Regulatory Mechanism Studies Of Liver-gut Axis And DNA Methylation In Autoimmune Diseases

Autoimmune diseases,including inflammatory bowel disease(IBD),psoriasis,systemic lupus erythematosus(SLE),rheumatoid arthritis(RA)and so on,are a group of chronic and highly disabling diseases caused by immune disorders,and their pathogenesis is still unclear.These diseases are widely distributed,with an estimated incidence of 80 per 100,000 person-years.In addition,autoimmune diseases are usually accompanied by a variety of complications,such as metabolic abnormalities,cardiovascular diseases and cognitive defects,which have become one of the important factors to increase the global health burden.IBD and psoriasis are two common autoimmune diseases occurring in barrier organs with recurrent episodes.The main clinical manifestations are typical mucosal barrier dysfunction and abnormal permeability,accompanied by extensive infiltration of immune cells in the focus.Current treatments of these two diseases mainly focus on the rapid control of inflammation,prolongation of remission period and improvement of life quality,but the diseases can’t be cured completely.Therefore,aiming at the core pathological changes of IBD and psoriasis,that is,the destruction of the integrity of the"tissue-environment barrier"at the structural and functional levels,we can find the key molecules intervening in this pathological process,which will help to achieve epithelial cell regeneration and repair,correct abnormal differentiation of epidermal cells,and ameliorate mucosal barrier microenvironment,thereby improving the effectiveness of clinical therapeutic drugs.In recent years,the"liver-gut axis"has become a hot topic in the pathogenesis and treatment of digestive tract diseases such as IBD.IBD and hepatobiliary diseases may be interrelated via liver-gut communication and abnormal bile acids(BAs)metabolism is relevant in IBD pathogenesis.Cholic acid(CA)is the earliest primary BA found in human,which is synthesized by sterol 12α-hydroxylase(CYP8B1)through the classical pathway in liver.In this research,we found that BA metabolism was dysregulated in active IBD patients and dextran sulfate sodium(DSS)-induced experimental colitic mice.The contents and proportions of CA,as well as the ratios of12α-OH/non-12α-OH BAs were significantly higher compared to healthy controls.To determine the pathogenic role of CA in colitis,we orally administered CA to DSS-induced ulcerative colitis(UC)mice,trinitrobenzene sulfonic acid(TNBS)-induced Crohn’s disease(CD)mice and interleukin(IL)-10 knockout spontaneous colitic mice.The results showed that CA supplementation potentiated colitis mainly by inducing epithelial mucosal barrier dysfunction,rather than directly regulating immune cells.CYP8B1,which catalyzes CA synthesis,was induced in livers of colitic mice.Through liver-specific overexpression or interference with the expression of CYP8B1,it was found that liver Cyp8b1-overexpressing mice developed more severe colitis with compromised repair of the mucosal barrier,whereas Cyp8b1-knockdown mice were resistant to colitis.Mechanistically,the excessive activation of the CYP8B1-CA axis destroyed the intestinal mucosal barrier and damaged the renewal of leucine-rich repeat-containing G protein-coupled receptor 5 positive(Lgr5~+)intestinal stem cells(ISCs).Next,to gain mechanistic insight into how CA affected the function of Lgr5~+ISCs,metabolomics and lipidomics and RNA sequencing(RNA seq)analyses were applied on isolated crypts of CA-treated colitic mice.It was found that CA impaired Lgr5~+ISCs function through suppression on peroxisome proliferator-activated receptorα(PPARα)-mediated fatty acid oxidation(FAO).At present,there are no agents specifically targeting CYP8B1 on the market.FXR activation could downregulate CYP8B1 expression via different signaling pathways.In this research,to validate the translational significance of our finding,obeticholic acid(OCA),a potent and selective FXR agonist,was used to treat DSS-induced colitis in mice.OCA significantly inhibited the expression of CYP8B1 in liver,as well as decreased the accumulation of CA in intestine,reprogrammed Lgr5~+ISCs renewal,and thus relieved colitis.As a consequence,these results demonstrated that liver FXR and CYP8B1could regulate the development of IBD,and targeting liver FXR and CYP8B1 is expected to become a new strategy for the IBD treatment in the future.Methyl 4-(adenin-9-yl)-2-hydroxybutanoate(DZ2002)is a reversible type III S-adenosyl-l-homocysteine hydrolase(SAHH)inhibitor,which exhibited immunological regulation effects in vivo and in vitro.Previous studies from our team have showed that DZ2002 ameliorated imiquimod(IMQ)-induced psoriasis-like skin inflammation in mice.However,the molecular mechanisms of DZ2002 in the treatment of psoriasis remained to be illuminated.In this research,we further clarified the therapeutic mechanism of DZ2002 in improving psoriasis in mice.The results showed that:1)DZ2002 treatment attenuated hyperplasia,restored differentiation features of epidermis and constrained neutrophil infiltration into psoriasis-like skin;2)Whole genome bisulfite sequencing(WGBS)analysis showed that DZ2002 treatment reversed the abnormal DNA methylation patterns in the psoriatic skin.The methylation level of GATA3 which regulated epidermal differentiation and lipocalin-2(LCN2)which affects neutrophil chemotaxis changed significantly in psoriatic skin by DZ2002 treatment;3)DZ2002 rectified the differentiation of damaged keratinocytes by promoting GATA3 expression and inhibited the expression of chemokine in keratinocytes via reducing LCN2 production partly in psoriasis-like skin;4)DZ2002 directly reduced the levels of LCN2 and chemokine receptors in neutrophils and inhibited their chemotactic migration.In conclusion,our findings established a novel cross-organ regulatory mechanism in IBD pathogenesis,in which excessive CA synthesized by hepatic CYP8B1 impaired Lgr5~+ISCs renewal by repressing PPARα,thus hindering the damaged repair of the intestinal mucosal barrier during colitis.In addition,we used WGBS analysis,combined with keratinocytes and immune cells phenotype analysis,to explain the intervention mechanisms of inhibiting SAHH in psoriasis-like skin to correct abnormal DNA methylation patterns.The above research is of great significance for in-depth understanding of the disease mechanisms of IBD and psoriasis,and the discovery of new therapeutic targets to promote the development of innovative drugs.