transforming-growth-factor-beta and Enterobacteriaceae-Infections

Host defense mechanisms are multi-layered and involve constitutive as well as inducible components. The dissection of these complex processes can be greatly facilitated using a reporter gene strategy with a transparent animal. In this study, we use Caenorhabditis elegans as a model host and introduce a new pathogen-inducible fluorescent reporter involving the promoter of f57f4.4, a gene encoding a putative component of the glycocalyx. We show that this reporter construct does not respond to heavy metal or hypertonic environments, but is specifically and locally induced in the intestine upon Photorhabus luminescens and Pseudomonas aeruginosa infections. We further demonstrate that its upregulation requires live pathogens as well as elements of the nematode p38 MAP kinase and TGF-beta pathways. In addition to introducing a new tool for the study of the interactions between C. elegans and a pathogen, our results suggest a role for the glycocalyx in gut immunity.

Topics: Animals; Animals, Genetically Modified; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Enterobacteriaceae Infections; Genes, Reporter; Glycocalyx; Glycoproteins; Green Fluorescent Proteins; Intestines; MAP Kinase Signaling System; Membrane Proteins; p38 Mitogen-Activated Protein Kinases; Photorhabdus; Promoter Regions, Genetic; Pseudomonas aeruginosa; Pseudomonas Infections; RNA Interference; RNA, Small Interfering; Transforming Growth Factor beta

Proper immune responses are needed to control pathogen infection at mucosal surfaces. IL-22-producing CD4(+) T cells play an important role in controlling bacterial infection in the gut; however, transcriptional regulation of these cells remains elusive. In this study, we show that mice with targeted deletion of the fourth DNA-binding zinc finger of the transcription factor Ikaros had increased IL-22-producing, but not IL-17-producing, CD4(+) T cells in the gut. Adoptive transfer of CD4(+) T cells from these Ikaros-mutant mice conferred enhanced mucosal immunity against Citrobacter rodentium infection. Despite an intact in vivo thymic-derived regulatory T cell (Treg) compartment in these Ikaros-mutant mice, TGF-β, a cytokine well known for induction of Tregs, failed to induce Foxp3 expression in Ikaros-mutant CD4(+) T cells in vitro and, instead, promoted IL-22. Aberrant upregulation of IL-21 in CD4(+) T cells expressing mutant Ikaros was responsible, at least in part, for the enhanced IL-22 expression in a Stat3-dependent manner. Genetic analysis using compound mutations further demonstrated that the aryl hydrocarbon receptor, but not RORγt, was required for aberrant IL-22 expression by Ikaros-mutant CD4(+) T cells, whereas forced expression of Foxp3 was sufficient to inhibit this aberrant cytokine production. Together, our data identified new functions for Ikaros in maintaining mucosal immune homeostasis by restricting IL-22 production by CD4(+) T cells.

Topics: Animals; Citrobacter rodentium; Enterobacteriaceae Infections; Forkhead Transcription Factors; Ikaros Transcription Factor; Immunity, Mucosal; Interleukin-17; Interleukin-22; Interleukins; Mice; Mice, Inbred C57BL; Mice, Knockout; Mutation; Nuclear Receptor Subfamily 1, Group F, Member 3; Receptors, Aryl Hydrocarbon; STAT3 Transcription Factor; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

The gut mucosa hosts large numbers of activated lymphocytes that are exposed to stimuli from the diet, microbiota and pathogens. Although CD4(+) T cells are crucial for defense, intestinal homeostasis precludes exaggerated responses to luminal contents, whether they are harmful or not. We investigated mechanisms used by CD4(+) T cells to avoid excessive activation in the intestine. Using genetic tools to label and interfere with T cell-development transcription factors, we found that CD4(+) T cells acquired the CD8-lineage transcription factor Runx3 and lost the CD4-lineage transcription factor ThPOK and their differentiation into the T(H)17 subset of helper T cells and colitogenic potential, in a manner dependent on transforming growth factor-β (TGF-β) and retinoic acid. Our results demonstrate considerable plasticity in the CD4(+) T cell lineage that allows chronic exposure to luminal antigens without pathological inflammation.

Topics: Animals; CD4-Positive T-Lymphocytes; CD8 Antigens; Cell Differentiation; Cells, Cultured; Citrobacter rodentium; Colitis; Core Binding Factor Alpha 3 Subunit; Enterobacteriaceae Infections; Homeodomain Proteins; Inflammation; Intestinal Mucosa; Intestines; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Signal Transduction; Tamoxifen; Transcription Factors; Transforming Growth Factor beta; Tretinoin

Immaturity of gut-associated immunity may contribute to pediatric mortality associated with enteric infections. A murine model to parallel infantile enteric disease was used to determine the effects of probiotic, Lactobacillus acidophilus (La), prebiotic, inulin, or both (synbiotic, syn) on pathogen-induced inflammatory responses, NF-κB, and Smad 7 signaling. Newborn mice were inoculated bi-weekly for 4 weeks with La, inulin, or syn and challenged with Citrobacter rodentium (Cr) at 5 weeks. Mouse intestinal epithelial cells (CMT93) were exposed to Cr to determine temporal alterations in NF-Kappa B and Smad 7 levels. Mice with pretreatment of La, inulin, and syn show reduced intestinal inflammation following Cr infection compared with controls, which is associated with significantly reduced bacterial colonization in La, inulin, and syn animals. Our results further show that host defense against Cr infection correlated with enhanced colonic IL-10 and transforming growth factor-β expression and inhibition of NF-κB in syn-treated mice, whereas mice pretreated with syn, La, or inulin had attenuation of Cr-induced Smad 7 expression. There was a temporal Smad 7 and NF-κB intracellular accumulation post-Cr infection and post-tumor necrosis factor stimulation in CMT93 cells. These results, therefore, suggest that probiotic, La, prebiotic inulin, or synbiotic may promote host-protective immunity and attenuate Cr-induced intestinal inflammation through mechanisms affecting NF-κB and Smad 7 signaling.

Topics: Animals; Animals, Newborn; Cell Line; Citrobacter rodentium; Enterobacteriaceae Infections; Epithelial Cells; Gastroenteritis; Gene Expression Regulation; Interleukin-10; Intestines; Inulin; Lactobacillus acidophilus; Mice; NF-kappa B; Prebiotics; Probiotics; Signal Transduction; Smad7 Protein; Transforming Growth Factor beta

Enterobacter sakazakii (ES) is an emerging pathogen that causes meningitis and necrotizing enterocolitis in infants. Dendritic cells (DCs) are professional phagocytic cells that play an essential role in host defense against invading pathogens; however, the interaction of ES with DCs is not known. In this study, we demonstrate that ES targets DC-specific ICAM nonintegrin (DC-SIGN) to survive in myeloid DCs for which outer membrane protein A (OmpA) expression in ES is critical, although it is not required for uptake. In addition, DC-SIGN expression was sufficient to cause a significant invasion by ES in HeLa cells and intestinal epithelial cells, which are normally not invaded by ES. OmpA(+) ES prevented the maturation of DCs by triggering the production of high levels of IL-10 and TGF-beta and by suppressing the activation of MAPKs. Pretreatment of DCs with Abs to IL-10 and TGF-beta or of bacteria with anti-OmpA Abs significantly enhanced the maturation markers on DCs. Furthermore, DCs pretreated with various inhibitors of MAPKs prohibited the increased production of proinflammatory cytokines stimulated by LPS or OmpA(-) ES. LPS pretreatment followed by OmpA(+) ES infection of DCs failed to induce maturation of DCs, indicating that OmpA(+) ES renders the cells in immunosuppressive state to external stimuli. Similarly, OmpA(+) ES-infected DCs failed to present Ag to T cells as indicated by the inability of T cells to proliferate in MLR. We conclude that ES interacts with DC-SIGN to subvert the host immune responses by disarming MAPK pathway in DCs.

Topics: Animals; Anthracenes; Bacterial Outer Membrane Proteins; Butadienes; Cell Adhesion Molecules; Cell Survival; Cronobacter sakazakii; Dendritic Cells; Enterobacteriaceae Infections; Enzyme Inhibitors; Epithelial Cells; Flavonoids; HeLa Cells; Humans; Imidazoles; Interleukin-10; Lectins, C-Type; Lipopolysaccharides; Lymphocyte Activation; Microscopy, Electron, Scanning; Microscopy, Electron, Transmission; Mitogen-Activated Protein Kinase Kinases; Nitriles; Pyridines; Rats; Receptors, Cell Surface; Transfection; Transforming Growth Factor beta

Enteropathogenic Escherichia coli (EPEC) is a common pathogen in infantile diarrhea, causing a characteristic histopathologic attaching and effacing (A/E) lesion in the intestinal mucosa. The mouse pathogen Citrobacter rodentium causes a similar A/E lesion in the murine intestine. Like EPEC, C. rodentium infection results in colonic crypt hyperplasia, goblet cell depletion, epithelial proliferation, and mucosal disruption. Using this murine model, we tested the hypothesis that preinoculation of murine gut with Lactobacillus acidophilus early in life can enhance host defense against enteric bacterial infection and attenuate bacteria-mediated colitis. Two-week old BALB/c mice were inoculated with L. acidophilus twice per week for 4 weeks before C. rodentium infection or concomitantly with the exposure to C. rodentium at 6-8 weeks of age. The probiotics were administered twice weekly thereafter. We observed that L. acidophilus inoculation in mice inhibits C. rodentium-induced colitis, which is associated with a decrease in C. rodentium colonization and translocation, an increase in its clearance, and a suppression of colonic myeloperoxidase (MPO) activity. Probiotic treatment also stimulates regulatory cytokine expression in the colon [transforming growth factor beta (TGF-beta), interleukin (IL)-10]. Preinoculation with L. acidophilus is more effective than concomitant use of probiotics in the induction of intestinal IgA secretion and in the downregulation of proinflammatory cytokine expression [tumor necrosis factor alpha (TNF-alpha), IL-6, and IL-12]. These observations suggest that inoculation with probiotics can effectively prevent bacteria-induced colitis by limiting enteric bacteria infection and promoting mucosal protective regulatory immune responses. This study may have ramifications for prevention of infectious diarrhea in human infants and children, particularly in developing countries.

Topics: Animals; Cell Proliferation; Citrobacter rodentium; Colitis; Colon; Enterobacteriaceae Infections; Interleukin-10; Intestinal Mucosa; Lactobacillus acidophilus; Mice; Mice, Inbred BALB C; Probiotics; Transforming Growth Factor beta