Effects Of NMN On Intestinal Barrier And Intestinal Flora In Mice With DSS Induced Enteritis

Objective:Nicotinamide mononucleotides(NMN)have physiological effects on mammals by converting to nicotinamide adenine dinucleotides(NAD~+).Inflammatory bowel disease(IBD)is a chronic nonspecific inflammatory bowel disease with multiple causes.The purpose of this study was to observe the therapeutic effect of NMN on IBD in mice and its effects on intestinal mucosal barrier and intestinal flora.Methods:Thirty-two 8-week-old C57BL/6J male mice were divided into normal control group(CON),disease group(DSS),early treatment group(N+D)and post-disease treatment group(D+N),with 8 mice in each group,administered daily for 3 weeks.Starting from the modeling day,mice in DSS group,N+D group and D+N group were given free drinking solution of 3%sodium dextran sulfate(DSS)for7 days,while mice in CON group were given distilled water,and mice in N+D group were given NMN.On day 8,the rats were fed by gavage,the CON and DSS groups were given distilled water,and the N+D and D+N groups were given NMN.The concentration of NMN was 2.5mg/m L,and the intragastric dose was 0.2ml.The treatment group included full 21 days of administration(N+D)and the last 14 days of administration(D+N).Body weight,fecal characteristics and occult blood of mice were recorded daily for 3 weeks to calculate DAI score.6 hours before the end of the experiment,FD4 was administered to mice to conduct in vivo imaging tracer experiment,and the effect of NMN on intestinal mucosal permeability of mice was preliminatively analyzed.Subsequently,the eye blood of mice was collected,the supernatant was collected,and the expression levels of serum inflammatory cytokines and NAD+were detected by ELISA.Mice feces were collected for 16S r DNA sequencing to analyze the intestinal microbial composition and expression differences,and to explore the effect of NMN on intestinal microflora of mice with enteritis.Colon tissue of mice was isolated and colon length was measured.HE staining was used to observe enteritis,and Alcian blue staining was used to observe changes in intestinal permeability.Subsequently,Immunohistochemical staining further investigated the role of NMN in the expression and localization of junction proteins(Claudin-1,ZO-1,Ocludin and E-Cadherin).Finally,correlation analysis was conducted between intestinal flora with significant abundance changes and serum inflammatory factors and intestinal mucosal connexin content obtained by detection to explore their correlation.Results:(1)ELISA showed that NMN increased serum NAD~+concentration in mice;Serum proinflammatory factors(IL-6,TNF-α)expression decreased.(2)Colonic length of mice treated with NMN recovered slightly and disease activity was relieved.HE staining showed that the pattern of intestinal inflammation was improved.(3)In vivo imaging experiments preliminatively showed that NMN could reduce intestinal mucosal permeability,and Alcian blue staining showed NMN intake increased the number of goblet cells and mucus level.Immunohistochemical analysis showed that NMN increased protein levels of claudin-1,Occludin,ZO-1 and E-Cadherin(E-Ca)and improved intestinal barrier function in mice.(4)16Sr DNA sequencing results showed that the composition and abundance of intestinal flora in IBD mice also changed significantly.Increased abundance of Firmicutes,Verrucomicrobia,Akkermansia and Lactobacillus,which are considered beneficial bacteria,But the abundance of Bacteroidetes and Muribaculaceae decreased obviously.(5)After NMN treatment,the levels of serum inflammatory factors(IL-6,TNF-α)in IBD mice were significantly positively correlated with Bacteroides,etc.And(Firmicutes,Verrucomicrobia,Akkermansia,Lactobacillus)were significantly negatively correlated.The content of intestinal mucosal compact junction protein(Claudin-1,E-Ca,Occludin)has a significant negative correlation with Bacteroides,etc.It was significantly positively correlated with the intestinal microbe(Firmicutes,Verrucomicrobia,Akkermansia and Lactobacillus).Conclusions:After NMN treatment,intestinal flora was correlated with intestinal mucosal barrier and intestinal inflammation,respectively.NMN can relieve inflammation,repair intestinal mucosal barrier and improve intestinal flora disorder in mice with DSS induced colitis.