Objective:To investigate the effect of co-colonization with Helicobacter hepaticus ( Hh) and Citrobacter rodentium ( Cr) on CD4 + T cell differentiation. Methods:A HH7-2tg mouse model was employed in this study. Approaches including antibiotic treatment, bacterial colonization, and dextran sulfate sodium (DSS)-induced inflammation were combined to study the impact of specific microbiota on CD4 + T cell differentiation. Flow cytometry was used to analyze the changes in CD4 + T cell subsets in mouse intestinal tissues. Differences between groups were analyzed using the unpaired Mann-Whitney U test (two-tailed). Results:Compared with mice with antibiotic-depleted gut microbiota, co-colonization with Hh and the gut microbiota significantly increased both the proportion and the absolute number of intestinal RORγt + T-bet + Foxp3 + CD4 + T cells (both P<0.01). Co-colonization with Hh and Cr induced significantly higher proportion and absolute number of RORγt + T-bet + Foxp3 + CD4 + T cells as compared with colonization with either Hh or Cr alone following antibiotic treatment (all P<0.05). Under DSS-induced inflammatory conditions, Hh failed to effectively induce RORγt + T-bet + Foxp3 + CD4 + T cells, suggesting that Cr metabolites or inflammatory signals were essential for the induction of this specific cell population. Recombinant Cr strains expressing Hh antigen did not exhibit the induction effect of co-colonization with both wild-type strains in terms of induction efficiency. The induction of this specific T cell population was asynchronous with that of other CD4 + T cell subsets, suggesting the uniqueness of their regulatory mechanism. Conclusion:Co-colonization with Hh and Cr induces RORγt + T-bet + Foxp3 + CD4 + T cells, and thus can serve as a viable in vivo induction model for studying RORγt + T-bet + Foxp3 + CD4 + T cells. This study provides reference for further research into the immune functions and regulatory mechanisms of this unique CD4 + T cell subset.

Objective:To investigate the effect of co-colonization with Helicobacter hepaticus ( Hh) and Citrobacter rodentium ( Cr) on CD4 + T cell differentiation. Methods:A HH7-2tg mouse model was employed in this study. Approaches including antibiotic treatment, bacterial colonization, and dextran sulfate sodium (DSS)-induced inflammation were combined to study the impact of specific microbiota on CD4 + T cell differentiation. Flow cytometry was used to analyze the changes in CD4 + T cell subsets in mouse intestinal tissues. Differences between groups were analyzed using the unpaired Mann-Whitney U test (two-tailed). Results:Compared with mice with antibiotic-depleted gut microbiota, co-colonization with Hh and the gut microbiota significantly increased both the proportion and the absolute number of intestinal RORγt + T-bet + Foxp3 + CD4 + T cells (both P<0.01). Co-colonization with Hh and Cr induced significantly higher proportion and absolute number of RORγt + T-bet + Foxp3 + CD4 + T cells as compared with colonization with either Hh or Cr alone following antibiotic treatment (all P<0.05). Under DSS-induced inflammatory conditions, Hh failed to effectively induce RORγt + T-bet + Foxp3 + CD4 + T cells, suggesting that Cr metabolites or inflammatory signals were essential for the induction of this specific cell population. Recombinant Cr strains expressing Hh antigen did not exhibit the induction effect of co-colonization with both wild-type strains in terms of induction efficiency. The induction of this specific T cell population was asynchronous with that of other CD4 + T cell subsets, suggesting the uniqueness of their regulatory mechanism. Conclusion:Co-colonization with Hh and Cr induces RORγt + T-bet + Foxp3 + CD4 + T cells, and thus can serve as a viable in vivo induction model for studying RORγt + T-bet + Foxp3 + CD4 + T cells. This study provides reference for further research into the immune functions and regulatory mechanisms of this unique CD4 + T cell subset.