Study On The Hypoglycemic Effect And Mechanism Of Marine Natural Product BDDPM Targeting PTP1B

In this thesis,the enzymatic activity and hypoglycemic activity in vivo of a marine-derived natural product were studied by pharmacokinetic experiments,cell biological experiments and animal experiments.The two main research objects are Protein Tyrosine Phosphatase 1B?PTP1B?and the natural product BDDPM.Type 2 diabetes mellitus?T2DM?is a severe metabolic syndrome caused primarily by insufficient insulin secretion and insulin resistance and characterized by hyperglycemia and hyperinsulinemia.PTP1B is a widely-recognized potential therapeutic target for type 2 diabetes,which dephosphorylates key phosphorylated proteins in the insulin signaling pathway,causing insulin resistance in tissues and organs involved in glucose metabolism,thus inducing T2DM.The marine natural product BDDPM exhibits potent PTP1B inhibitory activity(IC₅₀=2.4?M)in enzymology and has significant pharmacological research value.The research on the binding characteristics between BDDPM and PTP1B,the cytotoxicity and cellular activity of BDDPM as well as the hypoglycemic effect of BDDPM in animals are beneficial to provide scientific basis and guidance for the development of new hypoglycemic agents.In this study,we verified the inhibitory type of BDDPM against PTP1B and the binding characteristics in vitro by enzyme kinetics experiments and Biacore molecular interaction technique?SPR?.The molecular docking simulation technique was conducted to predict and analyze the binding site and mode of action between them.Then the toxicity of BDDPM to C2C12 cells was tested by cytotoxic MTT assay,and the effect of BDDPM on intracellular insulin signaling pathway was verified by Western blotting.Based on enzymology and cell biology experiments,we further studied the hypoglycemic effect of oral BDDPM in type 2 diabetic mice through mouse experiments?including fasting blood glucose measurement,insulin tolerance test,tissue protein Western blotting test,HE staining and immunohistochemistry experiments?such as lowering fasting blood glucose levels,ameliorating insulin resistance,lipid-lowering effects,and effects on the structure and function of various tissues and organs in diabetic mice.The experimental results demonstrate that BDDPM is a PTP1B competitive inhibitor which is able to specifically bind to PTP1B in vitro?affinity constant:2.9?M?.BDDPM can also embed the catalytically active region of PTP1B,and the binding between them is maintained by hydrogen bonds.Compared with the representative proteins of the PTPs protein family,BDDPM has a significantly high selectivity for the inhibition of PTP1B.Therefore,BDDPM is a highly specific competitive inhibitor of PTP1B.BDDPM has almost no significant effect on the cell viability of C2C12,and can significantly increase the phosphorylation level of proteins associated with the insulin signaling pathway,thus improving Insulin resistance status of C2C12 cells.BDDPM administration is able to significantly reduce fasting blood glucose levels in type 2 diabetic mice,and can improve insulin resistance and lipid metabolism abnormalities in tissues and organs associated with glucose metabolism.BDDPM not only does not adversely affect vital organs in animals,but functions to improve the structure of liver and pancreas tissues as well as promote hepatic glycogen production and improve pancreatic insulin secretion.In summary,we explored the enzymatic and binding kinetics characteristics,cellular cytotoxicity and activity as well as hypoglycemic and lipid-lowering effects of the marine-dervied natural product BDDPM,providing a valuable experimental evidence for exploring it as a potential hypoglycemic medicine.