Oral tolerance

Abstract

Multiple mechanisms of tolerance are induced by oral antigen. Low doses favor active suppression, whereas higher doses favor clonal anergy/deletion. Oral antigen induces T-helper 2 [interleukin (IL)-4/IL-10] and Th3 [transforming growth factor (TGF)-beta] T cells plus CD4+CD25+ regulatory cells and latency-associated peptide+ T cells. Induction of oral tolerance is enhanced by IL-4, IL-10, anti-IL-12, TGF-beta, cholera toxin B subunit, Flt-3 ligand, and anti-CD40 ligand. Oral (and nasal) antigen administration suppresses animal models of autoimmune diseases including experimental autoimmune encephalitis, uveitis, thyroiditis, myasthenia, arthritis, and diabetes in the non-obese diabetic (NOD) mouse, plus non-autoimmune diseases such as asthma, atherosclerosis, graft rejection, allergy, colitis, stroke, and models of Alzheimer's disease. Oral tolerance has been tested in human autoimmune diseases including multiple sclerosis (MS), arthritis, uveitis, and diabetes and in allergy, contact sensitivity to dinitrochlorobenzene (DNCB), and nickel allergy. Although positive results have been observed in phase II trials, no effect was observed in phase III trials of CII in rheumatoid arthritis or oral myelin and glatiramer acetate (GA) in MS. Large placebo effects were observed, and new trials of oral GA are underway. Oral insulin has recently been shown to delay onset of diabetes in at-risk populations, and confirmatory trials of oral insulin are being planned. Mucosal tolerance is an attractive approach for treatment of autoimmune and inflammatory diseases because of lack of toxicity, ease of administration over time, and antigen-specific mechanisms of action. The successful application of oral tolerance for the treatment of human diseases will depend on dose, developing immune markers to assess immunologic effects, route (nasal versus oral), formulation, mucosal adjuvants, combination therapy, and early therapy.

Figures

Fig. 1. Mechanisms of oral tolerance induction

Oral antigen crosses from the intestine into the gut associated lymphoid tissue in a number of ways. It can enter via M cells, be sampled by DC processes that penetrate the lumen, or be taken up by intestinal epithelial cells. DCs in the gut are unique in that they can drive Treg differentiation from Foxp3− cells. These properties of DCs relate to their being conditioned by commensal bacteria, TGFβ and IL-10 from gut epithelial cells, and their expression of retinoic acid, which is provided in the form of vitamin A in the diet and appears to be constitutively expressed by gut DCs. CD11b monocytes may also play a role in the induction of Tregs, and the induction of Tregs occurs in the MLNs and involves both CCR7 and CCR9. Costimulation by PDL1-PDL are also important for the induction of Tregs. Macrophages are stimulated to produce TGF-β after uptaking apoptotic epithelial cells or apoptotic T cells following high dose tolerance. Lower doses of antigen favor the induction of Tregs, whereas higher doses of antigen favor anergy/deletion as a mechanism of tolerance induction. The liver may also play a role in oral tolerance induction and antigen (high dose) may be rapidly taken up by the liver, where it is processed by plasmacytoid DCs that induce anergy and deletion. A number of different types of Tregs may be induced or expanded in the gut including CD4+CD25+Foxp3+ iTregs, nTregs, Tr1 cells, LAP+ Tregs (Th3 cells), CD8+ Tregs, and γδ T cells.

Fig. 2. Regulatory T-cell cascade following induction of Th3 type Tregs by oral antigen or oral anti-CD3

A Th3 cell is a CD4+CD25−Foxp3−LAP+ cell that exists in the peripheral immune compartment and is triggered by TCR signaling in the gut by oral antigen. Following triggering in the gut, the Th3 cell secretes TGF-β. Secreted TGF-β acts to maintain naturally occurring CD4+CD25+Foxp3+ Tregs, suppress Th1 and Th2 responses, and in concert with IL-6 may induce Th17 responses. Secreted TGF-β from Th3 cells also acts on CD4+Foxp3− cells and converts them to iTregs, which are Foxp3+CD25+LAP−. Depending on the milieu, these cells may become Foxp3+CD4+CD25+LAP+. These induced Tregs may also condition DCs to secrete IL-27 and in turn induce IL-10-secreting Tr1 cells.

Fig. 3. Intestinal closed loop experiments demonstrating binding of anti-CD3 to DCs in the gut

Mice with a targeted deletion of chemokine receptor CX3CR1 with eGFP insertion (HC. Reinecker, MGH) have all monocytes and DCs in the gut labeled with eGFP (green). Mice were anesthetized, stomach opened, and a closed loop was created in a part of the small intestine. Alexa Fluor 700 labeled anti-CD3 (2c11) antibody (blue) was injected into the loop and the intestinal loop harvested 45 min later. The intestinal content was cleared with PBS before imaging with confocal microscopy using multi-tracking for 2-color imaging. Image acquisition was carried out with Volocity software. DCs are labeled green and anti-CD3 antibody labeled blue. Green = DCs; Blue = anti-CD3 monoclonal antibody.

Fig. 4. Immune regulatory pathways in the gut induced by AHR ligands

AHR ligands such as ITE in the gut either from commensal bacteria or administered orally act directly on T cells and DCs in the gut to induce Foxp3iTregs and Foxp3-Tr1 cells. Both Tregs and AHR ligands condition DCs to amplify Treg induction through the production of IL-27 and retinoic acid.

Fig. 5. Oral insulin effect most evident in subjects with baseline IAA ≥ 300

A subgroup of islet autoantibody relatives with the highest level of insulin autoantibodies showed delayed progression to diabetes when treated with oral insulin to induce mucosal tolerance. Survival curve depicting time to diagnosis of type 1 diabetes in the DPT-1 oral insulin trial, for the subset of subjects with baseline-confirmed insulin autoantibody level of 300 nU/mL or above. Skyler, JSS Ann NY Acad Sci 2008. 1150: p 194.