The Study Of Protective Effect And Mechanism Of Berberine Targeting AMPK To Maintain Mitochondrial Homeostasis Against Myocardial Injury In Diabetic Cardiomyopathy

Background: DCM is a common cardiovascular complication of diabetes.Studies have shown that abnormal energy metabolism caused by long-term hyperglycemia plays an important role in myocardial injury in DCM,however,the mechanism has not been elucidated.Mitochondria are known to be the key organelles responsible for regulating energy metabolism in cells,so maintaining mitochondrial homeostasis is crucial for the treatment of DCM.AMPK pathway is important in regulating of mitochondrial homeostasis,suggesting that the regulation of mitochondrial homeostasis by the AMPK may be a potential therapeutic target of DCM.Berberine has been shown to effectively activate AMPK.Some studies found that berberine can effectively improve the cardiac function of DCM rats,but the mechanism needs to be further studied.Objective: To investigate the protective effect of berberine on myocardial injury in DCM model mice,and explore whether berberine exert its protective effect on DCM cardiomyocytes by targeting AMPK to maintain mitochondrial homeostasis.Methods: DCM animal and cell models were established,and berberine was used for intervention.Measurement of body weight,blood glucose,triglyceride,and total cholesterol concentrations was performed to evaluate glucose and lipid metabolism in mice.HE staining,WGA staining,real-time PCR,and Western blot were conducted to assess myocardial hypertrophy phenotype and marker expression.Exercise tolerance test and cardiac doppler ultrasound were used to detect cardiac function.Transcriptomics analysis was used to explore the relationship between berberine,AMPK,and mitochondrial homeostasis in DCM cardiomyocytes.Metabolomic and lipidomic analyses were conducted to investigate regulatory effects of berberine on overall metabolism.In cell model,AMPK activators,inhibitors,and mitochondrial fission inhibitors were used for intervention,and CCK-8 assay and LDH release were measured to evaluate cardiomyocytes damage.Immunofluorescence,Confocal laser scanning microscopy,Western blot,real-time PCR,and in vitro drug-protein binding assays were used to investigate the protective effects of berberine on cardiomyocytes under high-glucose conditions.Results:Berberine could significantly improve glucose and lipid metabolism and enhanced cardiac function in DCM model mice,and effectively alleviate the cardiomyocyte hypertrophy and damage in both DCM model mice and high glucose-induced cardiomyocytes.Transcriptomic sequencing and enrichment analysis suggested that the cardioprotective effect of berberine may be closely related to the AMPK signaling pathway and the regulation of mitochondrial function.Molecular docking prediction,in vitro drugprotein binding experiments,and western blotting confirmed that berberine could bind to AMPK and effectively activate AMPK.Abnormal expression of mitochondrial fusion and fission-related proteins in cardiomyocytes induced by high glucose led to fragmentation of mitochondria and inhibition of both autophagy flux and mitochondrial biogenesis.Pre-treatment with berberine or AICAR,or co-treatment with Compound C and berberine revealed that berberine could promote the expression of mitochondrial fusion-related proteins,inhibit the expression of mitochondrial fission-related proteins,regulate mitophagy and restore mitochondrial biogenesis,then maintained mitochondrial quality and quantity to perform normal functions.Metabolomics and lipidomics analysis demonstrated that the metabolic regulation of berberine was related to the metabolism of unsaturated fatty acids,choline,triglycerides,and phosphatidylcholine.Berberine had broad and significant metabolic regulatory capabilities.Conclusion: Berberine can significantly improve the cardiac function in DCM model mice,and has a protective effect on high glucose-induced cardiomyocytes injury.Berberine targets and binds to AMPK,then activate the AMPK,and maintain mitochondrial homeostasis by regulating the three key parts(mitochondrial fission/fusion dynamic balance,mitophagy and mitochondrial biogenesis)to protect against DCM cardiomyocytes injury.Moreover,it reveals that berberine also has a good "metabolic regulation" effect at the overall metabolic level,restoring the balance of abnormal choline metabolism and various lipid metabolism in DCM.