Effect Of Sitagliptin On Expression Of PEDF And The JAK/STAT Pathway In Renal Tissues Of Diabetic Rats

Objectives:To explore the effect of Sitagliptin on expressions of PEDF and JAK/STAT pathway in renal tissues of diabetic rats.Methods:After adaptivie feed for 1 week,60 male Sprague-dawlry?SD?rats were randomly divided into normal group?group A,n=10?and model group?n=50?.Normal control group was fed with the normal diet,drinking freely,model group was fed with high sucrose-fat diets?HSFDs?,also drinking freely.After HSFDs fed for 4 weeks,weight of rats were tested,blood samples of rats were collected and the fast blood glucose?FBG?,fasting insulin?FINS?level,total cholesterol?TC?,triglyceride?TG?,low density lipoprotein cholesterol?LDL-C?were assayed to determined whether the insulin resistance was induced.Two rats died of infection.Model group was fed with high sucrose-fat diets for4 weeks,the subjects in model group were injected intraperitoneally with streptozotocin?STZ,35mg/kg?after fasting for 12 hours.These diabetic rats were determined by randomly blood higher than 16.7mmol/L of three consecutive times after 72 hours of STZ injection?two rats was excluded because of failing to achieve the diagnosis standard of diabetes,two rats was excluded because of died of infection or DKD.?Rats in the normal control group were injected with citric acid buffer solution of the same volume.Rats of group A were given conventional diet;after successful modeling of diabetic SD rats,they were given high-fat diet and randomly divided into 5 groups:the non-intervention group?group B,n=10?,the group treated with sitagliptin?5mg/kg.d??group C,n=9?,the group treated with sitagliptin?10mg/kg.d??group D,n=9?,the group treated with sitagliptin?5mg/kg.d?+specific JAK2 ingibitor-AG490?0.1mg/kg.d??group E,n=8?and the group treated with sitagliptin?10 mg/kg.d?+specific JAK2 ingibitor-AG490?0.1mg/kg.d??group F,n=8?;rats of group A were given intragastric administration of normal saline of the same volume;rats in all the groups were intervened for 8 weeks.Rats of group E and group F were given intraperitoneal injection of specific JAK2ingibitor-AG490?0.1mg/kg.d?every day since the 10th week of the experiment;at the end of the 12th week,body weight and 24-hour urine microalbumin of rats of each group were measured;and after their blood was obtained to measure the levels of fasting blood glucose?FBG?,fasting insulin?FINS?,serum creatinine?SCr?,blood urea nitrogen?BUN?,TG,TC and LDL-C,rats were dissected and their right and left kidneys were taken out;The morphological changes of kidney were observed through light microscopy after hematoxylin-eosin?HE?staining.Immunohistochemistry staining was used to examine the expression of PEDF,P-STAT3,TGF?₁ and FN protein in the kidney.The real-time PCR method was used to measure the mRNA expression levels of PEDF in renal tissues of diabetic rat models.Results:?1?Body weights and biochemical indexes of rats after 4 weeks:Compared to normal group,BW,FBG,FINS,HOMA-IR,TG,TC and LDL-C of rats in the model group were increased,with significant statistical difference?P<0.05?;?2?After 12 weeks,compared with group A,BW of the rats in group B,C,D,E,F were significantly lower?P<0.05?.and FBG,TG,TC,LDL-C,SCR,CRP,BUN,24h urine protein of group B,C,D,E,F were significantly higher?P<0.05?.Compared with group B,FBG,TG,TC,LDL-C,SCR,CRP,BUN,24h urine protein of group C,D,E,F were significantly lower?P<0.05?;?3?The histopathological examination of rat renal by the naked eye and light microscopy:the normal control group A with glomerular morphology neat,glomerular cells arranged in rules,basement membrane was clear and tidy,no thickening,glomerular capillary lumens stenosis and renal tubular-interstitial inflammatory cell infiltration.Group B with glomerular volume increase,glomerular capillary loops collapse,occlusion of the lumen,thickening of basement membrane increased and a large number of inflammatory cell infiltration in renal interstitial.And the lesions in kidney of the rats of group C,D,E and group F with drug intervention was significantly lighter than that of group B;?4?Immunohistochemical results,compared with group A,the expressions of PEDF in group B,C,D,E rats were decreased?P<0.05?;and the expressions of p-STAT3,TGF-?₁and FN in group B,C,D,E rats were increased?P<0.05?;Compared with group B,all the indexes but the expressions of PEDF mentioned above of rats in group C,D,E,F were decreased,the expressions of PEDF were increased,with significant statistical difference?P<0.05?;group E compared with group C,and group F compared with group D,the expressions of PEDF was increased,and the expressions of p-STAT3,TGF-?₁and FN was decreased,with significant statistical difference?P<0.05?;?5?Rat kidney PEDF mRNA expression level:compared with group A,group B and C,D,E,F group in the rat kidney PEDF mRNA expression were significantly lower?P<0.05?;Compared with group B,PEDF mRNA expression of the rats of group C,D,E,F were significantly higher?P<0.05?.Compared with group C,Group D PEDF mRNA expression had increased?P<0.05?;While using the JAK2inhibitor AG490,Group E compared with group C,group F compared with group D,PEDF mRNA expression significantly increased?P<0.05?.?6?Pearson correlation analysis showed that expression of PEDF in renal tissues of diabetic rats was negatively correlated with p-STAT3,TGF-?1 and FN?r=-0.762,-0.831,-0.714,R²=0.581,0.690,0.510,P<0.05?;expression of p-STAT3 was positively correlated with that of TGF-?₁ and FN?r=0.877,0.851,R²=0.770,0.724 P<0.05?.Conclusions:Sitagliptin may up-regulate the expression of PEDF,inhibits the JAK/STAT signaling pathway in renal tissues of diabetic rats,and then inhibits the expression of the downstream factor TGF-?₁and FN to reduce the rinflammatory reaction of DN.