The Research Of Total Flavonoids Of Selaginella Tamariscina (Beauv.) Spring And Amentoflavone On Improving Insulin Resistance By The Method Of Proteomics

Objective: Using Type2diabetes model and proteomics technology to research TFST’seffect of hypoglycemic, and building insulin resistance model of HepG2cells to find themechanism of TFST and AME on improving insulin resistance.Methods:1, Wistar rats were fed with high-sugar and high-fat diet to induced insulinresistance in rats for8weeks, intraperitoneal injection STZ of35mg/kg to establish Type2diabetes model TFST (100,200and400mg/kg/day) were administered orally for8weeks.After the animals were killed by cervical, liver total protein were isolated by proteomicsmethod, and detect changes in each group with PDQuest software. Protein spots were cutwith3-fold difference to detect mass spectrometry. The protein’s information were searchedin the protein database and obtained the mechanisms and difference of TFST onhypoglycemic effect.We detected the levels of blood glucose, blood lipid, insulin, glucagonto prove the effect characteristics of TFST. Slice observed repair of pancreas, liver andkidney cells’ damage, and verify the results by detecting related indicators of liver andkidney function and inflammatory factors.2, Model of insulin resistance cells wereestablished, total protein was extracted to do the experiment of proteomics research, andobtain high resolution two-dimensional electrophoresis patterns. Using PDQuest software tocomparing changes in each group, and Protein spots were cut with3-fold difference to detectmass spectrometry. We could search protein information through protein database, and thenuse Western-blot and ELISA methiods to verify the results of proteomics. The mechanism ofTFST and AME on improve insulin resistance were probed through these methods.Results:1, Proteomics results showed that there are195protein spots isolated with the usingof two-dimensional electrophoresis,30protein spots were cut with3-fold difference by the software analyze to detect mass spectrometry. We analyzed the information and found themechanism of TFST’s hypoglycemic effect was related with PI3K-Akt signaling pathway,NF-κB pathway, P53pathway, and also can improve glucose metabolism in diabetic rats,regulating endoplasmic reticulum function. After the rats were sacrificed for the detection ofbiochemical markers, we found TFST can significantly reduce the level of FBG(P<0.01),improve insulin resistance conditions, reduce glucagon (P<0.05or P<0.01), increased HDLlevel(P<0.05or P<0.01), reduce TG, CHOL, LDL levels(P<0.05or P <0.01), TFST canstimulate insulin secretion (P<0.05). HE staining showed that TFST can repair the damagedislet cells and liver cells. In order to validate the proteomics results, we examined the levelsof NF-κB signaling pathway downstream inflammatory factors, the results show TFST cansignificantly reduce the level of IL-8, TNF-α, CRP(P<0.05or P<0.01).2, We use highglucose and high insulin to induced insulin resistance of HepG2cells to study themechanism of TFST and AME on the hypoglycemic function.We found that TFST and AMEcan significantly increased glucose consumption rate of insulin resistance cells.148proteinspots were isolated with the method of two-dimensional electrophoresis,20protein spots cutwith3-fold difference by the software analyze to detect mass spectrometry. Results showedthe TFST and AME improve insulin resistance by modulating the pathway of PI3K-Akt,NF-κB and regulating glucose metabolism and reducing liver cell injury. In order to validatethe results of proteomics, we found that TFST and AME can upregulate the expression ofPI3K proteins in the PI3K-Akt signaling pathway, and regulate glucose metabolism-relatedproteins and inflammatory cytokines’ expression.Conclusion: The intervention of TFST and AME on improving insulin resistance ismulti-target and related with PI3K-Akt signaling pathway, NF-κB signaling pathway andglucose metabolism.