| Objective:By establishing a model of non-alcoholic fatty liver disease(NAFLD)rats,the effects of different doses of lycopene supplementation on the glycolipid metabolism and iron overload of non-alcoholic fatty liver disease rats were analyzed,and the possible mechanism of action was explored.Methods:1.Animal model establishment and grouping: Sixty 6-week-old SPF male SD rats were selected,weighing 180-220 g.After one week of adaptive feeding,rats were randomly divided into blank control group,model group,lycopene 20mg/(kg·d)group and lycopene60mg/(kg·d)group according to body weight,15 rats in each group.The blank control group was fed with ordinary feed,and the remaining 3 groups were fed with high-fat and high-fructose feed for 4 weeks to establish a non-alcoholic fatty liver disease model.After successful modeling,the model group: continued to be fed with high-fat and high-fructose feed and given equal volume of olive oil;lycopene 20 mg/(kg·d)group: high-fat and high-fructose diet,20 mg/(kg·d)lycopene intragastric administration;lycopene 60mg/(kg·d)group: high-fat and high-fructose diet,60 mg/(kg·d)lycopene intragastric administration.The blank control group continued to be fed with ordinary feed and given equal volume of olive oil;A total of 8 weeks of intervention.After the last gastric irrigation,fasted for 12 h,the abdominal cavity was injected with 10% hydrate chlorinal anesthetic,taking blood from the abdominal aorta,separating the serum for cryopreservation,and the liver was left to make pathological section specimens,the remaining tissue was frozen in liquid nitrogen and turns to the refrigerator at-80 ?for storage.2.Detection of blood lipid and liver function indexes: Serum levels of high-density lipoprotein cholesterol(HDL-C),low-density lipoprotein cholesterol(LDL-C),total cholesterol(TC),triglyceride(TG),alanine aminotransferase(ALT)and aspartic acid aminotransferase(AST)were measured by automatic biochemical analyzer.3.Blood glucose and insulin levels: Blood glucose test paper method,radioimmunity method were used respectively to detect Fasting glucose(FBG),Fasting insulin(INS)level in each group,and insulin resistance index(HOMA-IR)was calculated.4.Detection of inflammatory factor level: Serum interleukin-6(IL-6),interleukin-18(IL-18)and interleukin-1?(IL-1?)expression were measured by flow cytometry.5.Pathological observation of liver tissue: HE staining was used to observe the pathological changes of liver tissue in rats.6.Detection of iron level: Inductively coupled plasma-mass spectrometry(ICP-MS)was used to detect serum iron and liver iron in each group of rats.7.Real-time quantitative PCR method was used to detect the gene expression levels of Hemojuvelin(HJV),Hepcidin,and Transferrin receptor 1(Tf R1)genes in the liver of rats.8.Western blotting was used to detect the expression levels of Hepcidin and Tf R1 protein in the liver.Results:1.Establishment of non-alcoholic fatty liver disease model: After 4 weeks of feeding with high-fat and high-fructose diet,compared with the control group,the serum levels of TG and LDL-C in the model group were significantly increased(P<0.05),indicating that lipid metabolism disorders occurred;The liver of rats in the blank control group had thin edges,bright red and smooth surface;In the model group,the liver had significantly larger,slightly blunt edges and pale yellowish colors with a greasy cut surface.Observation under light microscope showed that the hepatic lobules in the blank control group were clear,the hepatic cords were arranged neatly,and there were almost no fat vacuoles;While in the model group,hepatic cells with a unit area of more than 1/3 were fatty,and hepatic cells were significantly enlarged.2.Blood lipid and liver function indexes: Compared with the blank control group,the levels of LDL-C,TG,TC and ALT in the model group increased by 83.78%,93.02%,38.58% and 34.21%,respectively,and the differences were statistically significant(P<0.05).Compared with the model group,the serum TG and ALT levels of rats in the 20 and 60 mg /(kg·d)lycopene intervention groups were significantly reduced after 8 weeks of intervention(P<0.05);The levels of LDL-C and AST showed a decrease,but the differences were not statistically significant(P>0.05).3.Blood glucose and insulin: Compared with the blank control group,the FBG,INS,and HOMA-IR of the model group rats increased by 6%,36%,and 38%,but the differences were not statistically significant(P>0.05);Compared with the model group,the FBG level of the 20 and 60 mg/(kg·d)lycopene intervention groups decreased,and the differences were statistically significant(P<0.05).The INS level and HOMA-IR showed a downward trend,but the differences were not statistically significant(P>0.05).4.Inflammatory factors: Compared with the blank control group,IL-6,IL-1? and IL-18 levels in model group were significantly increased(P<0.05).Compared with model group,levels of IL-6 and IL-1? in the 20,60 mg/(kg·d)lycopene intervention groups decreased significantly(P<0.05),and level of IL-18 decreased,but there was no significant difference(P>0.05).5.The results of HE staining showed that: Under the light microscope,the liver tissue structure of rats in blank control group was not significantly abnormal,and the hepatic cords were arranged neatly.In the model group,the liver cells of the model group became larger,and most of the cells appeared fatty degeneration,hepatic cords were arranged disorderly,and there was a lot of fat Vesicles and inflammatory cell infiltration.20,60mg/(kg·d)lycopene intervention groups reduced the degree of fatty degeneration of liver cells,reduced inflammatory cells,and significantly reduced the number of fat vacuoles in the cytoplasm,and the improvement effect of liver tissues was more significant in60mg/(kg·d)lycopene intervention group.6.Iron level index: Compared with the blank control group,the serum iron level of the model group increased slightly,but the difference was not statistically significant(P>0.05),liver iron level increased by 77.06%(P<0.05);Compared with the model group,the liver iron levels of the 20 and 60 mg/(kg·d)lycopene intervention group were significantly reduced(P<0.05).7.PCR results showed that: Compared with the blank control group,the HJV and Hepcidin gene expression levels in the liver tissue of the model group were significantly reduced,and the Tf R1 gene expression level was significantly increased(P values are less than 0.05);Compared with the model group,Tf R1 gene expression levels in 20,60mg/(kg·d)lycopene intervention groups liver tissue were significantly reduced,HJV and Hepcidin gene expression expression levels in 60 mg/(kg·d)lycopene intervention group were significantly increased(P values are less than 0.05).8.Western Blotting results showed that: Compared with the blank control group,the expression level of Hepcidin protein in the liver tissue of the model group was significantly reduced,and the expression of Tf R1 protein was significantly increased(P values are less than 0.05);Compared with the model group,the expression of Tf R1 protein in liver tissue of the 20 and 60 mg/(kg·d)lycopene intervention groups were significantly reduced,and the expression level of Hepcidin protein was significantly increased(P values were less than 0.05).Conclusion:1.Lycopene supplementation can effectively improve metabolism of glycolipid and liver damage in NAFLD rats,reduce inflammation,and the improvement effect of high-dose lycopene is more significant.2.Lycopene supplementation can improve iron overload in NAFLD rats,and the mechanism may be related to the regulation of the expression of liver Hepcidin,HJV,Tf R1 and others. |
PDF Full Text Request