| ObjectiveTo explore the targets and underlying mechanism of resveratrol(RSV)in regulating glycolipid metabolism in diabetic nephropathy(DN).Methods1.PPI networks of common targets of RSV and DN was conducted by STRING 11.0.Cytoscape 3.7.0 and VarElect database were applied to screen out the core targets of RSV in the treatment of DN.And GO and KEGG enrichment analysis of core targets were performed by DAVID database.At last,the binding models of RSV to the core targets were simulated by molecular docking using AutoDock Vina and the results were visualized.2.Based on the prediction results of the "drug-target-disease" network constructed by network pharmacology,male Wistar rats were selected to construct a DN model with streptozotocin(STZ)intraperitoneal injection combined with a high-fat diet for in vivo experimental validation.And they were divided into three groups:normal control(NC)group,model(DM)group and DM+RSV(DR)group(30 mg/kg/d by gavage).3.After 12 weeks,blood samples of rats were collected and livers and kidneys of rats were sacrificed.Fasting blood glucose(FBG),TC,TG,LDL-C,HDL-C,blood urea nitrogen(BUN)and serum creatinine(Scr)were assessed by an automatic biochemical analyzer.Liver weight,kidney weight and body weight were measured to calculate liver index and kidney index.Histopathological changes in livers and kidneys were evaluated by HE staining and PAS staining.The proteins expression of p-PDK1 and p-AKT in livers were assessed by Western blot.Additionally,expression of HIF-1α in livers and kidneys was detected by immunofluorescence and Western blot.The mRNA levels of Glut1 and Ldlr in the livers and kidneys were determined by RT-qPCR.Results1.There were 128 targets of RSV against DN,of which the core targets were AKT1,IL1B,INS,JUN and STAT3.Furthermore,the mechanism of resveratrol for regulating glucose and lipid metabolism in DN was associated with biological processes such as positive regulation of smooth muscle cell proliferation,response to drug and inflammatory response as well as seven signal pathways such as HIF1 pathway,FoxO pathway and TNF pathway.And molecular docking results showed that the affinities of the five core targets were all lower than-5.0 kcal/mol with multiple hydrogen bonds.Among all the core targets,AKT1 had the best binding activity to RSV.2.Compared to NC group,FBG,TC,TG,LDL-C,HDL-C,BUN,Scr,liver index and kidney index were increased significantly in rats in DM group,while body weight was decreased in DM group.After 12 weeks of RSV administration,all above indexes except HDL-C,kidney index and body weight showed significant improvement(P<0.05 or P<0.01).3.The livers of DN rats showed obvious pathological changes of steatosis,inflammatory infiltration and reduction of hepatic glycogen.And their kidneys showed glycogen accumulation,glomerular enlargement,tubular basement membrane thickening and renal interstitial focal inflammatory cell infiltration.But these pathological changes were effectively improved in the liver and kidney tissues of rats in the DR group.4.Compared to NC group,the proteins expression of p-PDK1 and p-AKT in the livers as well as HIF-1α in the livers and kidneys were significantly enhanced.And the mRNA level of Glut1 in the livers of DM group was significantly increased,while the mRNA level of Ldlr was significantly decreased.The levels of the above protein and mRNA were significantly improved in the DR group(P<0.05 or P<0.01).Conclusions1.Network pharmacology revealed that RSV may act on core targets such as AKT1,IL1B,INS,JUN and STAT3 and involve in regulation of HIF-1 signal pathway and FoxO signal pathway,thereby regulating glycolipid metabolism in DN.2.The prediction of network pharmacology was preliminary verified by animal experiments.And the results suggested that RSV could ameliorate glycolipid metabolism disorders in DN by inhibiting PDK1/AKT phosphorylation and HIF1α expression with down-regulation of Glut1 and up-regulation of Ldlr,respectively,thereby improved kidney injury to slow down the progression of DN. |
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