Chronic Jet-lag Changes Colon Microbiome And Mycobiome And Promotes Progression Of MAFLD In HFHFD-fed Mice

Background and objective: With the development of urbanization and the change of lifestyle,the incidence of metabolic associated fatty liver disease(MAFLD)has been increasing.About 25% of the world’s population is now affected by the disease.With the progression of MAFLD,it can even lead to hepatocellular carcinoma,which seriously endangers public health.The circadian clock is closely related to metabolic homeostasis,and circadian disruption aggravates the progression of metabolic diseases,but the specific mechanism is still unclear.This study aims to observe the effect of circadian disruption,induced by chronic jetlag,on the pathogenesis and progression of MAFLD in high-fat and highfructose diet(HFHFD)fed mice,and to explore the related microbiotic mechanisms.Methods: Forty male C57BL/6 mice were randomly divided into 4groups: 1)mice on a normal diet exposed to a normal circadian cycle(NDNC);2)mice on a normal diet undergo experimental chronic jet-lag(NDCJ);3)mice on a HFHFD and exposed to a normal circadian cycle(HFHFD-NC);4)mice on a HFHFD undergo experimental chronic jet-lag(HFHFD-CJ).Changes in food intake,water intake and body weight were recorded weekly.At the end of the 16 th week,mice were sacrificed.Then blood lipids,serum inflammatory factors,insulin resistance,serum transaminase,and liver pathology were compared among groups.Microbiome and mycobiome of colon contents were analyzed by 16 S and ITS2 sequencing.The microbial diversity,composition,relative abundance at different levels of gut bacteria and fungi were compared among groups,and network analysis was used to analyze the interaction between the microbial communities.Results:(1)Body weight of mice was significantly higher in ND-CJ group vs.ND-NC mice in 9-11 th week and after the 15 th week.Body weight of mice was significantly higher in HFHFD-CJ group vs.HFHFD-NC group at the8-11 th week and the 14 th week.(2)The body weight,liver weight,liver weight to body weight ratio,visceral fat weight and visceral fat to body weight ratio in HFHFD-CJ group were significantly higher than those in HFHFD-NC group.The area under the curve of IPGTT at 16 th week was significantly higher in the HFHFD-CJ group vs.HFHFD-NC group.(3)The serum inflammatory factors IL-6 and TNFα were significantly higher in HFHFD-CJ group vs.HFHFD-NC group.The fasting blood glucose and HOMA-IR index were significantly higher,and the QUICKI index was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.(4)The liver pathological damage was much more serious in HFHFDCJ group vs.HFHFD-NC group.The NAS was significantly higher in HFHFD-CJ group vs.HFHFD-NC group.(5)The m RNA expression of hepatic Clock was significantly higher in HFHFD-CJ group vs.HFHFD-NC group,the m RNA expression of hepatic Bmal,Cry1 and Cry2 was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.(6)The 16 S bacterial sequencing results showed that the α-diversity of intestinal bacteria was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.The relative abundance of genus Helicobacter,Enterococcus,Klebsiella and Rhodococcus was significantly higher in HFHFD-CJ group vs.HFHFD-NC group.The relative abundance of genus Lactococcus,Akkermansia,Sutterella,Adlercreutzia,Turicibacter,Clostridium and Bifidobacterium was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.(7)The results of ITS2 fungal sequencing results showed that the αdiversity of intestinal fungi was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.The relative abundance of genus Aspergillus and Blumeria was significantly higher in HFHFD-CJ group vs.HFHFD-NC group.The relative abundance of genus Wickerhamomyces,Ganoderma,Pestalotiopsis,Sarocladium and Preussia was significantly lower in HFHFD-CJ group vs.HFHFD-NC group.(8)KEGG pathway analysis showed that there were significant differences in amino acid metabolism,steroid synthesis,endoplasmic reticulum protein processing and other signaling pathways in HFHFD-CJ group vs.HFHFD-NC group.(9)Network correlation analysis of gut microbes showed that the interactions between bacteria were significantly decreased in HFHFD-CJ group vs.HFHFD-NC group;the positive related interactions between fungi and bacteria were significantly increased,while the negative related interactions between fungi and bacteria were significantly decreased in HFHFD-CJ group vs.HFHFD-NC group.Conclusion:(1)Chronic jet-lag promotes obesity,disorders of glucose metabolism,dyslipidemia,insulin resistance,liver inflammation,and fibrosis in HFHFD-fed mice,and accelerates the progression of MAFLD.(2)Construction of chronic jet-lag mice model in HFHFD-fed mice by periodically changing the onset of light lead to disturbed expression of circadian clock genes in the liver such as Clock,Bmal1,Cry et al.(3)Chronic jet-lag changes the diversity,composition,relative abundance of the intestinal microbiome and mycobiome and affects the interaction between gut microbes,which may be related to the progression of MAFLD.