| Objective:To observe the therapeutic effect of Astragalus Polysacharin(APS)on hyperlipidemia and investigate the molecular mechanism of APS in the treatment of hyperlipidemia rats based on miR-128-3p/NRF2/antioxidative pathway-mediated lipid metabolism pathway.Method:1.Animal experiments SD rats were randomly divided into normal group,disease model group,simvastatin group and APS group.Rats in the normal group were fed the basal diet,and rats in the other three groups were fed the high-fat diet for 8 weeks.After successful modeling,the APS group and the simvastatin group were intragastrically administered at a concentration of700 mg/kg and 6.7 mg/kg,respectively,while the normal and disease model groups were given equal volume of normal saline.During the experiment,daily indicators such as drinking water,diet,energy intake,excretion,and body weight were recorded.Serum samples were collected every 4 weeks to test blood glucose,triglyceride(TG),total cholesterol(TC),and high density.high-density lipoprotein cholestero(HDL-C)?low density lipoprotein-cholesterol(LDL-C)and malonaldehyde(MDA)levels and alanine aminotransferase(ALT)? aspertate aminotransferase(AST),serum amylase(AMS),and total SOD Enzyme(T-SOD)activity.At the end of the experiment,the rats were sacrificed,and the pancreas and liver tissues of the rats were removed.The pancreatic tissue of the experimental rats was sectioned,the morphology of the islets and acinars was observed by HE staining,and the number and function of islet ? cells were observed by immunohistochemical staining.The liver tissue of the experimental rats was sectioned,HE staining was used to observe the cell morphology and fat deposition of the liver,and hepatic steatosis was graded.At the same time,the contents of TC and TG in the liver tissue of the experimental rats were measured,and the liver and spleen weight index was calculated.In addition,the expression of miR-128-3p in liver tissues was detected by RT-q PCR,and the expression of miR-128-3p target gene NRF2 in liver tissues was detected by Western Blot.2.Liver cell experiment The hyperlipidemia rat hepatocytes(BRL)and human hepatocellular carcinoma(Hep G2)models were constructed at a 2:1 concentration using oleic acid(OA)and palmitic acid(PA).The CCK-8 method was used to determine the optimal drug concentration and time for APS to treat both cells.Two cell models were treated with the optimal concentration and duration of the drug.The levels of TC,TG,MDA and T-SOD,and the expression of miR-128-3p and NRF2 in APS-treated and control groups were determined.Results:1.Animal experiments(1)Dynamic observation of daily lipids and lipids in rats with hyperlipidemia by APS After 8 weeks of high-fat diet,model group had higher the energy intake,body weight and blood lipid levels,and lower food intake,water intake and excretion than in the normal group(p<0.05).After 4 weeks of drug intervention,the levels of TC,TG and LDL-C in the simvastatin group were significantly lower than those in the disease model group(p<0.05).There was no significant difference in lipid levels between the APS group and model group;after 8 weeks of drug intervention,The drinking water,excretion,body weight and blood lipid level of rats in the simvastatin and APS groups were significantly improved compared with those in the disease model group(p<0.05).(2)Effect of APS on pathological changes of pancreatic tissue in hyperlipidemic rats APS can significantly reduce blood glucose and AMS levels in hyperlipidemic rats,but there was no significant improvement in the simvastatin group.HE staining showed that,compared with the normal group,the islet in disease model group and the simvastatin group changed,which the boundary was irregular,the vascular lumen in the islets expanded,and the cells in the islet cells had infiltration of inflammatory cells;islet and acinar cells in APS group rats showed different degrees of improvement in cell arrangement and diffuse edema.Immunohistochemistry showed that,compared with the normal group,the islet cells in disease model group and simvastatin group were irregularly arranged,and there were many breakages and large gaps;APS group had deeper colored and more concentrated in the insulin particles of pancreatic islet ? cells compared with the model group,showing varying degrees of improvement.(3)The effect of APS on liver pathological changes in hyperlipidemic rats APS can reduce transaminase activity in hyperlipidemia rats(p<0.05),but not simvastatin.Morphological observations revealed that,8 weeks after the high-fat diet,the disease model group had an increase liver weight index,blunt margin and white erosion-like surface in liver,which hinted a typical hepatocyte steatosis.After drug treatment,the serum lipid levels,TC and TG contents in liver tissue,liver index,and the number and area of lipid droplets in hepatocytes were significantly reduced in the simvastatin and APS groups.However,only APS can improve the level of transaminase in rats.(4)The effect of APS on the expression of miR-128-3p and NRF2 in liver tissue of rats with hyperlipidemia APS can significantly reduce the serum MDA content and increase T-SOD activity in model rats.The miRNA detection results showed that the expression of miR-128-3p in the serum and liver tissue from the disease model group was significantly upregulated compared with the normal group,but APS could significantly improve this result.The protein expression results showed that the NRF2 protein expression in the liver s was significantly lower in disease model group rat than in the normal group,while the NRF2 protein expression in the liver was significantly higher in APS group rats than in model group.2.Liver cell experiment The results of CCK-8 showed that the optimum concentration of APS in BRL and Hep G2 cells was 150 ?g/m L,and the optimal treatment time was 24 hours.In the high-fat-induced BRL and Hep G2 cell models,the intracellular levels of TC,TG,MDA and miR-128-3p expression were significantly increased(p<0.05),whereas T-SOD activity and NRF2 protein expression were significantly reduce.After administration of APS,the contents of TC,TG,MDA and the expression of miR-128-3p in the two cell models were significantly decreased,while the T-SOD activity and NRF2 protein expression were significantly increased(p<0.05).Conclusion:APS can significantly reduce blood glucose and blood lipid levels in hyperlipidemia model rats.The mechanism may be to improve pancreatic islet and acinar cell pathological changes in pancreas of model rats,and to restore pancreatic function to improve glucose and lipid metabolism;on the other hand,APS inhibits the expression of miR-128-3p in liver tissue of model rats,increases the expression of NRF2 protein in liver tissue,decreases the content of MDA,increases the activity of T-SOD,and increases the antioxidant capacity of liver tissue.Thereby,APS can improve the fat deposition and steatosis of liver tissue,regulate the body's lipid metabolism,and ultimately achieve the purpose of treatment of hyperlipidemia. |
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