Research On Regulatory T Cell-mediated Anti-inflammatory Mucosal Immune Effect By Lactobacillus

Lactic acid bacteria (LAB) as a probiotics has important physiological role and potential applications on prevented and treatment in inflammatory bowel disease (IBD), but the mechanism of LAB on how to enhance or mediate the body’s immune system is not clear. In this study, we first screened the anti-inflammatory activity of lactic acid bacteria and used to intervene in healthy mice using immunomagnetic beads separate the mesenteric lymph node CD4+T cells, detected CD4+Foxp3+Tregs lymphocytes and cytokines by flow cytometry, analysis the effection between dendritic cell and Tregs on mediated ability. Application of mixture of lactic acid bacteria intervention Citrobacter rodentium induced colitis in mice, using immunomagnetic beads, flow cytometry, colon H&E staining analysis the CD4+Foxp3+Tregs in mesenteric lymph nodes of mice changes, and the impaction of inflammatory cytokines, in order to clarify the molecular mechanism of lactic acid bacteria play an anti-inflammatory effect on modulate the immune system, to observe the anti-inflammatory effect of lactic acid bacteria on prevention enteritis by biopsy. And lay the theoretical foundation for the further development of lactic acid bacteria as immunomodulatory agents prevention IBD.1. Screening anti-inflammatory lactic acid bacteria in vitroFreshly isolated MLN from healthy BALB/c mice were made of the single cell suspension and co-cultured with lactobacillus in the cell culture plate, then measured the expression levels of Foxp3, IL-10, IL-12by FACS.The result show that Lactobacillus rhamnosus GG ATCC53103could up-regulated the Foxp3expression. The combination with Lactobacillus plantarum A and Lactobacillus rhamnosus ATCC53103and Lactobacillus acidophilus ATCC4356were up-regulated the IL-10and Foxp3expression.2. Establish the Citrobacter rodentium induced colitis modelApplication the Citrobacter rodentium intervented on healthy Balb/c mice for14days. Then detected the expression of cytokines in the mesenteric lymph nodes of mice was by flow cytometry, H&E staining of mouse distal colonic inflammation situation. The results showed that after the intervention of Cr is mainly caused by a Thl type immune response in MLN and biopsy results show that Cr infected mouse distal colon goblet cells increased and inflammatory cell infiltration, local epithelial cells in the intestine necrosis significantly pathological changes.3. The Molecular Immunology pathway on lactobacillus anti-Cr-induced colitisApplication the anti-inflammatory lactobacillus (ATCC53103+ATCC4356+L.p A) on the female Balb/c mice for21days. Detect the expression of CD4+CD25+Foxp3+Treg and DC in spleen and MLN. Then application Cr intervention on the mice which intervented after the mixed lactobacillus, detection of Treg and related cytokines expression, specimens from the distal colon by H&E staining of colon pathological changes. The results show that mixed lactobacillus can up-regulate the expression of Treg in MLN, and the lactobacillus could induced the DC maturation and differentiation in spleen and MLN. And the lactobacillus mixture could down-regulate the expression of inflammatory cytokines in the MLN after Cr intervention, biopsy results showed that lactobacillus can alleviate Cr for small destruction of rat intestinal damage.