| Background:Type 2 diabetes mellitus（T2DM）is a common chronic metabolic disease,which is characterized by hyperglycemia and insulin resistance.Hepatic gluconeogenesis,as a result of insulin resistance,is an important cause of hyperglycemia.Inhibiting hepatic gluconeogenesis is an important treatment for T2DM.AKT signaling pathway plays a key role in insulin resistance and is an important marker of T2DM.PEPCK and G6Pase are the key rate limiting enzymes in hepatic gluconeogenesis.AKT/PEPCK/G6Pase axis plays a key role in liver gluconeogenesis,and is expected to become a target for the treatment and drug development of T2DM.The AKT/PEPCK/G6Pase axis is used as a target to screen drugs with anti hepatic gluconeogenesis from traditional Chinese medicine or an important means to promote drug research and development.Oxymatrine is widely used as an adjunctive drug for obesity related diseases,and it is a potential drug for the treatment of hepatic gluconeogenesis.However,there are few reports on whether Oxymatrine has anti gluconeogenesis effect and its mechanism.OBJECTIVE:The purpose of this study was to investigate the role of AKT/PEPCK/G6Pase axis in hepatic gluconeogenesis and the specific mechanism of oxymatrine against hepatic gluconeogenesis,so as to provide new therapeutic targets and drugs for the treatment of T2DM.METHODS:Animal experiment:The objective of this study was to investigate the role of AKT/PEPCK/G6Pase axis in hepatic gluconeogenesis and the specific mechanism of oxymatrine against hepatic gluconeogenesis,so as to provide new therapeutic targets and drugs for the treatment of T2DM.The experiments were divided into control group,T2DM model group,Low dose intervention group of oxymatrine（10 mg/kg BW）,High dose intervention group of oxymatrine（20 mg/kg BW）,and Intervention group of metformin（250 mg/kg BW）.The gluconeogenesis of HepG2 cells and the protective effect of oxymatrine were assessed by measuring the glucose production and the uptake of 2-NDBG;the phosphorylation level of AKT,the m RNA and protein expression levels of PEPCK and G6Pase were detected by real-time PCR and Western blotting,and the expression changes of the above molecules and the regulation of oxymatrine on the expression of them were analyzed.Cell experiment:HepG2 cells were treated with high concentration glucose（55 mm）to mimic the occurrence of T2DM.The experiments were divided into Control group,High glucose treated group（55 mm）,Low dose intervention group of oxymatrine(10-7 m),High dose intervention group of oxymatrine(10-6 m),Intervention group of oxymatrine combined with AKT inhibitor（mk-2206）and Intervention group of metformin(10-6m).The gluconeogenesis of HepG2 cells and the protective effect of oxymatrine were assessed by measuring the glucose production and the uptake of 2-NDBG;the phosphorylation level of AKT,the m RNA and protein expression levels of PEPCK and G6Pase were detected by real-time PCR and Western blotting,and the expression changes of the above molecules and the regulation of oxymatrine on the expression of them were analyzed.RESULTS:（1）The fasting blood glucose of T2DM rats was up to 16.9 m M compared with the fasting blood glucose of 6.1 m M in the control group,and Oxymatrine was able to reduce the fasting blood glucose of T2DM rats in a dose-dependent manner.（2）Glucose tolerance was significantly impaired and K values were significantly lower in T2DM rats compared with controls（P<0.001）,whereas Oxymatrine dose dependently improved glucose tolerance and increased K values in T2DM rats（P<0.05 or P<0.01）.（3）The phosphorylation of AKT was significantly decreased（P<0.001）and the expression levels of PEPCK and G6Pase were significantly upregulated（P<0.001）in T2DM rats compared with controls,whereas Oxymatrine could dose dependently increase the phosphorylation of AKT and decrease the expression of PEPCK and G6Pase（P<0.05 or P<0.01）.（4）Monocrotaline significantly reduced body weight（P<0.001）,increased liver weight（P<0.001）,and increased hepatic index（P<0.001）and induced fibroid like changes in liver tissue architecture compared with the control rats,whereas Oxymatrine effectively protected the liver in a dose-dependent manner Histoimmunomorphological alterations.（5）Compared with the control,the uptake of 2-NBDG by the high glucose treated HepG2 cells was significantly reduced（P<0.001）and the production of glucose was significantly increased（P<0.001）,while Oxymatrine could significantly increase2-NBDG uptake and reduce glucose production（P<0.05 or P<0.01）in a dose-dependent manner in HepG2 cells.CONCLUSION:AKT/PEPCK/G6Pase axis plays an important role in the pathogenesis of type 2 diabetes mellitus;The inhibitory effect of oxymatrine on hepatic gluconeogenesis involves the expression regulation of PEPCK and G6Pase,and its mechanism depends on AKT phosphorylation. |
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