The Role Of Interferon-inducible Protein 204 In DSS-induced Colitis Of Mice

Interferon-inducible protein 204(IFI204)is an intracellular DNA receptor that recognizes DNA viruses and intracellular bacteria.It is now known that IFI204 mainly activates two kinds of innate immune signals after recognizing DNA: interferon production and inflammasome activation.Intestinal diseases such as enteritis and colon cancer threaten the health of many animals and humans,and are still one of the major areas of research.In this dissertation,DSS-induced colitis in mice was used as a research model to investigate the regulation of IFI204 on colitis.The study found that IFI204 exerts anti-colitis effect mainly through regulating the composition of intestinal flora,and analyzes that Akkermansia muciniphilia(A.muciniphilia)has a protective effect on the occurrence of colitis.Firstly,the relative expression of IFI204 in various tissues was detected by RT-PCR.The results showed that IFI204 was expressed in the small intestine,caecum and colon.Subsequently,the colitis model was induced by DSS and the IFI204-deficient mice were used to analyze the role of IFI204 in maintaining intestinal homeostasis.As a result,it was found that IFI204-deficient mice are more susceptible to DSS-induced colitis.The main manifestations are: the survival of IFI204-deficient mice was significantly reduced;from the fifth day of DSS-induced colitis,the weight loss of mice was more obvious;the IFI204-deficient mice had higher clinical scores.On the 7th day of DSS-induced colitis,compared with WT mice,IFI204-deficient mice had more severe pathological lesions in the colon,severe edema in the submucosal mucosa,large ulcers in the intestinal mucosa,and severe structural destruction.Subsequently,we further explored the causes of increased susceptibility to DSS-induced colitis in IFI204-deficient mice.The intestinal mucosal barrier includes microbiota,mucus layer,epithelial cell layer,and a variety of immune cells distributed in the submucosa.The mucus layer covers the surface of epithelial cells and isolates microorganisms and food antigens interacts with intestinal epithelial cells to maintain a normal immune response in the intestine.The expression of mucin2,the thickness of the mucus layer,and the number of goblet cells secreting mucin were examined by immunohistochemical methods.The results showed that there was no significant difference in the expression of mucin2 between WT and IFI204-deficient mice in normal.The thickness of mucus layer was similar,and the number of goblet cells was similar.The analysis of the properties of mucins by AB and PAS staining revealed that there was no significant difference in the total mucin expression in IFI204-deficient mice compared to WT mice.However,the acid mucins that played an important role in maintaining the stability of the mucus layer were significantly reduced.The above results indicate that intestinal barrier function is disordered after IFI204 gene deletion.From this result,we can speculate that the increased susceptibility of DSS-induced colitis in IFI204-deficient mice may be due to decreased expression of acid mucin.A large number of intestinal microbes are distributed in the intestinal.These intestinal microbes can compete with exogenous microbes to produce space,and nutrients can then withstand the invasion of pathogenic microbes.In addition,the intestinal microflora interacts with the host to regulate intestinal barrier function and maintain intestinal homeostasis.Therefore,we further analyzed whether the susceptibility of IFI204-deficient mice to DSS-induced colitis was due to differences in the composition of the intestinal microflora.Sequencing analysis of the fecal DNA of two types mice revealed that the two types of the mice were different and located in different communities;the bacteria of the Firmicutes,Bacteroidetes,and Proteobacteria were responsible more than 98% of the total intestinal microflora.Analysis and statistics of the bacteria in these three phyla found that,compared with WT mice,Bacteroidetes and Proteobacteria in the intestinal microflora of IFI204-deficient mice were significantly different,and there was no significant difference in the Firmicutes.The WT and IFI204-deficient mice were then co-housed or cross-fostered to make the intestinal microbiota more consistent.Then colitis was induced by DSS and it was analyzed whether the difference in flora affected the susceptibility of IFI204-deficient mice to colitis.As a result,it was found that the susceptibility of DSS-induced colitis to the two genotyped mice tends to be uniform.Mainly reflected in the DSS induced colitis,after co-housed or cross-fostered the survival rates were consistent,weight loss was consistent,clinical scores were consistent;DSS induction of colitis on the 7th day,no significant difference in pathological lesions of the colon.In addition,there was no difference in mucin2 expression and total mucin expression,and the expression of acid mucins was consistent.Based on the above results,it is known that the increased susceptibility to DSS-induced colitis in IFI204-deficient mice and the decrease of acid mucin are mediated by intestinal microflora,and the susceptibility of DSS-induced colitis can be transferred by microorganisms.The IFI204-deficient mice had different microbiota from the WT mice.The comparison of fecal bacterial DNA16 srRNAs was performed to find the key different bacterial populations.The results showed that the difference among the IFI204-deficient mice was the largest among the bacteria belonging to the A.muciniphilia,its content increased significantly.Then we detected the relative abundance of A.muciniphilia in the faeces of mice by RT-PCR.Significantly,after cohousing and cross-fostering,there was no significant difference in A.muciniphilia detected by the same method.Based on the above results,we speculated whether A.muciniphilia is a key differentiating bacterium and could determine the susceptibility of IFS204-deficient mice to DSS-induced colitis.Therefore,we performed anaerobic culture on A.muciniphilia from the feces of IFI204-deficient mice.We isolate and purificate of A.muciniphilia successfully.Subsequently,A.muciniphilia was replenished into WT mice by gavage,and the effect of the bacterium on the susceptibility to colitis was analyzed,the rate of WT mice increased significantly.In summary,the results of this study show that IFI204 can inhibit DSS-induced colitis,A.muciniphilia one of gut microbiota may mediate the protection of IFI204.