Mechanisms Of Specnuezhenide On Improving Insulin Resistance In Type 2 Diabetic Mice

Background:The incidence of type 2 diabetes(Diabetes mellitus type 2,T2DM)has risen dramatically in recent decades.The main feature of T2 DM is insulin resistance(Insulin resistance,IR).During IR,glucose output from the liver increases and the ability of adipocytes to take up glucose decreases.Although various effective hypoglycemic drugs have been used clinically,these drugs have certain limitations.Traditional Chinese medicine is widely used in hypoglycemic therapy because of its good curative effect and less side effects.Therefore,the use of traditional Chinese medicine or its monomers to improve insulin resistance has become one of the effective strategies for the treatment of T2 DM.Compound Zhenshu Tiaozhi Fang is a representative prescription for the prevention and treatment of diabetes and lipid metabolism,pioneered by the discipline leader of this platform.It is mainly composed of Citri Sarcodactylis Fructus,Ligustri Lucidi Fructus,Largehead Atractylodes Rhizome and other traditional Chinese medicines,of which specnuezhenide is the characteristic active ingredient of Ligustri Lucidi Fructus.According to the previous research of our research group,it was found that the insulin resistance can be effectively alleviated after administration of specnuezhenide through the insulin resistance Hep G2 cell model,and further in vivo verification in animal models is still needed.Therefore,in this study,by constructing a T2 DM mouse model,we deeply explored the mechanism of specnuezhenide in improving insulin resistance in T2 DM mice.Objective:1.To investigate the mechanism of specnuezhenide to improve insulin resistance in liver and adipose tissue to prevent and treat T2 DM.2.To explore the mechanism of specnuezhenide to improve insulin resistance by promoting fat browning.3.Investigate the mechanism of specnuezhenide improving insulin resistance through changes in the structure of intestinal flora.Methods:1.C57BL/6J mice were fed with high-fat diet for 4 weeks + low-dose STZ to establish a type 2 diabetes model.According to the blood glucose level of blood collected from the tail tip of the mice,the mice with successful modeling were randomly divided into the model group,the low-dose specnuezhenide group,the high-dose specnuezhenide group,and the metformin group.2.After the drug intervention,the serum of the mice was taken to measure the four items of blood lipids and insulin;The fatty degeneration of liver tissue was observed by H&E and Oil Red O staining experiments;Transcriptome analysis was used to explore the mechanism of improving insulin resistance;Quantitative Real-time PCR(q PCR)technology was used to detect the transcription levels of genes related to inflammatory factors in liver;The transcription level and protein expression of genes related to insulin signaling pathway and lipid accumulation in liver were detected by q PCR and Western blot(WB).3.The protein expression level of p Ser473 AKT in brown fat and visceral fat was detected by WB technology;The transcription level of browning-related genes in adipose tissue was detected by q PCR technology.4.Fecal samples were collected and analyzed by 16 S DNA sequencing to explore the changes and dominant flora of intestinal flora after drug intervention.Results:1.After the mice were modeled,the mice in the model group had a fasting blood glucose ≥11.1 mmol/L,and the T2 DM mice model was successfully established.2.Specnuezhenide decreased the hepatic pro-inflammatory factor(TNF-α,IL1β,IL-6)and increased the transcription level of anti-inflammatory factor(IL-10).3.Specnuezhenide increased the expression of hepatic IRS,AKT and GSK3β phosphorylated protein and decreased the expression of GS phosphorylated protein.4.The expression levels of ACACA,FASN,SCD1 and CD36 in the liver were decreased by specnuezhenide.5.Specnuezhenide decreased the expression of AKT phosphorylated protein in brown fat and visceral fat and increased the transcription level of "browning" genes in brown fat and subcutaneous fat.6.Specnuezhenide increased the abundance of Akkermansia and decreased fatty acid biosynthesis.Conclusion:1.Specnuezhenide effectively reduces the genes related to fatty acid synthesis to improve liver lipid accumulation,and effectively regulates the activities of key factors in the liver insulin signaling pathway such as IRS,AKT,GSK3β and GS.2.Specnuezhenide improves the insulin sensitivity of fat by improving the activity of brown fat by activating the AMPK-SIRT3 pathway,and promoting the browning of subcutaneous fat.3.T2 DM can effectively regulate the abundance and function of intestinal flora in T2 DM mice.