treatment group.(3)Puerarin combined with alogliptin improves CVF,left ventricular hypertrophy and myocardial fibrosis on diabetic rats which induced by high sugar and high fat diet with intraperitoneal injection of STZ.The effect of combination therapy is superior to single drug treatment group and the alogliptin combined with metformin group.The mechanism may be related to down regulation the expression of TGF-beta 1,Smad3,CTGF protein and up regulation the expression of Smad7.