Study On The Effect Of DPP-4 Enzyme Activity And Insulin Resistance In HepG2 Cells By Compounds Of Potentilla Bifurca

Potentilla bifurca var.humilior R.is a variant of Potentilla bifurca var.humilior,which mainly contains flavonoids,polysaccharides,tannins,and other active ingredients.It can be used for anti-tumor,anti-viral,anti-hyperglycemic embolism.Based on the 28 compounds isolated by the research group,7 active compounds were screened out in the early stage of the experiment.Through network pharmacology prediction,DPP-4 enzymatic reaction,glycolysis pathway,and PI3K-AKT signaling pathway to explore the improving effect of 7 compounds of Potentilla bifurca on insulin resistance.1.The 7 compounds with Potentilla bifurca is QC3G(Quercetin-3-O-?-D-glucopyranoside),QC3X(Quercetin-3-O-?-D-xylose),QC3GP(Quercetin-3-O-(6''-O-trans-p-coumaroyl)-?-D-glucoside),MC(Myricetin),QCQ(quercetin),EC((-)-epicatechin),CG(1-O-coumaroyl-?-D-glucose).The prediction of disease targets was carried out,and 32 potential targets of 7 compounds of Potentilla bifurca were screened.Through KEGG pathway analysis,19 possible pathways for the improvement of diabetes were obtained.Using the compound-target-pathway network diagram analysis,the PI3K-AKT signaling pathway has the largest node among the 19pathways of the Potentilla bifurca and has a strong correlation with diabetes disease.2.By establishing the DPP-4 enzyme reaction system to determine the inhibitory activities of the DPP-4 enzyme.Among them,compounds MC,QC,QC3X,QC3G,and QC3GP gradually increased their inhibitory activity to DPP-4 enzyme with the increase of concentration.The results of enzyme inhibitor characteristics showed that the Potentilla bifurca compounds were competitive inhibitors.3.The insulin concentration,action time,and stability during the establishment of the insulin resistance model were investigated.The results show that 10-8?mol·L-1 insulin can reach the IR state in Hep G2 cells with 48h,and this IR state can remain stable within 48h after modeling.The toxicity test results showed that when the compound concentration was 100?mol·L^-1,the cell viability of the compound MC,EC,QC3G,QC3GP,and CG groups was lower than 90%;when the compound concentration was 1?mol·L^-1and 10?mol·L^-1,the cell viability in each group was greater than 100%.4.Based on the insulin resistance model,the sugar consumption,pyruvate kinase activity,and hexokinase activity in the glycolytic pathway in IR-Hep G2 cells were measured.Compared with the model group,the results showed that the compounds EC,QC3X,and QC3G significantly improved the sugar consumption(P<0.01,P<0.05),and the compounds MC,QC,QC3GP,and CG improved the IR-Hep G2 cells.The activity of pyruvate kinase was significant(P<0.01,P<0.05),and the compounds MC,CG,and QC3G improved the activity of hexokinase in IR-Hep G2 cells significantly(P<0.01,P<0.05).5.Based on the prediction of the PI3K-AKT signaling pathway by network pharmacology,the contents of PI3K,AKT,and GLUT2 proteins in the PI3K-AKT signaling pathway were determined.Compared with the model group,the results showed that the compounds MC,EC,QC,QC3X,and QC3G significantly improved the content of PI3K in IR-Hep G2 cells(P<0.01),and compounds MC?EC?QC?QC3X?QC3G?QC3GP?CG could significantly improve the content of AKT in IR-Hep G2 cells(P<0.01).P<0.01),compounds MC,QC3X,QC3GP and QC improved the GLUT2 content in IR-Hep G2 cells with significant difference(P<0.01,P<0.05).In summary,The compounds could inhibit the activities of DPP-4,improve the activities of pyruvate kinase and hexokinase in the glycolytic pathway,and regulate the PI3K-AKT signaling pathway to treat insulin resistance,to achieve the effect of improving type 2 diabetes.