Heterogeneity of non-lymphoid cells expressing HLA-D region antigens in human fetal gut.

Human fetal ileum contains an abundance of cells expressing HLA-D region (HLA-DR) antigens. In this study we characterized the HLA-DR positive cellular infiltrate in fetal ileum using a panel of monoclonal antibodies against cells of the macrophage/monocyte lineage. As well as anti-HLA-DR, the whole infiltrate was recognized by three of the antibodies in the panel studied: RFD1, reported to be an antibody to 'dendritic cells', leu3a which recognizes CD4 (an antigen expressed on helper/inducer T cells and macrophages), and PD7/26 which recognizes the leukocyte common antigen. The macrophage specific antibodies RFD7 and 3.9 stained fewer cells than the anti-HLA-DR. By sequential staining it was clear that most of the RFD7 positive macrophages in the fetal gut lamina propria were not recognized by 3.9 which is a broad specificity antimacrophage antibody in adult tissue. In contrast to this, more of the macrophages within the Peyer's patches of the fetal gut were recognized by 3.9 than by RFD7. In fetal lamina propria not all of the HLA-D region positive cells expressed macrophage markers and some expressed both markers associated with macrophages and 'dendritic' cells. Macrophages with different surface phenotypes were differentially distributed between the lamina propria and the primitive Peyer's patches; RFD7+, 3.9- macrophages were concentrated in the lamina propria whereas RFD7-, 3.9+ macrophages were abundant in Peyer's patches. Within the Peyer's patch lymphoid tissue RFD7+, 3.9- cells were present in the T cell zone whereas RFD7-, 3.9+ cells were concentrated in the dome region as they are in the adult. This suggests that functional heterogeneity of organized macrophage populations may occur in fetal ileum which is free of dietary and bacterial antigens.     

Functional analysis of TCR gamma delta+ T cells in tumour-infiltrating lymphocytes (TIL) of human pancreatic cancer.

The effect of HIV infection on intestinal lamina propria macrophage subsets was investigated in 41 patients at various stages of HIV infection (asymptomatic HIV infection, n = 17; AIDS, n = 24). Duodenal biopsies taken from HIV patients at endoscopy were snap frozen and cryostat sections cut for immunohistochemical staining. MoAbs CD68 (EBM11, pan-macrophage marker), RFD1 (antigen-presenting cells) and RFD7 (mature phagocytic macrophages) were used to identify cell subsets using indirect immunoperoxidase or alkaline phosphatase. Double immunofluorescence using MoAbs to HIV proteins (p24, p17 and gp120) and RFD1 were used to identify HIV-infected antigen-presenting cells. Double immunofluorescence was also used to identify macrophages that expressed both RFD1 and RFD7 ('suppressor' macrophages). Intensity of HLA-DR expression in lamina propria cells was investigated using a MoAb to HLA-DR directly conjugated to glucose oxidase. The results show that there was no difference in overall density of macrophages, but there was a significant decrease in dendritic cells (RFD1+) in all clinical stages of HIV. There was no difference in the density of RFD7+ macrophages, nor was there a difference intensity of HLA-DR expression in lamina propria cells. Only four HIV-infected cells were positively identified in the 41 patients. This result suggests that the antigen-presenting arm of mucosal immune defences may be seriously compromised in HIV infection, and represents a further insult to mucosal immunity already impaired as a result of loss of CD4+ T lymphocytes. This may contribute to development of opportunist infection in the gut.     

Antigen presenting cells in the nasal mucosa of patients with allergic rhinitis during allergen provocation

Background The role of antigen presenting cells (APC) in allergic rhinitis is underexposed. Allergen presentation to T lymphocytes is probably an important aspect of the pathophysiological mechanism of allergic rhinitis. Objectives The aim of the study was to investigate the presence and dynamics of APC with special emphasis on Langcrhans cells (LC) in the nasal mucosa of patients with an isolated grass pollen allergy during an out-of-season 2-week allergen exposure, mimicking the natural grass pollen season. Methods Seventeen patients with isolated grass pollen allergy and four control subjects were challenged daily with allergen during a 2-week period in the winter. Biopsy specimens were obtained once before, six times during and once after the provocation period. Biopsy sections were stained with monoclonal antibodies: OKT6 (CDla-Langerhans cells). Ki-M6 (CD68 macrophages), L25 (dendritic cells), anti-IgE, HLA-DR and HLA-DQ (Major Histoeompatibihty Complex Class II - antigen presenting eells), as well as staining with acid phosphatase. Results APC with different characteristics are present in the epithelium and lamina propria of the nasal mucosa. The number of LC increased significantly in epithelium and lamina propria. IgE⁺-LC were present in the nasal mucosa and increase during provocation, HLA-DR⁺ cells with dendritic and lymphocytic morphology and HLA-DQ⁺ cells were found. The number of these cells increased during provocation in epithelium and lamina propria. The number of HLA-DR⁺ epithelial cells did not change. A significant increase in the number of Ki-M6⁺ cells (macrophages) was found in the lamina propria. However, Ki-M6⁺ cells increased to the same extent in the lamina propria in the control group. Conclusion APC are influenced by allergen provocation. This study supports the hypothesis that (IgE⁺) LC are involved in allergic rhinitis. The role macrophages play remains doubtful.     

Mononuclear phagocyte system of human Peyer''s patches: an immunohistochemical study using monoclonal antibodies.

Macrophages in sections of human terminal ileal Peyer's patches were studied using a panel of monoclonal antibodies. Germinal centre macrophages were large and strongly positive for acid phosphatase and stained with antibodies RFD1, RFD9, UCHM1 and other macrophage-specific monoclonal antibodies, but not RFD7. In the macrophages of the dome region there was heterogeneity of size and staining for acid phosphatase. The majority of the RFD1 positive cells in this region appeared to be macrophages. However, no RFD9- or RFD7-positive cells were present. By contrast, RFD7-positive but RFD9-negative macrophages were present in the lamina propria. In the interfollicular areas there were cells with a dendritic morphology which were strongly HLA-DR, HLA-DQ and RFD1-positive, but which did not stain with the other macrophage specific monoclonal antibodies or for acid phosphatase. Some macrophages and lymphocytes in the germinal centre and dome regions expressed interleukin 2 and transferrin receptors. These cells were not present in the adjacent lamina propria.     

Lamina propria macrophages and dendritic cells differentially induce regulatory and interleukin 17-producing T cell responses

The intestinal immune system must elicit robust immunity against harmful pathogens but must also restrain immune responses directed against commensal microbes and dietary antigens. The mechanisms that maintain this dichotomy are poorly understood. Here we describe a population of CD11b+F4/80+CD11c- macrophages in the lamina propria that expressed several anti-inflammatory molecules, including interleukin 10 (IL-10), but little or no proinflammatory cytokines, even after stimulation with Toll-like receptor ligands. These macrophages induced, by a mechanism dependent on IL-10, retinoic acid and exogenous transforming growth factor-beta, the differentiation of Foxp3+ regulatory T cells. In contrast, lamina propria CD11b+ dendritic cells elicited IL-17 production. This IL-17 production was suppressed by lamina propria macrophages, indicating that a dynamic interaction between these subsets may influence the balance between immune activation and tolerance.     

Lamina Propria T Cell Subsets in the Small and Large Intestine of Euthymic and Athymic Mice

We investigated lamina propria T cells from the small intestine (jejunum/ileum) and the large intestine (colon) of euthymic (BALB/c, C. B-17, C57BL/6) and athymic (C57BL/6 nu/nu; BNX bg/bg nu/nu xid/ xid) mice. CD3⁺ T cells represented about 40% of the lamina propria lymphocytes (LPL) from the small or the large intestine of euthymic mice, and 20–30% of the LPL populations from the small or large intestine of athymic mice. In the lamina propria T cell population of the small intestine, 85% were of the αβ lineage in euthymic mice, but only 40% were of the αβ lineage in athymic mice. T cells of the αβ lineage were thus more frequent than T cells of the αβ lineage in the intestinal lamina propria T cells of extrathymic origin. CD4⁺ T cells represented 40% of the lamina propria T cells in the small as well as in the large intestine of euthymic mice, and 20–30% of the T cells in the lamina propria of the nude mouse gut. In euthymic mice, 40% of the T cells in the small intestine lamina propria, and 30% of the T cells in the colonic lamina propria were CD8⁺, In intestinal lamina propria T cell populations of athymic mice, the CD8⁺ T cell population was expanded. Most (60–70%) CD8+ T cells in the lamina propria of the small and the large intestine of euthymic and athymic mice expressed the homodimeric CD8α⁺β^? form of the CD8 coreceptor. A fraction of 15–20% of all CD3+ T cells in the lamina propria of the small and the large intestine of euthymic and athymic mice were ‘double negative’ CD4^? CD8^?. A large fraction of the TCRαβ⁺ T cells in the colonic lamina propria (but not in the small intestine lamina propria) of euthymic mice expressed the CD2 and the CD28 costimulator molecules, the adhesion molecule LECAM-1 (CD62 L), and could be activated in vitro by CD3 ligation. These data reveal a considerable heterogeneity in the surface phenotype and the functional phenotype of murine lamina propria T cells.     

IgG transport across mucosal barriers by neonatal Fc receptor for IgG and mucosal immunity

Mucosal secretions of the human gastrointestinal, respiratory, and genital tracts contain significant quantities of IgG. The neonatal Fc receptor for IgG (FcRn) plays a major role in regulating host IgG levels and transporting IgG and associated antigens across polarized epithelial barriers. The FcRn can then recycle the IgG/antigen complex back across the intestinal barrier into the lamina propria for processing by dendritic cells and presentation to CD4⁺ T cells in regional organized lymphoid structures. FcRn, through its ability to secrete and absorb IgG, thus integrates luminal antigen encounters with systemic immune compartments and, as such, provides essential host defense and immunoregulatory functions at the mucosal surfaces.     

The diverse ontogeny and function of murine small intestinal dendritic cell/macrophage subsets

Intestinal dendritic cell and macrophage subsets are believed to play key roles in maintaining intestinal homeostasis in the steady state and in driving protective immune responses in the setting of intestinal infection. This mini-review focuses on recent progress regarding the ontogeny and function of small intestinal lamina propria dendritic cell/macrophage subsets. In particular we discuss recent findings suggesting that small intestinal CD103⁺ dendritic cells and Cx3cr1⁺ cells derive from distinct precursor populations and that CD103⁺ dendritic cells represent the major migratory population of cells with a key role in initiating adaptive immune responses in the draining mesenteric lymph node. In contrast, Cx3cr1⁺ cells appear to represent a tissue resident population, phenotypically indistinguishable from tissue resident macrophages. These latter observations suggest an important division of labour between dendritic cell/macrophage subsets in the regulation of intestinal immune responses in the steady state.     

CD4+ T lymphocytes injected into severe combined immunodeficient (SCID) mice lead to an inflammatory and lethal bowel disease

Transfer of 2 × 10⁵ congenic or semiallogenic purified TCRαβ⁺ CD4⁺ T cells to SCID mice leads to an infiltration of the recipient gut lamina propria and epithelium with a donor-derived CD4⁺ T cell subset which induces a lethal inflammatory bowel disease (IBD) in the recipients. In contrast, IBD was not observed in SCID mice transplanted with unfractionated splenic cells. The earliest detectable pathological changes after CD4⁺ T cell transfer were proliferation and hypertrophy of the entire colonic epithelial layer, including increased mitotic activity, increased expression of epithelial nuclear proliferation antigen, and elongation of the crypts. Later on, massive mononuclear cell infiltration, hypertrophy of all layers of the colon and occasional epithelial ulcerations were observed. At this stage, accumulations of IgA, IgM and small numbers of IgG1-, IgG2-and IgG3-secreting plasma cells were present in the lamina propria of both the small and large intestine. We conclude that low numbers of intraveneously transferred CD4⁺ T cells induce IBD in SCID mice. In the late stages of CD4⁺ T cell-induced IBD, the colonic lamina propria becomes infiltrated with macrophages, neutrophils and plasma cells secreting IgA, IgM, and to a lesser degree IgG antibodies which might play an accessory role in the pathogenesis of IBD.     

Changes in the Intestinal Lymphoid Compartment Throughout Life: Implications for the Local Generation of Intestinal T Cells

The intestinal lymphoid compartment has a rather stable composition throughout life. However, both during early neonatal development and at high age unique cell populations can be recognized at distinct sites in the intestinal tissue. Directly after birth all intestinal CD3⁺ cells are found in the lamina propria. At this time the epithelium does not contain T cells. These CD3⁺ lamina propria lymphocytes co-express both TCRβ and TCRδ chains, probably reflecting the expression of a TCRβδ heterodimer on the cell surface. Cells with this particular phenotype are present in comparable numbers in the lamina propria of both neonatal euthymic and athymic mice, indicating the thymus-independent nature of these cells. During aging the frequency of TCRαβ⁺ CD8αα⁺ intestinal T cells increases. These cells are also considered to be thymus-independent. The appearance of high numbers of CD4⁺ CD8αα⁺ intestinal T cells in aged mice is even more striking. It is postulated that the neonatal TCRβδ⁺ cells, and probably also the CD4⁺ CD8αα⁺ cells as found in old mice, are intermediates in the extrathymic differentiation pathway of TCRαβ⁺ CD8αα⁺ intestinal T cells.     

Proteomic composition and immunomodulatory properties of urinary bladder matrix scaffolds in homeostasis and injury

Urinary bladder matrix (UBM) is used clinically for management of wounds and reinforcement of surgical soft tissue repair, among other applications. UBM consists of the lamina propria and basal lamina of the porcine urinary bladder, and is decellularized as part of the process to manufacture the medical device. UBM is composed mainly of Collagen I, but also contains a wide variety of fibrillar and basement membrane collagens, glycoproteins, proteoglycans and ECM-associated factors. Upon application of the biomaterial in a traumatic or non-traumatic setting in a mouse model, there is a cascade of immune cells that respond to the damaged tissue and biomaterial. Here, through the use of multicolor flow cytometry, we describe the various cells that infiltrate the UBM scaffold in a subcutaneous and volumetric muscle injury model. A wide variety of immune cells are found in the UBM scaffold immune microenvironment (SIM) including F4/80⁺ macrophages, CD11c⁺ dendritic cells, CD3⁺ T cells and CD19⁺ B cells. A systemic IL-4 upregulation and a local M2-macrophage response were observed in the proximity of the implanted UBM. The recruitment and activation of these cells is dependent upon signals from the scaffold and communication between the different cell types present.     

Visualizing Cytokine-Secreting Cells In Situ in the Rhesus Macaque Model of Chronic Gut Inflammation

Cytokine-producing cells in gut-associated lymphoid tissues of rhesus macaques with chronic enterocolitis were studied. The confocal microscopy technique that we developed enables simultaneous in situ visualization of multiple extra- and/or intracellular antigens at a resolution higher than that allowed by light or epifluorescence microscopy. The presence of interleukin-6 (IL-6)-, tumor necrosis factor alpha (TNF-α)-, and IL-1-α-producing cells was focally intense in the colon lamina propria of the affected animals. The IL-1-α-producing cells were T lymphocytes (CD3⁺), while the TNF-α-producing cells were both macrophages (CD68⁺/HAM56⁺/LN5⁺) and T lymphocytes (CD3⁺). The IL-6-producing cells within the colon consisted of T lymphocytes and macrophages. The amount of IL-6-producing cells seen in macaques with enterocolitis was significantly higher (P < 0.001) than that seen in the healthy control animal, while TNF-α- and IL-1-α-producing cells were seen only in macaques with enterocolitis. Most of the T lymphocytes that produced cytokines were detected in the lamina propria, while the macrophages were most prominent in highly inflamed regions of the lamina propria. Taken together, our findings indicate that there might be immunological similarity between chronic enterocolitis of rhesus macaques and humans, suggesting the potential use of the nonhuman primate model for the validation of novel therapies.     

Isolation and characterization of antigen-presenting dendritic cells from the mouse intestinal lamina propria

A method was developed for the isolation of antigen-presenting dendritic cells, and macrophages, from mouse intestinal lamina propria and Peyer's patches. Peyer's patches, and the lamina propria of both the small and large intestine, contained cells with potent stimulatory activity in the allogeneic mixed leucocyte reaction. These cells were separated from macrophages by fibronectin adherence and further enriched by density centrifugation. The isolated stimulatory cells expressed high levels of class II major histocompatibility complex (MHC) antigens, and resembled splenic dendritic cells in both morphology and function. Macrophages were recovered from the lamina propria but not Peyer's patches. These cells also expressed class II MHC antigens, but failed to stimulate the mixed leucocyte reaction and, instead, induced indomethacin-sensitive suppression.     

The identification and developmental requirements of colonic CD169+ macrophages

CD169‐positive macrophages in the marginal zone of the spleen and subcapsular sinus of lymph nodes play an important role as gatekeepers, strategically located to capture pathogens. Here we identified a population of CD169‐positive macrophages in the colon and investigated which factors influenced their development. Murine colonic CD115⁺ F4/80^lo CD11c^lo macrophages expressing CD169 were present in the lamina propria, mainly surrounding the crypts. In spite of the high levels of bacterial flora in the colon and the importance of Toll‐like receptor signalling in mucosal homeostasis, the presence of CD169⁺ macrophages was not affected in mice that were deficient in MyD88‐mediated Toll‐like receptor signalling and in mice in which the bacterial flora was eradicated. Whereas the development of splenic CD169⁺ macrophages was dependent on lymphotoxin α, colonic CD169⁺ macrophages were present in normal numbers in lymphotoxin α‐deficient mice. In contrast, reduced numbers of CD169⁺ macrophages were found in the colon of mice deficient in vitamin A, whereas CD169⁺ macrophages in the spleen were unaffected. In conclusion, we identified a new macrophage subset in the lamina propria of the colon characterized by the expression of CD169. Its differentiation, unlike CD169⁺ macrophages in lymphoid organs, is independent of lymphotoxin α signalling, but requires vitamin A.     

Distribution of beta 7 integrins in human intestinal mucosa and organized gut-associated lymphoid tissue.

Two alternative integrins involved in mucosal homing (alpha 4 beta 7) or epithelial retention (alpha E beta 7) of lymphocytes were examined in the human gut. The distribution of the beta 7 subunit [monoclonal antibody (mAb) M301] was bimodal in that it was strongly expressed by alpha E beta 7 + cells but weakly by alpha 4 beta 7 + cells. More than 90% of intraepithelial lymphocytes (IEL), including the minor subsets of CD4+, T-cell receptor (TCR) gamma/delta +, and CD3- cells, expressed alpha E beta 7 as did most lamina propria CD8+ (88%) and a fraction (36%) of CD4+ lymphocytes. Conversely, B-lineage cells (CD19+) and macrophages (CD68+) were negative. In gut-associated lymphoid tissue (GALT: Peyer's patches and appendix) only a few (< 5%) cells were positive for alpha E beta 7 (confined to CD8+ lymphocytes and CD11c+ putative dendritic cells). A relatively small fraction of IEL (30-50%) expressed alpha 4 beta 7 (mAb Act-1), while most (70%) lamina propria T and B lymphocytes, blasts, plasma cells and macrophages were positive. In GALT, T lymphocytes expressed similar levels of alpha 4 beta 7 as in the lamina propria whereas relatively few B lymphocytes (< 50%) were positive. Isolated lamina propria CD8+, CD4+, CD19+, and CD38+ cells contained mRNA for alpha 4 and the former three subsets as well as appendix CD8+ cells also for beta 7 while only lamina propria CD8+ cells had mRNA for alpha E. Together, the results suggested that alpha E beta 7 and alpha 4 beta 7 are differentially regulated in inductive sites and effector sites of the human gut. Because lymphoid cells at both sites expressed mainly alpha 4 beta 7, this integrin may be a homing receptor on memory and effector cells bound for lamina propria as well as on naive lymphocytes extravasating in GALT. Conversely, because alpha E beta 7 was mainly expressed by CD8+ cells in epithelium and lamina propria, it was probably induced after extravasation, in agreement with the observation that IEL and a fraction of lamina propria T lymphocytes (mainly CD8+ cells) generally expressed higher levels of beta 7 than most CD4+ and B cells. Also a subset of putative dendritic cells located near the follicle-associated epithelium of GALT expressed alpha E beta 7, perhaps reflecting epithelial interaction during primary immune responses.     

Cytotoxic reactivity of gut lamina propria CD4+ αβ T cells in SCID mice with colitis

Polyclonal, mucosa-seeking memory/effector CD4⁺ T cells containing a large fraction of blasts activated in situ accumulate in the gut lamina propria of severe-combined immunodeficient (SCID) mice developing colitis after CD4⁺ T cell transplantation. CD4⁺ T cells isolated from different repopulated lymphoid tissues of transplanted SCID mice proliferate in vitro in the presence of interleukin (IL)-2 + IL-7. CD3 ligation enhances this cytokine-supported proliferation in CD4⁺ T cells from the spleen and the mesenteric lymph node of transplanted SCID mice; CD3 ligation suppresses the cytokine-supported proliferation in CD4⁺ T cells from the gut lamina propria in a cell density- and dose-dependent manner. Almost all CD4⁺ T cells from repopulated lymphoid tissues of transplanted SCID mice express CD95 (Fas) on the cell surface, and a large fraction of CD4⁺ T cells from the gut lamina propria of transplanted SCID mice express the Fas ligand on the surface. Gut lamina propria CD4⁺ T cells show Fas-dependent cytotoxicity. A large fraction of gut lamina propria CD4⁺ T cells that infiltrate the inflamed colon in transplanted SCID mice are activated in situ and many CD4⁺ T cells are apoptotic. Hence, a large fraction of colitis-inducing CD4⁺ T cells undergo activation-induced cell death in situ and can damage other cells through Fas-dependent cytotoxicity.     

Functional maturation of lamina propria dendritic cells by activation of NKT cells mediates the abrogation of oral tolerance

We previously showed that although systemic administration of α‐galactosylceramide (αGalCer) or agonistic anti‐CD40 induced functional maturation of dendritic cells (DC) in mesenteric lymph nodes, only the former treatment succeeded in breaking the induction of oral tolerance. In this study, we looked for the essential factor responsible for the disruption of oral tolerance. We found that lamina propria (LP)‐DC was responsible for the oral OVA presentation and that Peyer's patch was not essential for the induction of oral tolerance. Therefore, we investigated the role of LP‐DC. Treatment with αGalCer but not with anti‐CD40 induced the full maturation of LP‐DC at an early time point. This functional activation of LP‐DC was mediated by strong activation of NKT cells that reside abundantly in the small intestinal lamina propria (SI‐LP) and interferon‐γ partially contributed to the LP‐DC activation. LP‐DC isolated from αGalCer‐treated OVA‐fed mice induced the differentiation of naïve CD4⁺ T cells into Th1 and Th2 and was associated with the reduced Foxp3⁺ population. In contrast, LP‐DC isolated from anti‐CD40‐treated OVA‐fed mice failed to generate Th cell differentiation but induced more Foxp3⁺ CD4⁺ T cells. Our results demonstrate that triggered by NKT cells in SI‐LP, functional maturation of Ag‐capturing DC from SI‐LP is necessary for the abrogation of oral tolerance induction.     

Macrophage accumulation in gut mucosa differentiates AIDS from chronic SIV infection in rhesus macaques

The relationship between recruitment of mononuclear phagocytes to lymphoid and gut tissues and disease in HIV and SIV infection remains unclear. To address this question, we conducted cross‐sectional analyses of dendritic cell (DC) subsets and CD163⁺ macrophages in lymph nodes (LNs) and ileum of rhesus macaques with acute and chronic SIV infection and AIDS. In LNs significant differences were only evident when comparing uninfected and AIDS groups, with loss of myeloid DCs and CD103⁺ DCs from peripheral and mesenteric LNs, respectively, and accumulation of plasmacytoid DCs and macrophages in mesenteric LNs. In contrast, there were fourfold more macrophages in ileum lamina propria in macaques with AIDS compared with chronic infection, and this increased to 40‐fold in Peyer's patches. Gut macrophages exceeded plasmacytoid DCs and CD103⁺ DCs by ten‐ to 17‐fold in monkeys with AIDS but were at similar low frequencies as DCs in chronic infection. Gut macrophages in macaques with AIDS expressed IFN‐α and TNF‐α consistent with cell activation. CD163⁺ macrophages also accumulated in gut mucosa in acute infection but lacked expression of IFN‐α and TNF‐α. These data reveal a relationship between inflammatory macrophage accumulation in gut mucosa and disease and suggest a role for macrophages in AIDS pathogenesis.     

Mechanisms of Oral Tolerance

Oral tolerance is a state of systemic unresponsiveness that is the default response to food antigens in the gastrointestinal tract, although immune tolerance can also be induced by other routes, such as the skin or inhalation. Antigen can be acquired directly by intestinal phagocytes, or pass through enterocytes or goblet cell-associated passages prior to capture by dendritic cells (DCs) in the lamina propria. Mucin from goblet cells acts on DCs to render them more tolerogenic. A subset of regulatory DCs expressing CD103 is responsible for delivery of antigen to the draining lymph node and induction of Tregs. These DCs also imprint gastrointestinal homing capacity, allowing the recently primed Tregs to home back to the lamina propria where they interact with macrophages that produce IL-10 and expand. Tregs induced by dietary antigen include Foxp3⁺ Tregs and Foxp3^? Tregs. In addition to Tregs, T cell anergy can also contribute to oral tolerance. The microbiota plays a key role in the development of oral tolerance, through regulation of macrophages and innate lymphoid cells that contribute to the regulatory phenotype of gastrointestinal dendritic cells. Absence of microbiota is associated with a susceptibility to food allergy, while presence of Clostridia strains can suppress development of food allergy through enhancement of Tregs and intestinal barrier function. It is not clear if feeding of antigens can also induce true immune tolerance after a memory immune response has been generated, but mechanistic studies of oral immunotherapy trials demonstrate shared pathways in oral tolerance and oral immunotherapy, with a role for Tregs and anergy. An important role for IgA and IgG antibodies in development of immune tolerance is also supported by studies of oral tolerance in humans. The elucidation of key pathways in oral tolerance could identify new strategies to increase efficacy of immunotherapy treatments for food allergy.