Active Ingredients And Mechanisms Of Mulberry Prevention And Treatment Of Diabetes Based On Network Pharmacology

OBJECTIVE: To use the network pharmacology technology,medium-pressure purification system technology,and HepG2 insulin resistance cell model to study the active ingredients and active substances of mulberry to prevent and treat DM on the regulation of key protein expression in PI3K-Akt-mTor,an important signaling pathway for glucose and lipid metabolism.Methods: 1.Search the database to obtain mulberry active ingredients and related target genes,conduct disease mapping,and use Cystoscope3.2.1 software to construct a "compound-target-disease" network diagram.Carry out Metascape channel enrichment analysis.2.Use the medium-pressure preparation and supporting purification system to separate and purify the chemical components of the mulberry alcohol extract,and combine HPLC and LC-MS to analyze the composition and identify the structure of the separated product.3.Establish HepG2 insulin resistance cell model,comprehensively evaluate the effect of each administration group on insulin resistance HepG2 cell status by measuring glucose consumption,HK,PK,TC,TG of each group after administration,using Western blotting(Western blotting,WB)method was used to detect the protein expression of Akt,P-Akt,PI3 K,P-PI3 K,mTor and P-mTor.Results: 1.The network modeling of mulberry screened a total of 22 active ingredients,16 high-frequency targets and 40 signal pathways related to diabetes.Enrichment analysis results show that the prevention and treatment of diabetes by mulberry may be caused by the synergy of its active ingredients,and the target may be closely related to the PI3K-Akt signaling pathway,nitrogen metabolism,and galactose metabolism.2.Established a rapid separation and purification method using medium-pressure purification system technology.A total of three compounds in the mulberry extract were obtained in this experiment.After HPLC standard control and molecular weight verification of mass spectrometry,they were proved to be neochlorogenic acid,rutin,and isoquercitrin.3.250 μmol / L palmitic acid-induced HepG2 cells for 24 h is the best plan for establishing insulin resistance model.The ethyl acetate layer,n-butanol layer,neochlorogenic acid,isoquercitrin,mulberry pigment,and myricetinhave good activity(P <0.05),indicating that they are the main active compounds of mulberry to improve insulin resistance.The mulberry extract can increase the HK and PK activities of insulin-resistant cells(P <0.05);significantly reduce the accumulation of TG in cells(P <0.05),and can clear TC(P <0.05),the difference is statistically significant.Western-blot method detected related proteins in cells.The results showed that after drug intervention,compared with the model group,the middle-dose ethyl acetate group and the n-butanol medium-dose group Akt,P-Akt,PI3 K,P-PI3 K,m-TOR,P-The expression level of mTor protein was significantly increased,and the difference was statistically significant(P <0.05).Conclusions: 1.Predicting and analyzing active components and diabetes targets through network pharmacological methods are helpful to analyze the multi-action targets of mulberry active ingredients in synergy with anti-diabetes.2.The different extraction sites of Mulberry and a variety of small molecular substances all have the effect of improving insulin resistance of HepG2 cells.This mechanism may be related to the enhancement of cellular lipid metabolism and the expression of related proteins(PI3K、Akt、mTor)in the insulin signaling pathway.Keywords: mulberry;diabetes;HepG2 cells;insulin resistance related to the enhancement of cell lipid metabolism and the expression of related proteins in the insulin signaling pathway.