Effects Of High Fat Meat Protein Diets On Gut Microbiota,Antioxidant Enzymes Activities And Redox Signaling In Mice

In developed countries,meat composes a significant proportion of the normal diet and consists of 15%of the daily energy intake,40%of daily protein,and 20%of daily fat.Recently,epidemiological investigations and increasing experimental studies have shown that high consumption of red meat,especially processed meat,has been linked to the immune dysfunction,oxidative stress,diabetes,and cardiovascular diseases.Therefore,using different methods and processing meat may aggravate the potential effect on protein oxidation and antioxidant enzyme activity.Whereas the molecular mechanistic events associated with redox status,mucosal immunosuppression and metabolomic markers associated with metabolic syndromes are of growing interest.In this study,the effects of high-fat diet(HFD)supplemented with meat protein on intestinal microorganisms,intestinal permeability,redox status and metabolic syndrome related metabolic markers were studied.1.Effects of high-fat meat protein diet on intestinal microorganisms and biomarkers of oxidative stressThe effects of meat protein in high fat diet on lipid balance,liver metabolism,antioxidant function and intestinal microorganism in mice were studied.Mice were fed a low fat diet contained casein protein(LFC)(Control),a HFD diet incorporated with casein(HFC),fish protein(HFF),or mutton protein(HFM),respectively.The intake of HFM diet increased body weight,caused liver steatosis and oxidative stress,and induced inflammation.The levels of AST,ALT,LPS and leptin,as well as the levels of oxidative damage biomarkers(dityr,AOPPs,3-NT)were significantly higher than those of HFF and HFC diets.In addition,compared with the LFC group,the composition of intestinal microflora in each group fed with HF protein changed a lot.At phylum level,the abundance of Firmicutes was significantly increased in the HFD groups compared with the LFC diet group.At genus level,the high fat fish(HFF)diet had the highest abundances of the beneficial bacteria Lactobacillus and mucin-degrading bacteria Akkermansia than that of the high fat casein(HFC)and high fat mutton(HFM)diets,whereas the abundances of a pro-inflammatory bacteria Escherichia-Shigella and intestinal permeability bacteria Prevotellaceae-003,Deferribacteres and Rikenellaceae RC9 were strikingly increased in HFM fed mice.Thus,HFM intake causes the accumulation of oxidative products,increases body weight,and caused oxidative damage,likely by altering gut microbiota.2.Effects of high-fat meat protein diet on redox state of liver and inflammatory response of intestineMetabolites were extracted and LC-MS-based metabolomic analysis of liver tissues from C57BL/6J mice on low-fat diet(LFD)or a high fat diet(HFD)with casein,or HFD with fish or HFD with mutton protein were performed.Mice fed HFF diet significantly ameliorates hepatic metabolites implicated in inflammation and oxidative stress(?ketohexanoic acid,ketoleucine,kynurenate,hydroxyphenyalacetic acid),gut-derived metabolites(hippurate,catechol indole-3-propinic acid,phenylacetic acid)and bile acid(taurocholate,glycochenodeoxycholate,glycocholate,taurine-BA1)metabolites metabolism that were otherwise increased by HFM diet.Furthermore,we analyzed the gene expression related to intestinal permeability,hepatic redox status,and inflammatory signaling by RT-qPCR.Results showed that HFM diet induced expressions of gene involved in hepatic inflammation enzymes and proteins(CD14,TLR-4,MYD88,TNF-?),while down-regulated expressions of gene involved in intestinal permeability enzymes and proteins(ZO-1,Occludin,Claudinl)than that of the HFC and HFF diets.The potential associations of these altered metabolites with serum endotoxin,inflammatory biomarkers,and intestinal permeability genes to further confirm their role in hepatic metabolism.Thus,meat proteins in high fat diet interact in host to create distinct responses in gut microbiota and metabolites.3.Glutaredoxin-1 deficiency causes hepatic lipogenesis and dyslipidemia in association with impaired antioxidant enzymes activities induced by high fat meat proteins in miceThe effects of high-fat meat protein diet on liver antioxidant capacity,proinflammatory cytokines and Nrf2/Keap1 signal pathway regulators in Glrx1-/-mice were studied.The results of biochemical and histological analyses indicated that intake of HFM increased hepatic total cholesterol,triglycerides,serum alanine transaminase and aspartate transaminase,and macro-and micro-vesicular lipid droplets accumulation,which are accompanied by altered gene expression associated with lipid(ACC1,Srebfl,CD36,FASn)and cholesterol metabolism(Srebf2,HMGCR,ACoXl).Furthermore,hepatic and colon gene expression of Nrf2/keapl signaling pathway and its downstream signaling targets were determined using RTqPCR.Glrx1 Knock Out(KO)increased Nrf2 activity and expression of its target genes(GPx,catalase,SOD1,G6pd,and Bbc3),which was exacerbated by intake of HFM diet.Glrx1-/-mice fed HFF diet significantly ameliorated diet-induced NAFLD biomarkers compared to HFC and HFM diets.4.Glutaredoxin-1 deficiency promotes potential metabolomic biomarkers associated with NAFLD induced by high fat meat proteinsThe purpose of this study was to verify the effect of high-fat meat protein diet on the metabolic pathway of Glrx1-/-mice.Glrx1-/-mice fed HFF diet significantly ameliorated diet-induced NAFLD biomarkers compared to HFC and HFM diets.In addition,serum metabolome profiling identified metabolites specifically associated with lipid metabolism(lysophosphatidyl cholines and lysophosphatidyl ethanolamine),bile acids metabolism,sphingolipid and amino acid metabolism pathways.A HFM diet increased the abundances of LysoPC(15:0),LysoPC(16:0),LysoPC(17:0),LysoPC(20:1),LysoPE(18:2),LysoPE(22:0),LysoPE(20:6),O-arachidonoylglycidol,12-ketodeoxycholic acid and sphinganine that are associated with NAFLD.The KEGG metabolic pathway of identified metabolites of high fat diets showed that the differential metabolites were associated with lipid metabolism,linoleic acid metabolism,amino acid metabolism,bile acid metabolism,sphingolipid metabolism,glycine and serine metabolism and glutathione metabolism pathways whereas HFF diet ameliorated NAFLD by modifying these pathways.In conclusion,dietary protein oxidative stress induced by mutton protein and glutaredoxin-1 deficiency may be the important reason of the potential metabolomic biomarkers associated with NAFLD.