| Objective:A mouse model of hyperglycemia and hyperlipidemia induced by diet.To study the regulatory effect of gypenosides,and to investigate the role of bile acid synthesis and transport pathway,which is closely related to lipid and glucose metabolism.Finally,to elaborate the mechanism of gypenosides on glucose and lipid metabolism.The aim of this study was to find a new target for the formulation of gypenosides and its main saponin components Methods:1.Establishment of animal model: Male C57BL/6J mice,freely intake of high fat and high sugar foods(fat: 60%;sugar: 20%;protein 20%)for 16 weeks.And according to the significant change of quantitative indicators,including body weight,serum total cholesterol,blood glucose,serum low density lipoprotein cholesterol.Beides,combining with histopathological examination to determine the end point of model.2.Pharmacodynamics and safety of gypenosides in regulation of glucose and lipid metabolism: After modeling,the mice in the test group were treated with gypenosides(250 mg/Kg,qd)for 2,4,10,16,and 22 weeks.Pathological changes of liver tissues were examined by HE staining.Three groups of positive control group were given rosiglitazone(hypoglycemic drugs,1.2 mg/Kg,qd),simvastatin(cholesterol lowering drugs,20 mg/Kg,qd),fenofibrate(triglyceride lowering drugs,35 mg/Kg,qd).Mice in model group were given equal volume sodium carboxymethyl cellulose solution as negative control.All mice above were fed with high fat and high sugar diet.Another group of mice were fed with normal diet to monitor the basic physiological data.Based on the quantitative indexes of body weight,serum total cholesterol level,blood glucose level,serum low density lipoprotein level.And the histopathological change as the qualitative index to study the effect of gypenosides.At the same time,the levels of serum transaminase and viscera index were monitored to evaluate the safety of long-term administration of gypenosides.3.Study on the expression of genes related to bile acid pathway: We extracted the mRNA from the liver and duodenum of each group of mice.To observe the changes of 11 liver gene expression,which is directly related to the regulation of bile acid synthesis and transport,and then to find out whether the regulation of glucose and lipid metabolism by gypenosides is related to the FXR mediated bile acid pathway.Since FXR→Fgf15 pathway is a unique regulatory pathway in intestinal tract,intestinal expression of FXR and Fgf15 is also included in the study.4.Study on the regulatory effect of gypenosides on bile acid molecules: Expect the expression of genes involved in the synthesis and transport of bile acids,bile acids were also determined in this study.In this part,the changes of the 16 major bile acids in the liver tissues of each group were analyzed by LC-MS/MS method.Then,the correlation analysis of the changes of mRNA and the changes of pharmacodynamics index was carried out to explain the mechanism of gypenosides.Results:1.Establishment of a mouse model of hyperlipidemia and hyperglycemia: Mice were fed with high fat and high sugar diet for 16 weeks,the body weight(P<0.01),blood lipids(P<0.01),blood glucose(P<0.05)levels were significantly increased.The main phenomenon of blood lipid is the increase of serum total cholesterol and low density lipoprotein cholesterol,and glucose tolerance and insulin resistance were also observed in mice fed a high-fat diet,this is similar to human hyperlipidemia.Histopathological examination showed that there were a lot of lipid droplets in the hepatocytes of mice,and the mice were in the early stage of nonalcoholic fatty liver disease.2.To determine the effect of gypenosides on blood lipid and blood glucose levels: After 10 weeks,the blood glucose was decreased significantly by gypenosides(P<0.05).The levels of serum total cholesterol and low density lipoprotein cholesterol were significantly decreased at 16 th weeks(P<0.01).Histopathological examination in 2 weeks showed that gypenosides reduced the size and number of lipid droplets in hepatocytes.The liver index(P<0.05)and aspartate aminotransferase(P<0.05)decreased significantly at 22 th weeks.3.The mechanism of action on glucose and lipid metabolism regulated by gypenosides: High fat diet significantly inhibited the gene expression of CYP7A1 and CYP8B1.As they are key enzyme to generate bile acids,this phenomenon could be explained by the negative regulation of mouse bodies in response to high fat diet pressure.Treated by gypenosides for 16 weeks,gene expression of CYP7A1 and CYP8B1 were elevated,indicating the role of gypenosides on bile acids related pathway.The content of bile acids in mouse liver showed consistent phenomenon with the gene expression data.Moreover,all data was used in principle component analysis and heat map visualization.In PCA result,mouse feed with normal diet,high fat diet,as well as high fat diet and gypenosides could be separated clearly.Thus,gypenosides effect on lipid and carbonhydrate regulation are highly related to bile acids related pathway.Conclusion:Gypenosides has a significant effect on the regulation of glucose and lipid metabolism.Significant changes related to enterohepatic bile acid pathway and the process is one of the current mechanism of gypenosides found reducing blood sugar most likely. |
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