Mechanism On HFD-induced NAFLD In Mice And The Intervention Effect Of Chrysin

Objective:In this study,High-fat diet(60%)was used to induced NAFLD in C57BL/6 mice to explore the mechanism in high-fat diet induced nonalcoholic fatty liver(NAFLD)disease,and to explore the improvement and mechanism of Chrysin on this basis.Method:NAFLD was induced in C57BL/6 mice by feeding a high-fat diet for up to 2months to induce NAFLD model.From the age of 7 weeks,the mice were fed with normal control diet(NCD)or high fat diet(HFD,60% fat content).After 8 weeks,the mice fed with high fat diet were randomly divided into 2 groups according to body weight,namely high fat diet group(HFD)Chrysin intervention group(40 mg/kg)From the beginning of the experiment,the mice were weighed once a week until 20 weeks after the end of the experiment,the fat was collected for weighing,and serum samples were collected for the detection of biochemical indicators(TG TC,HDL-C,LDL-C,ALS,ALT,LDH,HDBH)to evaluate the lipid/liver injury of mice.The levels of TG TC FFA and inflammatory factors such as IL-1β TNF-α and IL-6 in liver tissue were detected by kits.Oil Red O HE Masson Sirius red and F4/80 pathological staining were used to detect liver fat deposition,liver inflammation and liver fibrosis.In addition,we employed Western blot and proteomics to further elucidate the molecular mechanism of HFD-induced NAFLD and Chrysin intervention effect in mice using Graph Pad Prism 9.0 software was used for analysis and drawing.T-test or one-way ANOVA was used for statistical analysis,and the test level was 0.05.Results:1.High-fat diet caused mice hepatic steatosisThe study found that the HFD-fed mice had increased body weight,epididymal fat,FGB,and serum lipid content,and liver dysfunction.The levels of TG,TC FFA in the liver were also significantly increased compared with the NCD group.Western Blot results showed that HFD may induce increased lipogenesis through the AMPK/ACC/SREBP1-c pathway in hepatocytes.2.High-fat diet induced liver inflammation and fibrosis in miceThe Kuffers cells were significantly active by F4/80 staining in the HFD group.The levels of pro-inflammatory factors including IL-1β,TNF-α and IL-6 were increased and Masson and Sirius red staining showed that liver collagen deposition in HFD group.These results indicate that high-fat diet accelerates fatty liver lesions and induces inflammation and fibrosis in mice.In addition,NLRP3/Caspase-1 pathway was significantly activated in hepatocytes of mice in HFD group.These results suggest that high-fat diet may accelerate fatty liver lesion by activating NLRP3/Caspase-1 and induce liver inflammation and subsequent collagen fiber deposition in mice.3.Proteomics analysis of liver in HFD-induced NAFDL miceA total of 543 differentially expressed proteins were identified between NCD and HFD groups.The changes in biological processes were mainly related to lipid metabolism,and the molecular functions were mainly ribosome function,REDOX function,iron ion binding function,various enzyme activity function,etc.4.Chrysin(CN)improved hepatic steatosis in HFD-fed miceChrysin intervention significantly reduced the body weight,FGB,blood lipid and liver dysfunction of NAFLD mice.In addition,compared with the HFD group,the liver/body ratio of the intervention group was decreased,and the contents of TG,TC and FFA in liver tissue were also remarkable decreased.In addition,Chrysin treatment significantly active AMPK and ACC in liver,and reduced the protein expression of SREBP1-c.These results suggest that Chrysin may improve hepatic steatosis in NAFLD mice through AMPK/ACC/SREBP1-c pathway.5.Chrysin(CN)improves HFD-fed liver inflammation and fibrosis in miceChrysin treatment significantly reduced the number of Kuffers and the levels of cellular inflammatory factors.The Sirius red and Masson staining showed that the deposition of collagen fibers in the liver was decreased after Chrysin intervention.Western blot results showed that the expression of NLRP3 inflammasome in the hepatocytes intervention group was inhibited,suggesting that Chrysin may improve the liver inflammation of HFD-induced mice by inhibiting the NLRP3 inflammasome pathway.6.Chrysin(CN)altered the hepatic protein expression profile in HFD-fed mice.Cluster analysis showed that the main biological processes of down-regulated proteins were mainly related to inflammation,immune apoptosis and the up-regulated proteins were mainly related lipid metabolism.Conclusions:1.HFD may induce liver function and hepatic steatosis through AMPK/ACC/SREBP1-c pathway in mice.2.HFD induces liber inflammation and fibrosis in mice through NLRP3 pathway.3.Proteomics showed that HFD-induced hepatic steatosis in mice may be related to liver lipid metabolism,mitochondrial function,endoplasmic reticulum function, aerobic respiration and REDOX reactions.4.Chrysin may improve HFD-induced NAFLD mice through AMPK/ACC/SREBP1-c pathway and NLRP3 pathway.5.Chrysin(CN)altered the hepatic protein expression profile in HFD-fed mice.