Maintenance Of Oral Tolerance Research Articles

Goblet cell associated antigen passages support the induction and maintenance of oral tolerance.

Tolerance to innocuous antigens from the diet and the commensal microbiota is a fundamental process essential to health. Why tolerance is efficiently induced to substances arising from the hostile environment of the gut lumen is incompletely understood but may be related to how these antigens are encountered by the immune system. We observed that goblet cell associated antigen passages (GAPs), but not other pathways of luminal antigen capture, correlated with the acquisition of luminal substances by lamina propria (LP) antigen presenting cells (APCs) and with the sites of tolerance induction to luminal antigens. Strikingly this role extended beyond antigen delivery. The GAP function of goblet cells facilitated maintenance of pre-existing LP T regulatory cells (Tregs), imprinting LP-dendritic cells with tolerogenic properties, and facilitating LP macrophages to produce the immunomodulatory cytokine IL-10. Moreover, tolerance to dietary antigen was impaired in the absence of GAPs. Thus, by delivering luminal antigens, maintaining pre-existing LP Tregs, and imprinting tolerogenic properties on LP-APCs GAPs support tolerance to substances encountered in the hostile environment of the gut lumen.

Novel role of intestinal Goblet cells and Goblet Cell Associated Antigen Passages in induction of tolerance to dietary antigens and promoting intestinal homeostasis

Abstract Mucosal tolerance towards luminal antigens is a process fundamental to intestinal homeostasis, the breakdown of which results in onset of inflammatory bowel disease. How luminal antigens cross the intestinal epithelium to interact with the immune system in a manner capable of induction and maintenance of oral tolerance is unknown. We have identified a new pathway of luminal antigen delivery to dendritic cells (DC) in the lamina propria (LP), which was mediated by goblet cells (GC) associated antigen passages (GAPs). The role of GCs and GAPs in promoting mucosal tolerance is unexplored. We studied tolerogenic responses to dietary antigen, ovalbumin (Ova) in mice where GCs were deleted, or when GAPs are inhibited but GCs remain and compared to corresponding control mice. In the absence of GCs or functional GAPs, LP-DCs could not acquire luminal antigens in a manner capable of inducing antigen specific T cell responses in the draining mesenteric lymph nodes (MLN). Moreover, deletion of GCs/GAPs resulted in loss of induction of regulatory T cells in the MLN and abrogated tolerance to dietary antigens. In addition, loss of GCs/GAPs resulted in rapid reduction of pre-existing induced Treg population in the SI-LP. Notably, the in absence of GCs/GAPs resulted in rapid expansion of IL-17 producing T cells in the small intestine, indicating that GCs and GAPs play a crucial role in balancing the Treg/Th-17 axis in the intestine. Hence, suggesting that GCs and GAPs play a crucial role in the induction and maintenance of oral tolerance. These findings identify a previously unappreciated role of GCs in maintaining the Treg/Th17 balance and that processes resulting in inadequate formation of GAPs may underlie the loss of tolerance to luminal antigens.

Role of the Microbiome in Food Allergy.

Resident microbial communities likely modify risk for allergic disorders, including food allergy. We review epidemiologic studies linking microbial exposures to food allergy risk and discuss the mechanisms by which the microbiome may modulate oral tolerance. We additionally address ongoing translational efforts in human microbiome studies. Epidemiologic studies and murine models support that altered microbial exposures and colonization in early life modify food allergy risk. Differential microbiota confer protection or susceptibility to food allergy by modulating the regulatory tone of the mucosal immune system. Recent efforts are focused on the identification of bacterial strains necessary for oral tolerance in human and microbial-based clinical trials. Early childhood appears to be critical for the colonization of a diverse microbiota necessary for the induction and maintenance of oral tolerance. Identification and functional evaluation of protective commensal microbes will inform strategies for the prevention and treatment of food allergy.

The Dietary Paradox in Food Allergy: Yesterday's Mistakes, Today's Evidence and Lessons for Tomorrow

During the last decades the prevalence of food allergy has significantly increased among children and antigen avoidance still remains the standard care for the management of this condition. Most reactions are IgE-mediated with a high risk of anaphylaxis requiring emergency medications in case of inadvertent ingestion. Recent studies showed that continuous administration of the offending food, rather than an elimination diet, could promote the development and maintenance of oral tolerance. Indeed, intestinal transit of food proteins and their interaction with gut-associated lymphoid tissue (GALT) is the essential prerequisite for oral tolerance. On the contrary, low-dose cutaneous exposure to environmental foods in children with atopic dermatitis and altered skin barrier facilitates allergic sensitization. The timing and the amount of cutaneous and oral exposure determine whether a child will have allergy or tolerance. Furthermore, previous preventive strategies such as the elimination diet during pregnancy and breastfeeding, prolonged exclusive breastfeeding and delayed weaning to solid foods did not succeed in preventing the development of food allergy. On the other hand, there could be an early narrow window of immunological opportunity to expose children to allergenic foods and induce natural tolerance. Finally, the gradual exposure to the offending food through special protocols of specific oral tolerance induction (SOTI) may be a promising approach to a proactive treatment of food allergy.

Interferon-λ Mediates Oral Tolerance and Inhibits Antigen-Specific, T-Helper 2 Cell–Mediated Inflammation in Mouse Intestine

Oral tolerance is an important component of gastrointestinal homeostasis, but mechanisms of its development are not fully understood. Loss of oral tolerance occurs during food allergen-related inflammation in the gastrointestinal tract. Interferon (IFN)-λ regulates immunity, but its role in oral tolerance is not clear. We investigated the role and the mechanism of IFN-λ in the development of oral tolerance and its effect on antigen-induced, T-helper (Th)-2 cell-mediated inflammation in the intestine. Expression of IFN-λ and its receptor were analyzed by immunohistochemical, flow cytometric, or immunoblot analyses. Tolerogenic dendritic cells (DCs) and regulatory T cells were examined in vitro and in vivo. A mouse model of antigen-induced, Th2 cell-mediated intestinal inflammation was used to examine the role of IFN-λ and T cells in oral tolerance in the intestine. CD3+ cells expressed the IFN-λ receptor, which was up-regulated following antigen-specific or nonspecific activation. Interaction between IFN-λ and its receptor induced apoptosis of T cells and their subsequent phagocytosis by DCs. This led to the generation of tolerogenic DCs and T regulatory cells in vitro and in vivo. Passive transfer of IFN-λ-primed CD3+ cells inhibited Th2 cell-mediated inflammation in the intestine. IFN-λ is involved in development and maintenance of oral tolerance in the intestines of mice; it might be used to suppress antigen-specific Th2 cell-mediated inflammation in patients.

Intestinal Immune Regulation as a Potential Diet-Modifiable Feature of Gut Inflammation and Autoimmunity

The gastrointestinal tract represents the largest immune interface with the environment. Exposure to large numbers of dietary and microbial antigens requires complex and highly regulated intestinal immune responses by different immune cell types for the maintenance of oral tolerance. Defective immune homeostasis can cause gut barrier dysfunction and breakdown of tolerance, leading to chronic inflammation and autoimmunity. In this review, we summarize the key immune cell populations involved in oral tolerance. We also describe diet-modifiable aspects of gut immunity that alter the intricate balance between inflammatory and tolerogenic immune responses in the gut and contribute to disease development.

Steady-state migrating intestinal dendritic cells induce potent inflammatory responses in naive CD4+T cells

Steady-state dendritic cells (DCs) migrating in the lymph from the intestine induce tolerance to harmless intestinal antigens, preventing inflammatory responses. To determine if such DCs are inherently tolerogenic we collected intestinal lymph DCs (L-DCs) by cannulation of the thoracic duct of rats after mesenteric lymphadenectomy, and examined their capacity to activate naive CD4+ lymphocytes in an allogeneic mixed leucocyte reaction. L-DCs stimulated strong proliferative responses, induced secretion of inflammatory cytokines including interferon-γ, and induced FoxP3-positive lymphocytes to divide. To determine if the activated CD4+ T cells had been tolerized, they were rested and restimulated with irradiated splenocytes. The restimulated CD4+ T cells again proliferated and secreted inflammatory cytokines. These data demonstrate that the DCs, which migrate from the intestine in the steady state, are paradoxically able to induce strong inflammatory responses from naive T cells, despite their role in the maintenance of oral tolerance.

Health, Probiotics, and Inflammation

Probiotic bacteria are normal inhabitants of microflora and may confer several benefits, including prevention against intestinal inflammation. However, the exact mode of action of probiotics is still largely unknown. The first line of defense against the entry of pathogens is represented by the gut membrane barrier and probiotics may prevent pathogen-induced membrane damage by inhibiting pathogen adhesion and maintaining the correct organization of the tight junction and cytoskeleton proteins. The gut immune system should not only protect the mucosa against pathogens, but also avoid hypersensitivity reactions to food proteins and normal microflora. Failure of induction or maintenance of oral tolerance has been postulated to be a cause of food allergy. Feeding probiotic bacteria may prevent or ameliorate the onset of allergic disease and the associated inflammatory reactions through mechanisms involving modulation of T regulatory cells. Breakdown in tolerance toward intestinal bacteria is a primary cause of inflammatory bowel disease. Recent studies have shown that probiotics may ameliorate experimental colitis in mice by inducing interleukin 10 and interleukin 10-dependent T regulatory cells. In this article, an update of the anti-inflammatory activity of different probiotics and of the more accredited mechanisms underlying such activities are reported.

Oral Administration of IL-12 Abrogates the Induction but Not the Maintenance of Oral Tolerance

Background: IL-12 is a Th1-inducing cytokine and known to be induced by some food factors. This function of such foods is expected to be applied for antiallergic materials. However, the influence of IL-12 on the immune responses has not been fully investigated. Oral tolerance is an immunologically unresponsive state induced by orally administered food antigens. Although a failure in the induction of oral tolerance would result in food allergy, the mechanisms for the induction and abrogation of oral tolerance have not been clarified. IL-12 induced by food factors may also affect the induction of oral tolerance. In this study we examined the effect of the oral administration of IL-12 on the induction and maintenance of oral tolerance. Methods: BALB/c mice were fed β-lactoglobulin with or without IL-12. After immunizing them with the antigen, the serum antibody titer of these mice was measured to evaluate the induction of oral tolerance. Results: The induction of oral tolerance was prevented in the mice that had been simultaneously fed the antigen and IL-12. On the other hand IL-12 did not abrogate the already established oral tolerance when it was administered after feeding the antigen. We also found that oral tolerance could be induced normally in mice that had failed to induce such tolerance by simultaneous feeding of IL-12 and the antigen. Conclusions: The results indicate that IL-12 induced in the intestine by some food factors is involved in the regulation of oral tolerance.

Literature review: The best new articles in the specialty of allergy, asthma, and immunology, 2004–2005

A series of eight articles from the published literature for the period 2004-2005 was selected for review in this article. Small-airway obstruction in chronic obstructive pulmonary disease (COPD) and its management were the subjects of the first two articles reviewed in this commentary. Progression of COPD from the Global Initiative for Chronic Obstructive Lung Disease stages 0-4 was found to be most strongly associated with thickening of the airway wall and each of its compartments by a repair or remodeling process. Management of COPD, including the addition of pulmonary rehabilitation to treatment regimens, may reduce symptoms and improve exercise performance. The addition of theophylline or an inhaled corticosteroid (or both) to optimal inhaled bronchodilator therapy together with aggressive treatment of hypoxemia were suggested to provide additional benefits and lung-volume-reduction surgery and transplantation were recommended as other treatment options for a subgroup of patients with very severe disease. The next three articles addressed the impact of outdoor and indoor air pollution and the effects of childhood asthma as antecedents of asthma in the adult in later life. Current levels of air pollution are known to have chronic, adverse effects on lung development in children from age 10 to 18 years, leading to clinically significant deficits in attained forced expiratory volume in one second as children reach adulthood. Among inner-city children with atopic asthma, an individualized, home-based, comprehensive environmental intervention resulted in reduced asthma-associated morbidity and supported that a multifaceted, home-based, environmental intervention approach may be beneficial. The prognosis of childhood allergic asthma in adulthood was found to be determined largely early in life and the degree of atopy appeared to be a critical determinant of asthma persistence. The next article reviewed the controversial area of the treatment of chronic rhinosinusitis caused by fungi. Intranasal Amphotericin B was shown to reduce inflammatory mucosal thickening on both CT scan and nasal endoscopy and decreased the levels of intranasal markers for eosinophilic inflammation in patients with chronic rhinosinusitis. The last two articles reviewed two papers dealing with the role that gastrointestinal immune responses play in maintaining protective immunity in health and a critical role in the pathogenesis of a wide variety of clinical disorders associated with food allergy as well as the suspected pivotal role of oral tolerance to dietary proteins for the prevention of food allergy. The mode of antigen uptake in the gut and different regulatory immune cells appear to play critical roles in maintenance of oral tolerance as shown in many animal model systems.

Potentiation of immunological tolerance induction in adult mice by co-administration of pooled normal IgG and oral tolerogens: a potential therapeutic approach for autoimmune diseases

Oral tolerance can be defined as the inability of an adult animal to produce specific antibodies or cellular immune responses upon conventional immunization, after oral antigenic administration. Recently, the oral administration of antigens has gained renewed interest because of the possibility of inducing tolerance in nonimmunized adult animals and, consequently, opening up the theoretical possibility of preventing or treating diseases caused by malfunction of the immune system. This strategy has been proven to be useful in the prevention of allergic and autoimmune diseases in rodents, as well as in the amelioration of certain autoimmune diseases in humans. Although there is experimental and clinical evidence for the usefulness of oral tolerance in medical practice, the mechanisms responsible for this phenomenon are still poorly understood, and the results obtained are not always satisfactory. Herein, we show that the thymus is required for the induction and maintenance of oral tolerance, providing evidence that it is not a pure form of clonal deletion-based peripheral tolerance. Oral tolerance could therefore depend on the formation and release to the periphery of regulatory T cells, such as gammadelta or alphabeta T cells, by the thymus. This finding may have profound implications for the treatment of autoimmune diseases, since most of them are associated with thymic hypofunction. On the other hand, due to so far unknown mechanisms, the intraperitoneal co-administration of normal IgG to mice orally treated with tolerogen leads to a sustained and intense immunological tolerance, both in euthymic and thymectomized mice, including those of the lupus erythematosus-prone NZB x NZW lineage. This approach for inducing and maintaining tolerance in thymus-deficient conditions is discussed and put forth herein as a new evidence-based proposition for the therapy of autoimmune diseases.

Commonly Used Drugs Impair Oral Tolerance in Mice

Ibuprofen and antibiotics are commonly prescribed during early childhood. When given to mice at the time at which oral tolerance is induced, both treatments affect either the induction or the maintenance of oral tolerance. These results suggest that the coadministration of these and similarly acting drugs should be considered cautiously for infants at risk of allergy.

Cow's Milk Allergy in Infants with Atopic Eczema is Associated with Aberrant Production of Interleukin-4 During Oral Cow's Milk Challenge

A failure in the establishment and maintenance of oral tolerance in infancy may result in food allergy. To further assess the role of the intestinal immune system in cow's milk allergy (CMA), we investigated the systemic production of the pro-allergenic Th2 cytokine interleukin (IL)-4 and anti-allergenic cytokines IL-10, transforming growth factor (TGF)-beta1 and TGF-beta2 in infants suffering from atopic eczema with and without CMA during antigen elimination diet and oral antigen exposure. 18 infants (mean age, 9.6 months; 95% confidence interval 8.1-11.1 months) with atopic eczema and CMA and 17 infants (mean age, 9.7 months; 95% confidence interval 8.6-10.9 months) with atopic eczema tolerant to milk as assessed by a double blind, placebo-controlled cow's milk challenge were investigated. Peripheral blood mononuclear cells were obtained during antigen elimination diet and during oral cow's milk challenge and stimulated with Concanavalin-A or cow's milk or were left unstimulated. The cytokine concentrations were measured by enzyme-linked immunosorbent assay. During antigen elimination, the Concanavalin A-stimulated production of TGF-beta2 was significantly lower in infants with CMA as compared with infants without CMA: 129 pg/mL (interquartile ratio, 124-144 pg/mL) vs. 149 pg/mL (interquartile ratio, 133-169 pg/mL); P = 0.016. During oral antigen exposure, the immune responses in infants with CMA were characterized by significantly higher spontaneous production of IL-4 as compared with those without CMA: 12.0 pg/mL (interquartile ratio, 5.2-28.3 pg/mL) vs. 4.2 pg/mL (interquartile ratio, 1.5-7.6 pg/mL); P = 0.018. Infants with atopic eczema and CMA exhibit markedly increased systemic pro-allergenic IL-4 responses on intestinal antigen contact, which may partially be explained by a defective ability to launch anti-allergenic TGF-beta2 responses.

Cow's Milk Allergy in Infants with Atopic Eczema is Associated with Aberrant Production of Interleukin‐4 During Oral Cow's Milk Challenge

ABSTRACTObjectives:A failure in the establishment and maintenance of oral tolerance in infancy may result in food allergy. To further assess the role of the intestinal immune system in cow's milk allergy (CMA), we investigated the systemic production of the pro‐allergenic Th2 cytokine interleukin (IL)‐4 and antiallergenic cytokines IL‐10, transforming growth factor (TGF)‐β1 and TGF‐β2 in infants suffering from atopic eczema with and without CMA during antigen elimination diet and oral antigen exposure.Methods:18 infants (mean age, 9.6 months; 95% confidence interval 8.1‐11.1 months) with atopic eczema and CMA and 17 infants (mean age, 9.7 months; 95% confidence interval 8.6‐10.9 months) with atopic eczema tolerant to milk as assessed by a double blind, placebo‐controlled cow's milk challenge were investigated. Peripheral blood mononuclear cells were obtained during antigen elimination diet and during oral cow's milk challenge and stimulated with Concanavalin‐A or cow's milk or were left unstimulated. The cytokine concentrations were measured by enzyme‐linked immunosorbent assay.Results:During antigen elimination, the Concanavalin A‐stimulated production of TGF‐β2 was significantly lower in infants with CMA as compared with infants without CMA: 129 pg/mL (interquartile ratio, 124‐144 pg/mL) vs. 149 pg/mL (interquartile ratio, 133‐169 pg/mL); P = 0.016. During oral antigen exposure, the immune responses in infants with CMA were characterized by significantly higher spontaneous production of IL‐4 as compared with those without CMA: 12.0 pg/mL (interquartile ratio, 5.2‐28.3 pg/mL) vs. 4.2 pg/mL (interquartile ratio, 1.5‐7.6 pg/mL); P = 0.018.Conclusions:Infants with atopic eczema and CMA exhibit markedly increased systemic pro‐allergenic IL‐4 responses on intestinal antigen contact, which may partially be explained by a defective ability to launch anti‐allergenic TGF‐β2 responses.

Identification of the genes specifically expressed in orally tolerized T cells.

Oral tolerance is the systemic immunological unresponsiveness that occurs after feeding protein antigens. Its physiological role is thought to be the prevention of hypersensitivity to food antigens, and its therapeutic use to treat inflammatory diseases has been suggested. Although it has been shown that CD4(+) T cells mediate oral tolerance, the precise molecular mechanisms remain unclear. In the present study, we employed suppression subtractive hybridization and identified 10 genes specifically expressed in orally tolerized T cells. These included genes that were interesting in terms of their putative functions in the negative regulation of T cell activation, e.g. Culin 1, LAX, and Zfhx1b, as well as four genes that encoded unknown proteins. We further investigated the expression of these genes in hyporesponsive T cells induced in vitro (in vitro anergized T cells). We found that six of the 10 genes were highly expressed in these cells, and kinetic studies suggested that one was associated with the induction of anergy, while the other five were associated with the maintenance of anergy. The remaining 4 genes that were not expressed in in vitro anergized T cells are also of interest as they may play a specific role in in vivo T cell tolerance. Functional analysis of these genes should help to understand the complex mechanisms underlying the induction and maintenance of oral tolerance, and moreover, in vivo immune tolerance in general.

CC chemokine ligand 2 and its receptor regulate mucosal production of IL-12 and TGF-beta in high dose oral tolerance.

Oral tolerance is the result of a complex immunoregulatory strategy used by the gut and its associated lymphoid tissues to render the peripheral immune system unresponsive to nonpathogenic proteins, such as food or commensal bacteria. The mechanism of oral tolerance induction and maintenance is not well understood. We have previously shown that the chemokine, CC chemokine ligand 2 (CCL2), is important for the induction and maintenance of oral tolerance. To address the role CCL2 plays in oral tolerance, we used both CCL2(-/-) and CCR2(-/-) mice. Cells from the spleen, mesenteric lymph nodes, and peripheral lymph nodes of CCL2(-/-) and CCR2(-/-) mice fed high doses of OVA showed robust proliferative responses compared with cells from Ag-fed wild-type mice. CCL2(-/-) and CCR2(-/-) mice also produced high amounts of Th1 cytokines such as IL-2 and IFN-gamma and very low amounts of IL-4 and IL-10. The ability of APCs from the gut of CCL2(-/-) and CCR2(-/-) OVA-fed mice to stimulate an indicator T cell line was evaluated. APCs from the Peyer's patch of OVA-fed knockout animals could induce a T cell response measured by an increase in proliferation and generation of IL-12 and IFN-gamma with a concomitant reduction of TGF-beta compared with wild-type controls that did not induce a Th1 response. These data indicate that CCL2 and signaling through its receptor CCR2 is critical for the induction of oral tolerance by regulating Ag presentation leading to a disruption in the balance of inflammatory and regulatory cytokines.

A revisit of mucosal IgA immunity and oral tolerance

Induction of mucosal immunity by oral immunization with protein antigen alone is difficult: potent mucosal adjuvants, vectors, or other special delivery systems are required. Cholera toxin (CT) has been shown to be an effective adjuvant for the development of mucosal vaccines and, when given with vaccine, induces both mucosal and systemic immune responses via a Th2 cell-dependent pathway. However, and in addition to potential type-I hypersensitivity, a major concern for use of mucosal adjuvants such as CT is that this molecule is not suitable for use in humans because of its inherent toxicity. When we examined the potential toxicity of CT for the central nervous system, both CT and CT-B accumulated in the olfactory nerves/epithelium and olfactory bulbs of mice when given by the nasal route. The development of effective mucosal vaccines for the elderly is also an important issue; however, only limited information is available. When mucosal adjuvanticity of CT was evaluated in aged mice, an early immune dysregulation was evident in the mucosal immune system. The present review discusses these potential problems for effective mucosal vaccine development. Tolerance represents the most common and important response of the host to environmental antigens, including food and commensal bacterial components, for the maintenance of an appropriate immunological homeostasis. We have examined whether Peyer patches could play a more important role for the maintenance of oral tolerance. Using Peyer patch-null mice, we found that mice lacking this gut-associated lymphoid tissue retained their capability to produce secretory IgA antibodies but did not develop normal oral tolerance to protein antigens.

Intraepithelial lymphocytes: origins, distribution, and function.

Intraepithelial lymphocytes (IEL) are associated with the intestinal tract, respiratory tract, genitourinary tract epithelium, and the skin and are the first immune system cells to encounter pathogens that have invaded an epithelial surface. IEL are predominantly T cells (CD3+) with CD8+ cells predominating at most, but not all, sites. Both TCR alphabeta+ and TCR gammadelta+ cells are found within IEL populations and an increasing body of evidence suggests that some IEL may arise extrathymically. The presence within intestinal IEL of cells expressing potentially self-reactive TCR suggests that T cell selection within epithelia may differ from thymic T cell selection although recent evidence suggests that these cells may in fact be nonresponsive. IEL exhibit various cytotoxic activities including alloreactive and virus-specific CTL activity, NK activity and spontaneous cytotoxicity, activities consistent with an immune surveillance or first line of defence role. IEL also appear activated in vivo and secrete a variety of cytokines. Subsets of IEL have been shown to provide B cell help, to play a role in the maintenance of oral tolerance and to regulate epithelial cell function. In this review the morphology, distribution and phenotype of IEL, the potential for extrathymic development and possible functions of this unique lymphoid population are discussed.

Gamma delta T lymphocytes regulate the induction and maintenance of oral tolerance.

Oral administration of Ag, over a period of several days, induces a state of tolerance that is associated with activation of CD8+ T cells that can transfer unresponsiveness to naive syngeneic hosts. We previously demonstrated that these T cells are not CTL precursors and that they inhibit responses by CD8+ CTL, as well as Ab and CD4+ T cell responses. Activation of noncytolytic, CD8+ suppressor T cells by oral Ag is a process that is not understood. In these studies, we asked whether depletion of the gamma delta T cells altered induction of oral tolerance. Injection of the anti-delta-chain Ab (GL3) down-modulated the expression of gamma delta TCR and inhibited the induction of oral tolerance to OVA, as measured by Ab, CD4+, and CD8+ T cell responses. GL3 did not activate IL-2 secretion that could be detected in the serum, nor did it induce IL-2R expression by intraepithelial lymphocytes, suggesting that GL3 inhibited the function of gamma delta T cells rather than activating them. This interpretation is supported by our observation that oral administration of Ag did not induce tolerance in TCR-delta knockout mice. These data suggest that gamma delta T cells play a critical, active role in tolerance induced by orally administered Ag.

Effect of in Vivo Depletion of CD4+ and CD8+ Cells on the Induction and Maintenance of Oral Tolerance

The role of CD8+ and CD4+ cells in the generation and maintenance of high-dose oral tolerance has been investigated. Mice depleted of CD8+ cells prior to or subsequent to oral tolerization with OVA continued to express suppressed humoral and/or T cell proliferative responses to the homologous antigen. To assess the effect of CD4+ cell depletion on oral tolerance, mice were depleted of CD4+ cells prior to or subsequent to oral tolerization with OVA. After 50% regeneration of CD4+ cells was achieved, OVA-immunized mice produced anti-OVA immune responses that were comparable to those of water-fed control mice. Thus, under our experimental conditions the presence of CD4+ but not CD8+ cells is required to both establish and maintain oral tolerance.