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Background Early life stress (ELS) is an environmental trigger believed to promote increased risk of IBD. Our goal was to identify mechanisms whereby ELS in mice affects susceptibility to and/or severity of gut inflammation. Methods We utilized 2 published animal models of ELS. In the first model, newborn mice were separated from the dam daily for 4 to 8 hours starting on postnatal day 2 and then weaned early on postnatal day 17. Control mice were left undisturbed with the dams until weaning on postnatal day 21. In the second model, dams were fed dexamethasone or vehicle ad libitum in drinking water on postpartum days 1 to 14. Plasma and colonic corticosterone were measured in juvenile and adult mice. Colitis was induced in 4-week-old mice via intraperitoneal injection of interleukin (IL)-10 receptor blocking antibody every 5 days for 15 days. Five or 15 days later, colitis scores and transcripts for Tnf, glucocorticoid receptors, and steroidogenic enzymes were measured. Results Mice exposed to ELS displayed reduced plasma and colonic corticosterone. Control animals showed improvements in indices of inflammation following cessation of interleukin-10 receptor blockade, whereas ELS-exposed animals maintained high levels of Tnf and histological signs of colitis. In colitic animals, prior exposure to ELS was associated with significantly lower expression of genes associated with corticosterone synthesis and responsiveness. Finally, TNF stimulation of colonic crypt cells from ELS mice led to increased inhibition of corticosterone synthesis. Conclusions Our study identifies impaired local glucocorticoid production and responsiveness as a potential mechanism whereby ELS predisposes to chronic colitis in susceptible hosts.

The ICOS pathway has been implicated in the development and functions of regulatory T (Treg) cells, including those producing IL-10. Treg cell–derived IL-10 is indispensable for the establishment and maintenance of intestinal immune homeostasis. We examined the possible involvement of the ICOS pathway in the accumulation of murine colonic Foxp3- and/or IL-10–expressing cells. We show that ICOS deficiency does not impair induction of IL-10 by intestinal CD4 T cells but, instead, triggers substantial reductions in gut-resident and peripherally derived Foxp3+ Treg cells. ICOS deficiency is associated with reduced demethylation of Foxp3 CNS2 and enhanced loss of Foxp3. This instability significantly limits the ability of ICOS-deficient Treg cells to reverse ongoing inflammation. Collectively, our results identify a novel role for ICOS costimulation in imprinting the functional stability of Foxp3 that is required for the retention of full Treg cell function in the periphery.

Tuberculosis (TB), caused by Mycobacterium tuberculosis infection, is a leading cause of mortality and morbidity, causing ∼1.5 million deaths annually. CD4+ T cells and several cytokines, such as the Th1 cytokine IFN-γ, are critical in the control of this infection. Conversely, the immunosuppressive cytokine IL-10 has been shown to dampen Th1 cell responses to M. tuberculosis infection impairing bacterial clearance. However, the critical cellular source of IL-10 during M. tuberculosis infection is still unknown. Using IL-10 reporter mice, we show in this article that during the first 14 d of M. tuberculosis infection, the predominant cells expressing IL-10 in the lung were Ly6C+ monocytes. However, after day 21 postinfection, IL-10–expressing T cells were also highly represented. Notably, mice deficient in T cell–derived IL-10, but not mice deficient in monocyte-derived IL-10, showed a significant reduction in lung bacterial loads during chronic M. tuberculosis infection compared with fully IL-10–competent mice, indicating a major role for T cell–derived IL-10 in TB susceptibility. IL-10–expressing cells were detected among both CD4+ and CD8+ T cells, expressed high levels of CD44 and Tbet, and were able to coproduce IFN-γ and IL-10 upon ex vivo stimulation. Furthermore, during M. tuberculosis infection, Il10 expression in CD4+ T cells was partially regulated by both IL-27 and type I IFN signaling. Together, our data reveal that, despite the multiple immune sources of IL-10 during M. tuberculosis infection, activated effector T cells are the major source accounting for IL-10–induced TB susceptibility.

Inflammatory bowel diseases (IBD) are chronic disorders of the gastrointestinal tract with peak onset during adolescence. The etiology of IBD remains poorly understood, but is believed to involve genetic susceptibility, improper immune regulation, and environmental factors. Psychological stress is one environmental trigger that has been associated with an increased risk of inflammatory diseases. Specifically, previous studies have linked early life stress (ELS) with hypothalamic–pituitary–adrenal (HPA) axis dysfunction leading to adverse health outcomes later in life. Our goal was to determine whether ELS in mice affects susceptibility to, and severity of, colitis – induced subsequent to stress exposure. We used an established mouse model of ELS, maternal separation with early weaning (MSEW), in which newborn mice are separated from their mothers for 4–8 hours daily from day of life (DOL) 2 until early weaning on DOL 17. Then, starting on DOL 28, colitis was induced in MSEW mice and their normally-reared (NR) counterparts by transient blockade of the interleukin-10 receptor (IL-10R). Prior to induction of colitis, MSEW mice showed systemic and colon-specific depletion of corticosterone relative to NR mice. In addition, MSEW mice with active colitis harbored significantly reduced numbers of colonic Foxp3+ T cells compared to colitic NR mice. Finally, whereas NR mice were mostly in remission by 15 days following cessation of IL-10R blockade, MSEW mice showed consistently elevated Tnf in the proximal colon and sustained epithelial damage. Our results suggest that ELS-mediated HPA-axis dysfunction causes hypo-corticosteronism, which may subsequently promote prolonged colonic inflammation in susceptible hosts.

Inflammatory bowel diseases (IBD) are chronic autoinflammatory conditions in which the immune system loses tolerance to gastrointestinal microbes. The role of humoral immunity in IBD is largely undefined. We examined the potential of T-dependent antibodies to prevent intestinal inflammation using mouse models of IBD. For this, we used mice without inducible T cell costimulator ligand (ICOSL). Icosl−/− mice produce significantly fewer T-dependent antibodies. When tested, we found Icosl−/− mice had similar IgA binding but significantly reduced IgG binding to mucosally-associated microbes. Additionally, Icosl−/− mice have more CD4 T cells producing IL-10, an immunosuppressive cytokine, in the colon. We hypothesized that T-dependent antibodies and IL-10 cooperatively promote intestinal homeostasis. To test this, we temporarily eliminated IL-10 producing cells from adult mice. Icosl−/−, but not wild type or IgA−/− mice, rapidly developed colitis under these conditions. This colitis could be prevented if mice were first depleted of Gram-positive or anaerobic bacterial flora. We further investigated the synergy between these T-dependent antibodies and IL-10 utilizing mice with a codeletion of CD4 T cell-derived IL-10 (Il10cKO) and ICOSL. This codeletion resulted in colitis before 28 days of age. However, fostering Icosl−/− Il10cKO mice on wild-type mothers delayed this early-onset inflammation. This suggests T-dependent antibodies conveyed through breastmilk are protective against early-onset colitis. Collectively, our results suggest that IL-10 and T-dependent antibodies toward mucosa-associated Gram-positive anaerobes continually cooperate to maintain intestinal homeostasis throughout the lifespan.

IL-10 produced by T cells and CD11b+ cells utilizes TLR2 signaling and FimA antigen to inhibit early IFN-γ T cell responses to Porphyromonas gingivalis. P.g., a Gram-negative bacterium, is one of the main etiological agents of the chronic inflammatory disease, periodontitis. Disease progression is thought to occur as a result of an inadequate immune response, which although happens locally, can also occur distally as a result of the dissemination of P.g. into the circulation. As IL-10 and TLR2 are pivotal molecules in the immune response that P.g. elicits, we hypothesized that TLR2-mediated IL-10 production, following the initial systemic exposure to P.g., inhibits the IFN-γ T cell response. To address this hypothesis, mice were primed with P.g., and the types of cells producing IL-10 and the capacity of T cells to produce IFN-γ following blocking or neutralization of IL-10 were assessed. Our results showed that upon initial encounter with P.g., splenic T cells and CD11b+ cells produce IL-10, which when neutralized, resulted in a substantial increase in IFN-γ production by T cells. Furthermore, IL-10 production was dependent on TLR2/1 signaling, partly in response to the major surface protein, FimA of P.g. In addition, P.g. stimulation resulted in the up-regulation of PD-1 and its ligand PD-L1 on CD4 T cells and CD11b+ cells, respectively. Up-regulation of PD-1 was partially dependent on IL-10 but independent of TLR2 or FimA. These results highlight the role of IL-10 in inhibiting T cell responses to the initial systemic P.g. exposure and suggest multiple inhibitory mechanisms potentially used by P.g. to evade the hostˈs immune response, thus allowing its persistence in the host.

Extrathymic induction of regulatory T (T reg) cells is essential to the regulation of effector T cell responses in the periphery. In addition to Foxp3, T reg cell expression of suppressive cytokines, such as IL-10, is essential for peripheral tolerance, particularly in the intestines. TGF-β has been shown to induce expression of Foxp3 as well as IL10 and the vitamin A metabolite; all-trans retinoic acid (RA [at-RA]) has been found to enhance the former. We report that in contrast to its enhancement of TGF-β–mediated Foxp3 induction, at-RA potently inhibits the TGF-β–mediated induction of Il10 in naive CD4 T cells. Thus, mucosal DC subsets that are active producers of at-RA inhibit induction of Il10 in naive CD4 T cells while promoting induction of Foxp3. Accordingly, mice with vitamin A deficiency have increased numbers of IL-10–competent T reg cells. Activation of DCs by certain Toll-like receptors (TLRs), particularly TLR9, suppresses T cell induction of Foxp3 and enables induction of Il10. Collectively, our data indicate that at-RA has reciprocal effects on the induction of Foxp3 and Il10 in developing CD4+ T reg cells and suggest that TLR9-dependent inhibition of at-RA production by antigen-presenting cells might represent one mechanism to promote the development of IL-10–expressing T cells.

Development of T helper (Th) 17 cells requires transforming growth factor (TGF)-beta and interleukin (IL)-6 and is independent of the Th1 pathway. Although T cells that produce interferon (IFN)-gamma are a recognized feature of Th17 cell responses, mice deficient for STAT4 and T-bet-two prototypical Th1 transcription factors-are protected from autoimmunity associated with Th17 pathogenesis. To examine the fate and pathogenic potential of Th17 cells and origin of IFN-gamma-producing T cells that emerge during Th17 immunity, we developed IL-17F reporter mice that identify cells committed to expression of IL-17F and IL-17A. Th17 cells required TGF-beta for sustained expression of IL-17F and IL-17A. In the absence of TGF-beta, both IL-23 and IL-12 acted to suppress IL-17 and enhance IFN-gamma production in a STAT4- and T-bet-dependent manner, albeit with distinct efficiencies. These results support a model of late Th17 developmental plasticity with implications for autoimmunity and host defense.

Recent progress in our understanding of mechanisms by which the immunosuppressive cytokine interleukin-10 (IL-10) participates in an ever-increasing diversity of T-cell lineages to maintain immune homeostasis has broadened the framework for defining regulatory and effector T cells and has blurred the lines between them. In this review, we highlight established and emerging roles for IL-10 produced by distinct CD4(+) T-cell lineages that underlie its non-redundant role in curbing immune responses to the intestinal microbiota at steady state and its role to limit T-cell-driven inflammation in responses to pathogens.

CD4(+) regulatory T cells (T(reg) cells) that produce interleukin 10 (IL-10) are important contributors to immune homeostasis. We generated mice with a 'dual-reporter' system of the genes encoding IL-10 and the transcription factor Foxp3 to track T(reg) subsets based on coordinate or differential expression of these genes. Secondary lymphoid tissues, lung and liver had enrichment of Foxp3(+)IL-10(-) T(reg) cells, whereas the large and small intestine had enrichment of Foxp3(+)IL-10(+) and Foxp3(-)IL-10(+) T(reg) cells, respectively. Although negative for Il10 expression, both Foxp3(+) and Foxp3(-) CD4(+) thymic precursor cells gave rise to peripheral IL-10(+) T(reg) cells, with only Foxp3(-) precursor cells giving rise to all T(reg) subsets. Each T(reg) subset developed in IL-10-deficient mice, but this was blocked by treatment with antibody to transforming growth factor-beta. Thus, Foxp3(+) and Foxp3(-) precursor cells give rise to peripheral IL-10-expressing T(reg) cells by a mechanism dependent on transforming growth factor-beta and independent of IL-10.

Crohn's disease and ulcerative colitis are the two major forms of chronic relapsing inflammatory disorders of the human intestines collectively referred to as inflammatory bowel disease (IBD). Though a complex set of autoinflammatory disorders that can be precipitated by diverse genetic and environmental factors, a feature that appears common to IBD pathogenesis is a dysregulated effector T cell response to the commensal microbiota. Due to the heightened effector T cell activity in IBD, developmental and functional pathways that give rise to these cells are potential targets for therapeutic intervention. In this review, we highlight recent advances in our understanding of effector T cell biology in the context of intestinal immune regulation and speculate on their potential clinical significance.