| Object:To investigate whether Enteromorpha prolifera polysaccharide（EP）regulate lipid metabolism of Nonalcoholic fatty liver disease（NAFLD）rats by producing hydrogen sulfide（H2S）.On this basis,the underlying mechanism of hypolipidemic activity via the cystatin-γ-lyase/cystatin-β-synthetase/hydrogen sulfide（CSE/CBS/H2S）pathway was investigated to ameliorate NAFLD.Method:1.Animal model and experimental procedure:50 adult and female SD rats were randomly divided into 5 groups:Control group（Normal diet+H2O）,EP control group（Normal diet+EP）,Model group（High-fat diet+H2O）,EP intervention group（High-fat diet+EP）,Na HS group（High-fat diet+Na HS）,10 rats in each group.Control group and EP control group were fed with normal diet,and the other groups were fed with high fat diet.The other routine conditions of the five groups were the same during 35 days of continuous intervention.The dose of EP was 200mg/kg·BW.And the dose of Na HS was 56μmol/kg·BW.2.Tissue sample collection and the biochemical analysis:paraffin section,Hematoxylin and eosin（HE）staining.3.The levels of TC,TG,LDL-C,HDL-C,H2S and Hcy were measured.4.The activities of CSE and CBS in liver,brain,heart,kidney,spleen and lung were measured respectively.5.The relative expression levels of CSE and CBS protein in liver,brain,heart,kidney and spleen were measured by Western blot.6.The m RNA expression levels of CSE and CBS in liver and brain were measured by Real-time PCR.Result:1.Appearance of liver and Liver index:compared with the liver of the Control group,the liver of the Model group was lighter in color,softer in texture,larger in volume,greasy in cutting edge,heavier in liver weight and higher in liver index（P<0.05）;compared with the Model group,the liver of the EP intervention group and the Na HS group was more ruddy,lighter in weight and lower in liver index（P<0.05）.2.Liver histological alterations were observed by microscope after HE staining,it was showed the obvious histological alterations in the Model group compared with the Control group,such as inflammation,the loss of normal architecture,the increased number of adipodegenerative cells.Compared with the Model group,these structural characteristics were less pronounced in the EP intervention group and the Na HS group.3.The level of TC,TG and LDL-C in serum of the Model group were higher than those of the Control group（P<0.05）,while the level of TC,TG and LDL-C in serum of the EP intervention group and the Na HS group were lower than those of the Model group（P<0.05）.4.The result of the concentration of H2S in serum showed that,compared with the Model group,the EP intervention group and the Na HS group was higher in the rats fed with high-fat diet（P<0.05）.5.The activity of CSE,CBS enzyme in each organ tissue:compared with the Model group,CSE,CBS enzyme activity of liver,brain,spleen,heart and kidney of EP intervention group and the Na HS were higher（P<0.05）.The activitiy of CSE,CBS enzyme were lower than the detection limit（lower than 3.125μmol/L）.6.Compared with the Model group,the expression of CBS and CSE protein was increased in the EP intervention group and the Na HS group in the brain of rats（P<0.05）.Compared with the Model group,the expression of CBS in the EP intervention group was increased significantly（P<0.05）,but the expression of CSE was not significantly affected by Enteromorpha prolifera polysaccharide in the spleen and liver of rats（P>0.05）.At the same time,we observed different effects in the heart and kidney of rats,which the expression of CSE in the EP intervention group was higher than the Model group（P<0.05）,while there was no significant effect on the expression of CBS by Enteromorpha prolifera polysaccharide intervention（P>0.05）.7.Compared with the Model group,the expression level of CBS and CSEm RNA was increased in the EP intervention group and the Na HS group in the brain of rats（P<0.05）.Notably,Enteromorpha polysaccharide significantly increased the expression of CBSm RNA（P<0.05）,but had no significant effect on the expression of CSEm RNA in the liver of rats（P>0.05）.Conclusion:1.In the Model group,the liver index was increased obviously and histological alterations in hepatocytes were appeared.Compared with the Control group,and the levels of TC,TG and LDL-C in serum were increased,indicateing that NAFLD rat model was successfully established in this experiment.2.Enteromorpha prolifera polysaccharide made the liver index decreased,the level of TC,TG,LDL-C in serum decreased,and the histopathological state of hepatocyte steatosis improved in NAFLD rats by increasing the expression of CSE and CBS protein in some organs and tissues,the level of H2S in serum based on CSE/CBS/H2S pathway.3.Treatment with Enteromorpha prolifera polysaccharide has hypolipidemic activity to NAFLD rats,while it was safe to normal rats. |
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