The Study Of RegⅢγ On Antagonistic Colitis Mucosal Immune Effect Mediated By Lactobacillus

Intestinal epithelium as the body’s largest mucosal barrier can produce antibacterial factor regenerating islet-derived protein IIIγ(RegIIIγ) which is an important component in intestinal mucosal innate immunity. RegIIIγ as a directly bactericidal C-type lectin, mainly induced by MyD88 and IL-23/IL-22 cytokines, and has a strong bactericidal ability, play an important role in preventing the penetration of commensal bacteria and pathogens and maintaining homeostasis. However, the RegIIIγ expression is unclear in the occurrence and development stage of inflammatory bowel disease. Lactobacillus as a probiotics, have a variety of proposed beneficial effects, which can regulate the intestinal homeostasis and immune balance by interaction with intestinal epithelial thus promoting intestinal health.And in the treatment of inflammatory bowel disease Lactobacillus has also achieved certain efficacies. However, the mechanism of Lactobacillus is not entirely clear, especially for the antibacterial protein RegIIIγ regulatory mechanism, which in the process of interaction between Lactobacillus and ntestinal epithelial cells is not clear.Thus this study mainly analyzing that the expression of RegIIIγ in mice body intervention of Lactobacillus. Initial exploration the role and the molecular mechanism of RegIIIγ in Lactobacillus immune defense and antagonism colitis mucosal immunity. Contents:(1) Effects of multiple Lactobacillus on the expression of RegIIIγ in the normal mouse intestinalThen in this study, firstly used pre-screened the anti-inflammatory activity of multiple Lactobacillus and intervention in healthy mice, then we observed the amount of change of the organizational structure of mouse intestinal, RegIIIγ and cytokines including MyD88, IL-23(IL-12p40, IL-23p19), IL-22, IL-17 A and TNF-α expression before and after the intervention, by HE staining, fluorescence quantitative PCR, immunofluorescence and ELISA techniques.finally evaluated the impact of multiple Lactobacillus on RegIIIγ within health mice intestinal. Discussion the molecular mechanisms of Lactobacillus regulation on intestinal mucosal innate immune factors RegIIIγ and pro-inflammatory cytokines in normal intestinal. In addition used heat-inactivated multiple Lactobacillus and multiple Lactobacillus spent culture medium intervention in healthy mice, then we observed the amount of change of RegIIIγ mRNA by applying fluorescence quantitative PCR. To investigate the effect of multiple Lactobacillus actively secretes metabolites on intestinal RegIIIγ expression levels. The results showed that: when multiple Lactobacillus colonization, the RegIIIγ has high expressions in mice ileum and colon, and there is a significant difference than those in the control group. Histological analysis showed that the ileum and colon tissue was normal after Lactobacillus colonization, and no inflammation produced. For the regulation of the pro-inflammatory cytokines, the study found that multiple Lactobacillus can induce significantly expression of myeloid differentiation factor 88(MyD88), IL-23(IL-23p19 and IL-12p40) and IL-22 in mice ileum and colon compare with the control group. However the IL-17 Aand TNF-α, although has increased in a certain range, but not significantly different from the control group. Furthermore compared with the control group, only live and heat-inactivated multiple Lactobacillus could induce RegIIIγ higher expression in mice ileum and colon, while the multiple Lactobacillus spent culture medium there was no significant.(2) The study of RegIIIγ induced by multiple Lactobacillus on bacterial colitis mucosal immuneIn this study, firstly healthy mice infected with Citrobacter rodentium developed acute colitis and application of different doses of multiple Lactobacillus treatment for colitis in mice(1×1019cfu and 1×1010cfu). During the treatment, monitor weight changes throughout in each group of mice. During treatment every 7 days, mice were sacrificed and remove colon contents and the intestines of each group. Preliminarily research the molecular mechanisms of Lactobacillus regulation of intestinal mucosal innate immune factors RegIIIγ and related factors to play anti-inflammatory effects through measure the cfu of Citrobacter rodentium,determin histological detection of colon and decection the change of RegIIIγ and cytokines MyD88, IL-23(IL-12p40, IL-23p19), IL-22, IL-17 A and TNF-α in the intestinal of colitis model group and multiple Lactobacillus therapy group,by applying fluorescence quantitative PCR, immunofluorescence and ELISA techniques. The results showed that: The cfu of Citrobacter rodentium upregulation gradually in colon, weight lost and colon tissues were severely damaged by the increasing day of the infection. However with the increase of the days of multiple Lactobacillus treatment, the cfu of Citrobacter rodentium decreased significantly, weight gain and the intestinal lesions were significantly ameliorating compared with the colitis model group. Group comparison, the most significant effect of treatment is the group of high-dose multiple Lactobacillus; Compared with the normal control group, Colitis model mice colon RegIIIγ and related factors MyD88, IL-23(IL-23p19 and IL-12p40), IL-22, IL-17 A and TNF-α significantly increased and increased with the degree of inflammation. But colitis model mice after multiple Lactobacillus treatment, RegIIIγ and related factors MyD88, IL-23(IL-23p19 and IL-12p40), IL-22, IL-17 Aand TNF-α significantly decreased in colon, and with the increase of the days of multiple Lactobacillus treatment, the expression level further reduced. Group comparison, the most significant regulation to RegIIIγ and related factors is the group of 1×1010cfu multiple Lactobacillus. Conclusion:A specific component of the structure of Lactobacillus, and not its secreted metabolites may properly activate the cytokines MyD88, IL-23, IL-22 to induce RegIIIγ production in the healthy mouse intestinal, and maintain intestinal homeostasis.The colonization of Lactobacillus is the precondition of the intestinal inflammation remission. Further research showed the mechanism of Lactobacillus remission IBD related with reduction RegIIIγ abnormal expression,but the exact mechanism remains to be further studied.