| Objective:Xyloketal B is a novel marine compound with antioxidant and anti-apoptotic activity. Previous studies had found that it had the ability to antagonize the lipid aggregates. High fat and high fructose induced NAFLD mice model as well as oleic acid and palmitic acid induced non-alcoholic hepatic steatosis HepG2 cell model were established, then treated with Xyloketal B to observe its pharmacodynamics effects on NAFLD, and explore the mechanism of Xyloketal B treatment for non-alcoholic fatty liver in vitro experiments.Methods:(1) Pharmacodynamics research in vivo. After adapting one week, C57BL6/J mice were randomly divided into two groups:(1) Control group:normal chow diet (n= 16), (2) Model group:high fat diet plus 10% high fructose liquid (HFDL) (n= 48) for 40 days to induce NAFLD. Eight mice from control group and eight mice from model group on 8th weeks were sacrificed in order to confirm whether NAFLD developed or not. After the formation of NAFLD in mice, the model group animals were further randomly divided into five groups, named model group, ator-vastatin group (15 mg/kg/d, ip), low dose group (Xyloketal B,5 mg/kg/d, ip), middle dose group (Xyloketal B,10 mg/kg/d, ip), high dose group (Xyloketal B,20 mg/kg/d, ip). The animals were injected intraperitoneally once daily for 40 days. Fasting blood glucose, blood lipid and other biochemical indicators levels were examined, HE staining to observe liver biopsy, triglyceride (TG) and cholesterol (CHOL) of liver homogenates were detectedby using appropriate detection kit. Meanwhile, free amino acids in serum were analysed by HPLC.(2) Cytological study in vitro. HepG2 cells non-alcoholic hepatic steatosis model was in-duced by 0.5 mM free fatty acid (oleic acid:palmitic acid,2:1). After the model was successfully established, adding different concentrations of Xyloketal B (6.25～100 μM) into cells, TG con-tent in the cells was detected by using the TG kit and lipid accumulation was observed by Oil red O. p-AMPK, AMPK, PPARy, CPT1A, SREBP-1C and its downstream key enzymes in fatty acid synthesis (ACL, ACC1 FAS) protein expression were assayed by western blotting after Xyloket-al B (12.5,25,50 μM) intervented.Result:After feeding with high fat diet plus 10% high fructose liquid for 8 weeks, blood glucose and blood lipid of the mice is obviously increase, the liver of mice appeared the large diffuse fat vacuoles and inflammation, which indicated that the liver developed simple steatosis. Xyloketal B improved the lipid metabolic disorder in NAFLD mice. Xyloketal B administration can signif-icantly decrease BMI, hepatic lipid concentration, blood glucose and improve liver function (P< 0.05). HE staining showed that Xyloketal B with different doses significantly ameliorated fat vacuoles and inflammation in NAFLD mice liver. HPLC metabolism profile combined with pat-tern recognition technology screened out 5 kinds of amino acids with significant differences (glutamic acid, glycine, methionine, isoleucine and valine). Xyloketal B with different doses re-versed the levels of the amino acids in NAFLD mice. In addition,0.5 mM FFA (the ratio of OA and PA,2:1) were determined to treat HepG2 cells for 24 h in order to induce HepG2 cells model of NAFLD. After treated by Xyloketal B for 24 h, intracellular TG and lipid were amelioratived significantly. AMPK was activated, the key enzymes associated with liver fatty synthesis includ-ing SREBP-1C and its downstream targeting enzymes such as ACC1, ACL, FAS, were down-regulated significantly in addition, expression of p-AMPK、PPARy and CPT1A were up-regulated.Conclusion:Xyloketal B showed the good therapeutic effects on NAFLD model in both vitro and vivo, the possible mechanism is AMPK target via down-regulate AMPK-SREBP-1C meta-bolic pathways to reduce fatty acid synthesis and up-regulate AMPK-PPARy-CPTIA metabolic pathways to promote fatty acid oxidation, regulating lipid metabolism, and ultimately reducing the content of TG in the liver to the purpose of treating fatty liver. |
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