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Oral Tolerance: Immune Mechanisms and Treatment of Autoimmune Diseases
Abstract
Orally administered proteins induce systemic hyporesponsiveness to the fed protein. The mechanism underlying such ‘oral tolerance’ depends on the amount of antigen fed. with higher doses inducing deletion and anergy, and lower doses inducing regulatory cells. Orally administered autoantigens suppress many experimental autoimmune diseases, as discussed here by Howard Weiner.
... Upon stimulation of the undifferentiated, ThO CD4+ T cell, there is the capacity to develop, depending on the APC, antigen concentration, number of exposures and the cytokine environment, into either Thl or Th2 subsets (Constant and Bottomly, 1997). More recently, a Th3 subset has been reported, that secretes TGF-P and are thought to be a unique subset of regulatory cells (Weiner, 1997). ...
... The Th2 subset cytokine profile secretes cytokines such as IL-4 and IL-5 (in mice only), which help B cells to proliferate and differentiate and are associated with humoral type responses facilitating antibody production (Constant and Bottomly, 1997). Th3 cells have been implicated in the mechanisms involved in oral tolerance (Fukaura et al., 1996;Weiner, 1997). IL-12 released from APC is thought to direct ThO cells to Thl (Hsieh et al., 1993) and IL-4 is believed to direct the precursor ThO cells to Th2 (Le Gros et al, 1990) and Th3 cells (Inobe et al., 1998). ...
This study has demonstrated that a highly reproducible model of experimental autoimmune uveoretinitis (EAU) can be achieved in the BIO.RIII mice using a human interphotoreceptor retinoid binding protein peptide corresponding to amino acid residues 161-180. Using a histological grading system which graded both cellular and structural scores individually, the time course of disease in this model was examined and found to be predictable and consistent. Disease was acute in nature characterised by rapid onset of a massive inflammatory response which resulted in extensive damage to the rod outer segments and neuronal layers of the retina. Treatment with potent immunosuppressive agents, CD4 monoclonal resulted in the inhibition of disease and a reduction in disease incidence. In addition, treatment with p55-tumor necrosis factor receptor-Ig (p55-TNFR-Ig) fusion protein reduced structural damage to the retina despite a high level of cellular infiltration in the eye. This suggests that target organ damage in an acute model of EAU can be modulated. However, the value of the B10.RIII human IRBP161-180 induced model maybe of limited value in the study of immunosuppressive agents. The albino Biozzi ABHdql mice are susceptible to experimental allergic encephalomyelitis (EAE), however their susceptibility to EAU induction was unknown. Initial, studies demonstrated that ABH mice were susceptible to EAU induction with native bovine interphotoreceptor retinoid binding protein (IRBP), but not S-Antigen (S-Ag). The uveitogenic domain of interphotoreceptor retinoid binding protein was then identified for this mouse strain using recombinant domains of human IRBP expressed from an Escherichia coli expression system. Following sensitisation with the individual domains of IRBP, ABH mice were found to be susceptible to disease induction with IRBP domain 2. The mice developed disease with high incidence and moderate disease scores. Examination of the disease course over a six week period indicated that disease was of a chronic progressive nature. However, in this model although an inflammatory insult had occurred in the eye the resulting level of retinal destruction less severe than that seen in the B10.RIII IRBP161-180 induced model of EAU and an intact retina still remained at six weeks post-immunisation. Using synthetic overlapping peptides corresponding IRBP domain 2, a uveitogenic and immunogenic epitope was identified. The uveitogenic epitope corresponded to human IRBP511-530 and incidence disease with high incidence and mild disease scores. Further immunogenic epitopes residing in IRBP domain 2 were identified which may be involved in the amplification of disease in the ABH mice. IRBP511-530 also contains some characteristics which have been identified to be involved in the H-2Ag7 binding motif. However, further analysis of the amino acid residues is required to determine the important residues. This study also examined EAU in (ABH X BIO.RIII) FI mice and showed that these mice are susceptible to disease induction with retinal antigens which are positive in both B10.RIII and ABH mice. Furthermore, (ABH X BiO.RIII) FI mice are susceptible to a novel antigen, corresponding to IRBP domain 3, which is not uveitogenic in either the ABH or B10.RIII mice. The (ABH X B10.RIII) FI mice may provide a useful system for studying the effects of B10. background genes and bystander suppression studies.
... Ambivalent sexism, which has received much attention in the literature, is acknowledged as a determinant of attitudes toward SH. [24] examined the impact of gender and ambivalent sexism on perceptions of sexual harassment. Dominant authoritarianism, which emphasizes male dominance over women, is a form of hostile sexism. ...
... Механизм пероральной толерантности инициируется в лимфоидной ткани, ассоциированной с кишечником (gut-associated lymphoid tissues, GALT), но влияет на системный иммунитет [84]. Большие скопления GALT формируют пейеровы бляшки [85], а куполообразный эпителий, располагающийся над ними, содержит интраэпителиальные лимфоциты и эпителиальные клетки, часть из которых имеет микроскладки (М-клетки). М-клетки транспортируют Аг из просвета кишечника в пейеровы бляшки, где формируется иммунный ответ в слизистой оболочке. ...
The review considers the full spectrum of currently known autoantigens in osteoarthritis (OA) and discusses their role in the development and/or persistence of synovitis and the initiation of subsequent destruction of articular cartilage with the development of an autoimmune response and auto-inflammation. Of great interest are methods of drug prevention of OA considering autoimmunity responses and associated auto-inflammation, including the use of pharmaconutraceuticals.
Preclinical and clinical studies of the safety and efficacy of pharmaconutraceuticals containing native type II collagen are presented. A clear relationship between the composition/chemical structure of the collagen components and its mechanism of action and efficacy is discussed. Taking into account the autoimmune pathogenesis of OA, new combined pharmaconutraceuticals aimed at reducing the manifestations of autoinflammation (chondroitin sulfate, glucosamine sulfate) are developed. They have an optimal ratio of active ingredients with a sufficient level of evidence, which allows enhancing their beneficial pharmacological effects.
... Gut-Associated Lymphoid Tissue, GALT) в двенадцатиперстной кишке, индуцируют оральную толерантность к антигенам и снижают выраженность повреждающего действия Т-лимфоцитов на хрящевую ткань. Небольшие дозы перорального гликозилированного НК-II представляют собой активные эпитопы с правильными трехмерными структурами для GALT, что помогает сформировать иммунную толерантность при ОА [63]. ...
In recent decades, there has been an increase in the prevalence and medical and social burden of osteoarthritis (OA) and nonspecific back pain in all countries of the world. The First Multidisciplinary Bilateral Russia-Uzbekistan Expert Council presented innovations in the prognosing, personalized prevention and adjuvant therapy of degenerative-dystrophic diseases of the joints and spine, the evidence base for the effectiveness and safety of the use of drugs that modify the course of OA (Disease-modifying osteoarthritis drugs, DMOADs): chondroitin sulfate, glucosamine sulfate, undenatured type II collagen for adjuvant pharmaconutraceutical support – prevention and adjuvant therapy (treatment) of OA and nonspecific lumbosacral pain. The expert counsil resolution presents an optimized algorithm for the management, prevention and adjuvant therapy of OA and non-specific back pain, maintaining the function of healthy joints after intense physical activity with the inclusion of the drug Chondroguard solution for intra-articular and intramuscular administration (INN – chondroitin sulfate) and a new pharmaconutraceutical from the DMOADs group – TRIO trademark Chondroguard® (Chondroguard®TRIO).
... Immunosuppressive cytokines, such as TGF- and IL-10, have been implicated in the development of peripheral tolerance (5,6). To examine whether the presence of these cytokines was essential for the induction of unresponsiveness by i.n. ...
In this study we examined the immunological parameters underlying the natural immunity to inhaled nonpathogenic proteins. We addressed this question by examining the effect of intranasal exposure to OVA in both wild-type mice and mice reconstituted with OVA-TCR transgenic CD4+ T cells. Intranasal administration of OVA induced an initial phase of activation during which CD4+ T cells were capable of proliferating and producing cytokines. Although many of the OVA-specific CD4+ T cells were subsequently depleted from the lymphoid organs, a stable population of such T cells survived but remained refractory to antigenic rechallenge. The unresponsive state was not associated with immune deviation due to selective secretion of Th1- or Th2-type cytokines, and the presence of regulatory CD8+ T cells was not required. Moreover, neutralization of the immunosuppressive cytokines IL-10 and TGF-β did not abrogate the induction of tolerance. Inhibition of the interaction of T cells with CD86, but not CD80, at the time of exposure to intranasal Ag prevented the development of unresponsiveness, while selective blockade of CTLA-4 had no effect. Our results suggest that intranasal exposure to Ags results in immunological tolerance mediated by functionally impaired CD4+ T cells via a costimulatory pathway that requires CD86.
... Mucosal sites that are suitable for tolerance induction are both the gastro-intestinal tract (1,2) and the airway system (3)(4)(5)(6). Tolerance can involve clonal deletion, clonal anergy, and active T cell-mediated suppression (7). When tolerance is induced via the intranasal route, active suppression is involved as demonstrated by adoptive transfer of splenocytes (8). ...
Mucosal tolerance is a naturally occurring immunological phenomenon that prevents harmful inflammatory responses to ingested or inhaled environmental, predominantly nondangerous, Ags. The nasal mucosa is an extremely efficient compartment in the induction of immunological tolerance which can be exploited in Ag-specific treatment of autoimmune disease. With the use of a model Ag (OVA) and an Ag implicated in the autoimmune disease rheumatoid arthritis (human cartilage gp-39), we here show in a mouse model that the superficial cervical and internal jugular lymph nodes that drain the nasal mucosa are instrumental in the induction of tolerance. Removal of these lymph nodes abrogates tolerance induction, which can be restored by transplantation of superficial cervical lymph nodes, but not of peripheral lymph nodes. The results indicate that lymph nodes that directly drain the nasal mucosa constitute a unique microenvironment which favors the induction of immunological tolerance.
... Antigen has to be administered frequently in doses greater than immunogenic doses to induce tolerance. The autoimmune disorders which can be prevented are type I diabetes, multiple sclerosis, rheumatoid arthritis, graft rejection, etc. [53]. To suppress immune attack and to delay the onset of high blood sugar, potatoes expressing insulin and a protein called Glutamic acid Decarboxylase (GAD), linked to CT-B subunit were fed to mouse and the results were effective. ...
Infectious diseases cause more than one million deaths every year. Fifty percent of these diseases are caused by bacteria that infect the mucosal membrane of the mammalian host. Vaccines are recognized worldwide as one of the most effective resources against infectious diseases. It is a biological product that can improve the immune response to specific diseases. Edible vaccines are referred to the use of edible parts of the genetically modified plants. It effects on the lining of the gastrointestinal tract allows the activation of systemic immunity and mucosal immunity (GIT). Edible vaccines are used to prevent various diseases, such as hepatitis B, measles, malaria, cholera, Norwalk disease, anthrax, foot-and-mouth disease, rabies, rotavirus, HIV, HPV, diabetes, and sexually transmitted diseases. The purpose of this review is to introduce edible vaccines as a novel oral immunization method, the types and uses of edible vaccines in clinical trials.
... Rheumatoid arthritis, diabetes, multiple sclerosis, lupus, and transplant rejection are among the most common autoimmune disorders being researched (Weiner, 1997). In one clinical investigation, diabetic mice were fed potatoes that could produce glutamic acid decarboxylase (GAD) protein and insulin. ...
The researchers are still doing efforts to develop an effective, reliable, and easily accessible vaccine candidate to protect against COVID‐19. As of the August 2020, nearly 30 conventional vaccines have been emerged in clinical trials, and more than 200 vaccines are in various development stages. Nowadays, plants are also considered as a potential source for the production of monoclonal antibodies, vaccines, drugs, immunomodulatory proteins, as well as used as bioreactors or factories for their bulk production. The scientific evidences enlighten that plants are the rich source of oral vaccines, which can be given either by eating the edible parts of plants and/or by oral administration of highly refined proteins. The use of plant‐based edible vaccines is an emerging trend as it possesses minimum or no side effects compared with synthetic vaccines. This review article gives insights into different types of vaccines, the use of edible vaccines, advantages of edible vaccines over conventional vaccines, and mechanism of action of edible vaccines. This review article also focuses on the applications of edible vaccines in wide‐range of human diseases especially against COVID‐19 with emphasis on future perspectives of the use of edible vaccines.
... The Interleukin6 and Iinterleukin8 secreted by the epithelial cells favors inflammation, so a strong epithelial stimulation can cause an inflammatory effect on immune response. Weiner (1997) suggested three types of interaction with the intestinal cells to induce an immunity response: ...
Fermented milk products containing Lactic Acid Bacteria have been reported to possess a beneficial effect on human including enhanced immune performance, attenuation of lactose intolerance, increased resistance to diseases causing microbes, alleviation of food allergies and the suppression of cancer growth. An optimally working immune system is essential for defense against infectious diseases and cancers. Deficit in any component of the immune system can expose an individual to a greater risk of infection or may increase the severity of disease. LAB are extensively used for the manufacture of fermented foods, especially dairy products are known to have excellent nutritional and organoleptic effect on milk. In addition, LAB has been found to exhibit a range of physiological and therapeutic effects in the consumer including stimulation of immune system. In this chapter we will discuss the basics of human immunity and effects of consumption of fermented foods/functional foods on it.
... It has been shown in animal models that oral feeding of protein antigens can downregulate systemic immune responses, known as oral tolerance [14,15]. Oral administration of therapeutic molecules theoretically offers advantages such as ease of administration and reduction in adverse effects. ...
Allergic airway disease is the most common chronic airway inflammatory disorder in developed countries. House dust mite, cockroach, and mold are the leading allergens in most tropical and subtropical countries, including Taiwan. As allergen avoidance is difficult for patients allergic to these perennial indoor allergens, allergen-specific immunotherapy (ASIT) is the only available allergen-specific and disease-modifying treatment. However, for patients sensitized to multiple allergens, ASIT using each corresponding allergen is cumbersome. In the present study, we developed a recombinant L . lactis vaccine against the three most common indoor aeroallergens and investigated its effectiveness for preventing respiratory allergy and safety in mice. Three recombinant clones of Der p 2 (mite), Per a 2 (roach), and Cla c 14 (mold) were constructed individually in pNZ8149 vector and then electroporated into host strain L . lactis NZ3900. BALB/c mice were fed with the triple vaccine 5 times per week for 4 weeks prior to sensitization. The effectiveness and safety profile were then determined. Oral administration of the triple vaccine significantly alleviated allergen-induced airway hyper-responsiveness in the vaccinated mice. The allergen-specific IgG2a was upregulated. IL-4 and IL-13 mRNA expressions as well as inflammatory cell infiltration in the lungs decreased significantly in the vaccinated groups. No body weight loss or abnormal findings in the liver and kidneys were found in any of the groups of mice. This is the first report to describe a triple-aeroallergen vaccine using a food-grade lactococcal expression system. We developed a convenient oral delivery system and intend to extend this research to develop a vaccination that can be self-administered at home by patients.
... In the gastrointestinal (GI) tract, oral tolerance is mediated by antigen exposure to CD103+ DCs. Upon antigen uptake and processing, CD103+ DCs migrate to the mesenteric lymph nodes (MLNs), which are the lymphoid organs draining the gut, where they drive the differentiation of naïve T cells to different types of regulatory T cells (T regs ) with the help of cytokines, enzymes, and metabolite signaling such as TGF-β, indoleamine 2,3-dioxygenase, and retinoic acid 36,37 . However, the access of therapeutic proteins in their original forms to gut-associated lymphoid tissue (GALT) is challenging due to the harsh conditions of oral delivery. ...
The safety and efficacy of several life-saving therapeutic proteins are compromised due to their immunogenicity. Once a sustained immune response against a protein-based therapy is established, clinical options that are safe and cost-effective become limited. Prevention of immunogenicity of therapeutic proteins prior to their initial use is critical as it is often difficult to reverse an established immune response. Here, we discuss a rational design and testing of a phosphatidylserine-containing nanoparticle platform for novel oral prophylactic reverse vaccination approach, i.e., pre-treatment of a therapeutic protein in the presence of nanoparticles to prevent immunogenicity of protein therapies.
... Many studies have sought to remedy autoimmune diseases, like rheumatoid arthritis and encephalomyelitis, by inducing oral tolerance which is actively mediated by Tregs (8,9). However, IBD patients have defective oral tolerance responses (10). ...
Background: Inflammatory bowel disease (IBD) involves an increase in T effector cells in the intestines that disrupts the normal balance with T regulatory cells (Tregs). A therapy that restores this balance has the potential to treat IBD. We have shown that epicutaneous exposure to OVA induces Tregs that are able to induce tolerance. The Tregs also migrate to the intestines where they alleviate colitis in mice, demonstrating the potential for skin induced Tregs to treat intestinal inflammation. We investigated the role of Foxp3, IL-10, and TGF-β in the suppression of colitis by epicutaneous immunotherapy (ET).
Methods: RAG1 −/− mice were transferred with CD4 ⁺ CD45RB hi T cells from wild type mice to induce colitis. To determine whether Foxp3 ⁺ Tregs, IL-10-, or TGF-β-producing Tregs were necessary, Foxp3-DTR, IL-10 −/− , or CD4-dnTGFBRII mice were immunized with OVA and OVA TCR enriched T cells were added. As control groups, some mice were given OVA TCR enriched T cells from wild type mice or no OVA TCR enriched T cells. Half of the mice in each group were then exposed on the skin to Viaskin patches containing OVA weekly for 3 weeks. Mice given OVA TCR enriched T cells from Foxp3-DTR mice were given diphtheria toxin (DT) or not in addition to ET. Mice were assessed for weight loss, colon length, colonic cytokine production, and histological inflammation.
Results: ET, after injection with OVA TCR enriched T cells derived from wild type mice, prevented weight loss, decreased colonic inflammatory cytokine production and histological colitis. ET in the absence of the OVA TCR enriched T cells did not alleviate colitis. ET, after injection with OVA TCR enriched T cells derived from Foxp3-DTR mice, prevented weight loss, decreased colonic inflammatory cytokine production, and histological colitis. Ablation with DT did not impair the ability of ET to alleviate colitis. ET failed to alleviate colitis when OVA TCR enriched T cells were derived from IL-10 −/− or CD4-dnTGFBRII mice.
Conclusions: ET through induction of Tregs, which produce IL-10 and TGF-β, could be a promising treatment for IBD.
... It is well established that mucosal administration of soluble antigens induces systemic immunological unresponsiveness, a phenomenon known as mucosal tolerance (1,2). However, it has been recognized that the effectiveness of mucosal tolerance often requires repeated and large amounts of antigen, and sometimes the suppressed immune responses are of short duration. ...
... Another subset of adaptive Treg are the Th3 cells, which are a unique population of T helper cells induced by oral tolerance to non-self-antigens. Th3 cells produce high concentrations of TGFb and moderate amounts of IL-10 (38). In contrast to Tr1 cells which do not express Foxp3, some Th3 cells are TGFb-induced-Foxp3 + cells (51). ...
The immune system is a tightly regulated network which allows the development of defense mechanisms against foreign antigens and tolerance toward self-antigens. Regulatory T cells (Treg) contribute to immune homeostasis by maintaining unresponsiveness to self-antigens and suppressing exaggerated immune responses. Dysregulation of any of these processes can lead to serious consequences. Classically, Treg cell functions have been described in CD4+ T cells, but other immune cells also harbour the capacity to modulate immune responses. Regulatory functions have been described for different CD8+ T cell subsets, as well as other T cells such as γδT cells or NKT cells. In this review we describe the diverse populations of Treg cells and their role in different scenarios. Special attention is paid to the aging process, which is characterized by an altered composition of immune cells. Treg cells can contribute to the development of various age-related diseases but they are poorly characterized in aged individuals. The huge diversity of cells that display immune modulatory functions and the lack of universal markers to identify Treg make the expanding field of Treg research complex and challenging. There are still many open questions that need to be answered to solve the enigma of regulatory T cells.
... This process is mediated through different pathways, including immune checkpoints (ICs) (1). When the immune system is too activated, it may lead to autoimmune diseases (2,3); when it is too anergic, it contributes to infection and cancer spread (4). T lymphocytes have a central role in this homeostasis. ...
T cells have a central role in immune system balance. When activated, they may lead to autoimmune diseases. When too anergic, they contribute to infection spread and cancer proliferation. Immune checkpoint proteins regulate T cell function, including cytotoxic T lymphocyte antigen-4 (CTLA-4) and programmed cell death-1 (PD-1) and its ligand (PD-L1). These nodes of self-tolerance may be exploited pharmacologically to downregulate (CTLA-4 agonists) and activate [CTLA-4 and PD-1/PD-L1 antagonists, also called immune checkpoint inhibitors (ICIs)] the immune system. CTLA-4 agonists are used to treat rheumatologic immune disorders and graft rejection. CTLA-4, PD-1, and PD-L1 antagonists are approved for multiple cancer types and are being investigated for chronic viral infections. Notably, ICIs may be associated with immune-related adverse events (irAEs), which can be highly morbid or fatal. CTLA-4 agonism has been a promising method to reverse such life-threatening irAEs. Herein, we review the clinical pharmacology of these immune checkpoint agents with a focus on their interplay in human diseases.
Expected final online publication date for the Annual Review of Pharmacology and Toxicology, Volume 61 is January 8, 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.
... For instance, among the adjuvants, CFA and IFA preferably can stimulate Th1 and Th2 responses, respectively. Beside adjuvants, different routes of receiving the immunogens, such as IP (Wang et al., 1996), oral (Weiner, 1997), and nasal (Tian et al., 1996) discriminately can modulate Th2 type responses. For achieving a better vaccination results in this area, the vaccine should be designed to stimulate Th2 cells to obtain strong humoral immunity and decrease the side effect of Th1 cells such as delay type hyper sensitivity. ...
The functional competence of leukemia inhibitory factor (LIF), as immunocontraceptive vaccine in mice, was investigated. Balb/c mice were divided into two groups of vaccinated and controls. The recombinant human LIF (rhLIF) protein and phosphate buffer saline was emulsified with Freund's adjuvant and injected into vaccinated and control groups, respectively. The inhibition of implantation was evaluated in mice uterine. The concentration of secreted interferon-γ (IFN-γ) and interleukin (IL)-4 were measured in cultured splenocyte of mice stimulated by rhLIF. The expressions of immune responsive gene 1 (IRG-1), cochlin (COCH), amphiregulin (Ar), and heparin-binding EGF-like growth factor (HB-EGF) genes were determined. Mice were assessed for inhibition of fertility after delivery, reversibility of immune response against rhLIF, and survival rate. Active immunization of mice with rhLIF resulted in reduction of the implantation and fertility rate up to 80.49% and 75%, respectively. All mice produced a high titer of anti-rhLIF antibodies in serums and vaginal fluids washes after 16 weeks; however, these antibodies were cleared from vaginal fluid washes after six months. A significant down-regulation in mRNA levels of IRG-1, Ar and HB-EGF was observed in vaccinated group compared to controls; however, no significant change in the expression profile of cochlin gene was detected. The results showed that rhLIF prevented pregnancy in a high percentage of female mice. Although the immunization of female Balb/c mice with rhLIF inhibited fertility and expression of genes associated with this molecule, further studies are needed to support this protein as a suitable candidate for contraceptive vaccine in mammals.
... Cette compétence tol og e pe et à la g effe de foie d' t e u e des t a spla tatio s la mieux tolérée(Adams et al., systèmes HLA (Antigènes Leucocytaires Hu ai s e t e le do eu et le e eveu 'est pas directement impliquée dans le rejet de greffe. Le foie a également un rôle important dans la tolérance orale(Weiner, 1997). La situatio vas ulai e h pati ue pe et au a tig es i g s d' t e p se t s et tolérés lors de leur passage dans les sinusoïdes, et par extension dans le reste du corps. ...
Afin de lutter contre des agressions aiguës pouvant être induites par une hépatite virale, des toxines ou bien une surcharge médicamenteuse, le foie active différents processus interconnectés tels que la prolifération hépatocytaire, l'inflammation ou la mort cellulaire. Une protéine clé se trouve au carrefour de ces différents processus cellulaires : la protéine p38 alpha MAP (Mitogen Activated Protein) Kinase. Dans le foie, de nombreux travaux décrivent un double rôle de cette protéine. En effet, elle est considérée comme un suppresseur de tumeur en régulant négativement la prolifération des hépatocytes, cellules majoritaires du foie, mais peut aussi agir comme un oncogène en favorisant certains processus liés au cancer et notamment l'inflammation. Cependant, son action lors d'une agression aiguë du foie adulte reste peu caractérisée. L'objectif de mon travail de thèse était donc de caractériser le rôle de la protéine p38 alpha lors d"une agression hépatique aiguë, grâce à un modèle murin pour lequel l'expression de cette protéine est invalidée spécifiquement dans les hépatocytes (modèle murin KO). Pour répondre à cette question, nous avons utilisé le modèle expérimental d'hépatopathie classiquement utilisé pour reproduire la division hépatocytaire en contexte inflammatoire : le modèle du tétrachlorure de carbone (CCl4). L'injection de cet hépatotoxique à des souris entraîne une cytolyse hépatocytaire dans les zones centro-lobulaires rapidement suivie par une infiltration de cellules immunitaires et une prolifération compensatoire des hépatocytes sains, pour réparer le tissu lésé. Des souris contrôles et KO ont été sacrifiées à différents temps après injection permettant d'établir une cinétique post-CCl4, au cours de laquelle du sang et du tissu hépatique ont été collectés. Les lésions hépatiques, l'inflammation, l'apoptose et la prolifération ont ensuite été étudiées. Tout d'abord, nous avons observé que le nombre d'hépatocytes en phase S (marquage BrdU, taux de CyclinA2) est diminué en l'absence de p38a au cours de la phase de régénération post-CCl4. Par ailleurs, nous montrons que la perte d'expression de la protéine p38a confère un effet protecteur contre les lésions hépatiques induites par le CCl4. En effet, les zones de cytolyse et les taux d'alanine transaminase (reflet de la mort hépatocytaire) sont significativement diminués (de 40H à 60H post-CCl4) chez les souris KO. Pour expliquer ce phénotype, nous avons analysé l'apoptose au cours de la cinétique de régénération, mais nos résultats ne montrent pas de différences significatives dans la mise en place de ce processus entre les animaux contrôles et KO. En revanche, nous montrons que la réponse anti-oxydante dans le foie des animaux KO est amplifiée et pourrait donc participer à la protection du tissu hépatique. Enfin, nos observations immunohistochimiques montrent un recrutement massif de cellules inflammatoires dans les zones de cytolyse corrélé à une augmentation importante des niveaux de cytokines (comme le TNFa) et chimiokines pro-inflammatoires (CCL2 et CCL5). Laensemble de ces données suggèrent fortement que la déficience en p38a induit une réponse immunitaire spécifique favorisant le nettoyage et la réparation du tissu hépatique. En conclusion, mes travaux de thèse montrent que lors d'une lésion aiguë l'absence de la MAP Kinase p38a protège le foie en ajustant la balance « prolifération - réponse inflammatoire », favorisant ainsi une réparation précoce du tissu et le maintien de l'homéostasie hépatique.
... Evoking an immune response against such antigens would induce constant infl ammation in the gut, causing chronic infl ammatory diseases observed in food allergies or intolerances [13][14][15]. The phenomenon of suppression of immune response against non-harmful antigens from gut microbiota or food antigens is called oral tolerance [16,17] ( Figure 1). DCs also have the ability to extend their pseudopodia through the tight junctions to sample the antigens in the intestinal lumen [18,19]. ...
... Toxin binders in the diet help to reduce the absorption of toxin from the gut (Magnoli et al., 2011). Higher antibody titre is due to the binding ability and lower absorption of toxin from gut with mycotoxins (Weiner, 1997). Clay-based toxin binder also stimulates the indirect activation of B cells, T cells and macrophages (Lyons & Jacques, 1994). ...
The aim of this study was to evaluate the effects of commercial toxin binders on growth performance, organ morphology, immunity and serum biochemistry in broilers. Dietary treatments consisted of the negative control (NC): experimental diet with aflatoxin B1
... One potentially attractive therapy for autoimmunity is the induction of autoantigen-specific "mucosal tolerance" (Weiner 1997). In experimental studies, protein antigens administered orally (or nasally) have been found to produce suppression of humoral and cellular immune responses on subsequent immunisation. ...
Autoimmune uveitis is a non-infective ocular inflammation of humans that potentially causes blindness. It is mediated primarily by autoreactive CD4+ Tlymphocytes that target antigens within the retina. Autoantibody responses play a secondary role. This thesis describes various investigations into humoral and cellular immune responses in autoimmune uveitis and assesses several methodologies for their suitability to applied human research. Recombinant human retinal S antigen (RSAg), an important candidate autoantigen in uveitis, was cloned and expressed in bacterial and human cells. Purified RSAg was tested by ELISA against sera from uveitis patients and controls. The recombinant antigens performed well in ELISA. No significant differences in antibody titres were detected between the groups. B-cell epitope preferences of anti-RSAg polyclonal antibodies were investigated by screening several random phage display libraries. One library produced results, but no defining epitope was identified for either uveitis or control sera. In uveitis research this technique might be better suited to delineating minimal epitope requirements of monoclonal antibodies. As yet undiscovered uveitis autoantigens may exist. A human retinal complementary DNA library was constructed and screened with uveitis and control sera. Seven potentially autoantigenic peptides were identified and expressed as fusion proteins. At least one peptide displayed significantly higher ELISA readings for uveitis over control sera. The full potential of this technique is still to be realised. Responses of peripheral CD4+ T-lymphocytes to antigen-specific stimulation were studied at the single-cell level using cytokine flow cytometry. A definite response to RSAg was detected in human uveitis and control lymphocytes using this method. This technique has great potential for identifying autoantigenic proteins/epitopes, and analysing resultant cytokine profiles in uveitogenic T-cells. Several of the new strategies and techniques described here have already produced exciting findings. It is envisaged that they will make further significant contributions to applied human uveitis research in the near future.
... These results suggest that oral tolerance may develop towards HBeAg or against HBcAg (which shares around 80% of the HBeAg primary sequence) by virtue of the HBeAg being secreted by the salivary glands, swallowed and treated by the immune system as a dietary antigen. It is hypothesized that mucosal administration of antigens in a Th1immunomodulatory environment may help in the subversion of such oral tolerance(27).Recent reports from Boni et al. reveal an important degree of T-cell functional recovery after antiviral treatment with Lamivudine(28,29), suggesting that it is possible to rescue T cells from non-responsive states. Other reports demonstrate that in patients naturally recovering from HBV chronic infection, there is a rebound in cellular immunity that correlates with viral clearance(30). ...
Trial study protocol.
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... Mucosal vaccines, despite a number of potential advantages over injectable ones (such as no need of injections and hence no risk of transmitting bloodborne diseases, and easy way of administration), are much less common. Most soluble protein antigens introduced via the mucosal route are poorly immunogenic and induce specific, long-lasting tolerance [4][5][6]. Moreover, the problems with rapid antigen degradation on the mucosal surfaces and lack of appropriate mucosal adjuvants primarily contribute to their diminished usefulness [7]. ...
Mucosal immunizations are convenient ways of vaccination, which do not require any trained personnel for administration. One of the major challenges for developing an effective mucosal vaccine is finding appropriate adjuvant. Bacillus subtilis endospores have been shown to help solving these obstacles while serving as a platform for presentation of both, antigens and adjuvants. In this study, we have successfully designed and constructed recombinant spores displaying an antigen/adjuvant chimeric protein. We have used a fragment of Clostridium difficile flagellar cap FliD protein as antigen and VQGEESNDK peptide, a fragment of human IL-1β, as adjuvant. Recombinant spores presenting FliD were able to elicit immune response in orally immunized mice which could be evaluated by detection of FliD-specific IgA antibodies in feces of immunized animals. Moreover, the presence of IL-1β fragment significantly changed characteristics of elicited immune response. Obtained results show that recombinant spores presenting an antigen/adjuvant chimeric protein exhibit both properties in mucosal immunization of mice. Moreover, IL-1β fragment could serve as valuable adjuvant in B. subtilis spore-based mucosal vaccines.
Electronic supplementary material
The online version of this article (10.1007/s12033-018-0117-0) contains supplementary material, which is available to authorized users.
... Treg cells account for 5-10% of CD4 + T cells and may be divided into natural Treg (nTreg) cells and inducible Treg (iTreg) cells (3). nTreg mainly refers to CD4 + CD25 + Treg cells and iTreg refers to transforming growth factor (TGF)-β-secreting T helper lymphocyte 3 (Th3) and IL-10-secreting type 1 regulatory T (Tr1) cells (4)(5)(6). Furthermore, immunosuppressive CD8 + Treg and natural killer T cells, which exhibit bidirectional immunity, represent two categories of Tregs (7,8). Previous studies on Treg cells mainly focused on CD4 + CD25 + Treg, Th3 and Tr1 cells. ...
The purpose of the present study was to investigate the role of latency-associated peptide (LAP)⁺ CD4⁺ T cells in hepatocellular carcinoma (HCC) immunity. Flow cytometric analysis was performed to detect the proportion of LAP⁺ CD4⁺ T cells among the peripheral blood mononuclear cells (PBMCs) of 30 HBV-infected HCC patients at the pre-operative and post-operative stages, as well as 30 hepatitis B virus (HBV)-infected volunteers as a control group. Furthermore, tumor tissues and peri-tumor tissues from 28 patients with HCC, as well as hepatic tissues from 28 HBV-infected patients with benign lesions were subjected to immunohistochemical analysis with double staining for LAP and CD4, and the average number of the LAP⁺ CD4⁺ T cells in each visual field was quantified. The results indicated that the proportion of LAP+CD4+ T cells in the PBMCs of patients with HCC was significantly higher than that in the control group (1.84±0.85 vs. 0.73±0.39%, P=0.019), while it was significantly reduced after the operation (1.07±0.35, P=0.021), but still slightly, if not significantly, higher compared with that in the control group (P=0.342). Furthermore, the number of LAP⁺ CD4⁺ T cells per high-magnification microscopic field (magnification, x400) in the HCC tissues was 11.25±3.00, which was significantly higher than that in the peri-cancer tissues (5.75±1.00) and that in the HBV-infected hepatic tissues around benign lesions (2.61±0.83). In peri-cancer tissues, LAP⁺ CD4⁺ T cells were also significantly more abundant than in control tissues. Furthermore, in the HCC tissues, LAP⁺ CD4⁺ T cells were present as clusters in the tumor stroma and closely associated with CD4⁺ T lymphocytes. By contrast, in the peri-cancer liver tissues and HBV-infected hepatic tissues around benign lesions, LAP⁺ CD4⁺ T cells were sparsely distributed. LAP⁺ CD4⁺ T cells have marked inhibitory effects, and in the peripheral blood and tumor tissues of patients with HCC, they have an important role in the suppression of anti-tumor immunity and in the immune evasion of tumor cells.
... Compared to IgG, IgM, and IgE, antibodies of the IgA class are much less proinflammatory and can even be considered antiinflammatory (8). IgA antibodies thus contribute to the general downimmunoregulatory tone of the mucosal immune system (20), which has the dual role of protecting against ubiquitous foreign substances and microbes, while at the same time not subjecting the mucosa to undue inflammation. In a sense, the limited ability of the Fc portion of IgA to activate proinflammatory phenomena can be viewed as part of the evolutionary focusing of the secondary functions of IgA on the ability to bind to the pIgR, which provides a mechanism for transport across mucosal epithelium. ...
The humoral arm of the mucosal immune system is principally composed of locally synthesized polymeric IgA, whose Fc portion is adapted for binding to the polymeric immunoglobulin receptor that is expressed on the basolateral surface of mucosal epithelial cells, including enterocytes. This receptor mediates the endocytosis and transcytosis of polymeric IgA, which allows IgA to function in host defense at three anatomic levels in relation to mucosal epithelium: IgA antibodies in the lamina propria can bind antigens and excrete them through the epithelium into the lumen; antiviral IgA antibodies in transit through epithelial cells can inhibit virus production by an intracellular action; and IgA antibodies secreted into the lumen can prevent antigens and microbes from adhering to and penetrating the epithelium. The ways in which IgA antibodies function in mucous membranes provide challenging investigative opportunities for cell physiologists and cell biologists.
Phosphatidylserine (PS) is an anionic phospholipid exposed on the surface of apoptotic cells. The exposure of PS typically recruits and signals phagocytes to engulf and silently clear these dying cells to maintain tolerance via immunological ignorance. However, recent and emerging evidence has demonstrated that PS converts an "immunogen" into a "tolerogen", and PS exposure on the surface of cells or vesicles actively promotes a tolerogenic environment. This tolerogenic property depends on the biophysical characteristics of PS-containing vesicles, including PS density on the particle surface to effectively engage tolerogenic receptors, such as TIM-4, which is exclusively expressed on the surface of antigen-presenting cells. We harnessed the cellular and molecular mechanistic insight of PS-mediated immune regulation to design an effective oral tolerance approach. This immunotherapy has been shown to prevent/reduce immune response against life-saving protein-based therapies, food allergens, autoantigens, and the antigenic viral capsid peptide commonly used in gene therapy, suggesting a broad spectrum of potential clinical applications. Given the good safety profile of PS together with the ease of administration, oral tolerance achieved with PS-based nanoparticles has a very promising therapeutic impact.
Expansion of plant engineering technologies has developed new techniques to introduce new plant‐based pharmaceutical products. The approach to produce plant‐based edible vaccine emerged in the early 1990s after the worldwide attention to produce safer, easier, and more effective vaccines for the cure of severe diseases. Hiatt and coworkers made the first attempt to produce plant‐based vaccine in 1989. Many research groups around world are putting their best efforts to get the opportunity to produce plant‐based vaccines against numerous diseases. Several new approaches like biolistic, agroinfiltration, electroporation, sonication, and polyethylene glycol treatment have been established for the production of these vaccines. These vaccines are cost effective, heat stable, and less toxic, which makes them extremely useful against the conventional vaccines especially for the poor countries. Till date, numerous transgenic plants have been employed to produce these vaccines against bacterial, viral, and parasitic infections. Currently, many vaccines are in the clinical trial phase, and several have been produced. Despite the wonderful and motivating concept of producing plant‐based vaccines using genetic engineering and plant biotechnology, there are a number of challenges to reach the final goal of manufacturing commercially available vaccine.
Gut microbes comprise of millions of microbes together called gut microbiota. Their vast diversity plays an important role in maintenance of immune homeostasis. Gut microbes released several molecules against the antigenic pathogen and stabilize the self-tolerance of the immune system. Disturbance in the microbial community could be the major cause of loss of self-tolerance and thus results in autoimmunity. Literatures highlighted the number of diseases which are linked with the gut microbiota and its modulation. The various diseases are chronic obstructive pulmonary disease (COPD), ulcerative colitis, autism, type-1 diabetes, etc. Here, we present evidence mechanism links between gut microbiota dysbiosis and autoimmune diseases to understand the future treatment approaches based on gut microbiota for the autoimmune disease.KeywordsGut homeostasisAutoimmune diseasesGut dysbiosisImmune toleranceAutoimmunity
p>The hypothesis of this thesis was that 'the characteristics of early life exposure to dietary egg allergen determine infant atopic phenotype'.
Hen egg ovalbumin (OVA) was the dietary allergen investigated. Egg allergy is common in infancy and has implications for later inhalant sensitisation and respiratory allergic disease.
Women with a personal or partner history of atopy, were randomised to dietary egg exclusion or a normal healthy diet from 17-20 weeks of pregnancy till the end of breast-feeding. OVA was present in as many blood and breast milk samples of egg avoiding as control women. Atopic women had higher levels of serum OVA than non-atopic women, while atopic and egg avoiding women more often had OVA in breast milk, and in higher quantities, than non-atopic, egg avoiding women. These data suggest that dietary exclusion, particularly by atopic women, does NOT eliminate allergen exposure in early life. Antenatal OVA exposure, in the context of an egg-avoiding and atopic mother, was associated with a greater risk of an atopic phenotype at 6 months of age. Also, exposure via breast milk from an atopic mother suggested a greater risk of later atopy. Maternal serum OVA IgG concentration was shown to mark compliance to an egg exclusion diet and differences in cord concentrations were related to subsequent atopy. These data imply modulatory influences of maternal IgG and atopic environment over developing immune responses and raise the possibility that dietary exclusion as a primary allergy prevention strategy may have adverse consequences. Postnatally, differences in OVA IgG and IgG subclasses were identified for persistently egg sensitised children and elevated OVA IgG1 was associated with later asthma. This suggests that serum OVA IgG1 measurement might be used as an adjunct to skin testing and serum IgE measurement to predict allergic respiratory disease.
This work has provided insight into mechanisms that may modulate early life programming of atopy and has proposed factors for consideration in primary allergy prevention strategies. Furthermore, the potential for a serological measurement in infancy to predict long-lasting respiratory disease offers the prospect of early implementation of secondary allergy prevention measures.</p
Autoimmune diseases have induced a huge burden on individuals, families, economy, and the healthcare system. Considering their wide spectrum of manifestations and systemic involvement in some of the cases, different complications are associated with these diseases. On the other hand, since most of the current treatments systematically target the immune system, patients might experience a variety of adverse effects. Despite the progress in finding new targets and treatment strategies in animal studies, the development of treatments in clinical application is not seen to the same extent. One important explanation regarding this issue is the difference between the animal and human physiological systems. Moreover, the genetic and epigenetic differences between individuals potentially result in different disease manifestations even with the same underlying pathology besides different spectra of response to treatment. Therefore, finding proper treatment for autoimmune diseases by conducting translational studies that facilitate the application of current evidence from laboratory, animal studies, and preclinical findings in clinical practice is of interest. In this chapter, a brief review of currently available treatments for autoimmune diseases and existing challenges in this field are provided.
Objectives: Maintenance of immunological tolerance towards the semi-allogeneic fetus is important for successful pregnancy outcome. However, some women experience unexplained recurrent pregnancy losses (uRPL). It has been described that regulatory T-cell (Treg) percentages are decreased at the maternal-fetal interface (decidua) during miscarriage compared to elective termination pregnancies. We hypothesized that in women with uncomplicated term pregnancy after a previous history of uRPL a higher level of immune regulation is required at the fetal-maternal interface compared to women with uncomplicated pregnancy without any miscarriage in history.
Methods: We developed two antibody panels for in-depth immune profiling by mass cytometry using a total of 59 unique immune markers. The first panel focused on all immune lineages while the second had a focus on T cells, including intracellular markers. We analysed decidual immune cells isolated from placentas of 6 women with historic uRPL (cases) and 3 women without miscarriage history (controls).
Results: Analysis and visualization of the high-dimensional data was performed by applying HSNE and t-SNE for the case- and control groups combined. For this first analysis we focused on Tregs. In clustering analysis based on marker expression, FoxP3+ and FoxP3- Treg-like cells (CD25+CD127-CTLA-4+) clustered together. We observed a significantly higher Treg-like cell percentage in the decidua parietalis of cases compared to controls (2.35% vs. 0.18% within CD45+ cells, P=0.024). For both the decidua basalis and parietalis, 50% of cases had an increased Treg-like cell percentage compared to controls.
Conclusion: A proportion of women with an history of uRPL display increased Treg-like cell levels at the fetal-maternal interface, suggesting a possible compensatory role allowing for a successful pregnancy. We aim to further elucidate the immunological context of this potential regulatory compensation mechanism by analyzing the other immune cell types in the decidua.
In the majority of children with food allergy tolerance to food develops within the years. However in a number of patients the food allergy stays for life and patients are compelled to adhere an eliminative diet, during which there is a threat of the allergenic product occasional exposure constantly. Recent studies showed the efficacy of oral immunotherapy with different products in food allergy in children. In this article we reviewed the experience of oral allergen-specific immunotherapy with different foods.
Ischemic stroke triggers an inflammatory response that progresses for days to weeks and involves both tissue resident and peripheral immune cells. There is evidence that the acute inflammatory response contributes to the progression of ischemic brain injury, that ultimately exacerbates neurologic deficits. On the other hand, recent research points at a more multifaceted role of immune cells in brain ischemia, where immune cells participate in repair processes during the subacute and chronic stages. In addition, the interaction of the ischemic brain and the immune system is bidirectional, and while the peripheral immune system supplies immune cells that participate in the local inflammatory response, neural and humoral factors generated by the ischemic brain signal to the peripheral immune system leading to an acute systemic inflammatory response followed by immunosuppression and increased risk for nosocomial infections. Understanding these processes and the nature of immune cells involved during deleterious and reparatory phases of postischemic inflammation will be necessary to devise effective therapeutic strategies for human stroke. In this chapter, we review the features of postischemic inflammation, focusing on recent advances and insights on the potential molecular and cellular mechanisms by which such inflammation influences stroke pathophysiology. Furthermore, we examine the role of bidirectional signaling between the ischemic brain and the peripheral immune system in determining stroke outcome. Finally, we analyze the potential therapeutic implications of modulators of specific inflammatory targets from the perspective of near-future translational approaches.
A method for the preparation of smooth and spherical microspheres of the hydrophobic corn protein, zein, by a phase separation method, is reported in this thesis. Solubility studies on zein as well as amino acid and peptide composition analyses, together with studies on the formulation process were carried out to comprehend the mechanisms of zein microsphere formation. Microspheres were characterised in terms of guest molecule loading, size, charge, morphology, and composition. Ovalbumin, as a model protein/antigen guest, was incorporated into the zein microspheres during microsphere formation at different experimental loadings, and its in vitro release was examined in phosphate buffer at 37 °C. Microsphere degradation studies as a function of medium pH and in the presence of enzymes were also performed in vitro to understand the release of the guest molecule. Zein microspheres were found to be extremely resistant to degradation in the absence of enzymes, which was reflected in negligible release of ovalbumin. Finally, zein microspheres (both blank and ovalbumin-loaded) were administered orally, intramuscularly, rectally, and vaginally to investigate whether a zein microparticulate carrier system was capable to induce an enhanced immune response to the model antigen. In these studies, adjuvanticity of zein microspheres could not be proved due to the antigenicity of the zein protein.
Currently, more than 46 million adults in the United States suffer from some form of arthritis. Arthritis is also prevalent in other species, including canine and equine. Among all forms of arthritis, osteoarthritis (OA) is the most common form, afflicting nearly 27 million adults. The other common form of arthritis is rheumatoid arthritis. Among animal species, canine and equine are more prone to arthritis than other species. A large number of factors, such as aging, excessive exercise, obesity, genetic predisposition, immune disorders, poor nutrition, injury, and infection, can lead to OA. Presently, the use of nutraceuticals appears to be a good option to treat or manage OA, because they are taken orally, well-tolerated, and safe. However, some nutraceuticals exert adverse effects. This chapter describes, in brief, the characteristics of OA, pathophysiology of OA, and efficacy, safety, and toxicity of some commonly used nutraceuticals.
The immune system comprises two major arms: the innate immune system, which is activated following the detection of danger molecules by highly conserved receptors, and a second line of defense known as adaptive immunity, which provides an enormous anticipatory repertoire of antigen-specific effector cells and antibodies. The liver is an important immune organ, and liver-derived products can trigger the innate and adaptive immune system to initiate, mediate, regulate, and resolve systemic inflammation. The unique anatomy and microanatomy of the liver enables the generation of distinct immune responses. Generally, the immune responses elicited in the liver are skewed toward tolerance in response to the constant exposure of an enormous antigen load from the gut. An important mechanism leading to liver tolerance is antigen presentation by nonprofessional and/or immature hepatic antigen-presenting cells. Autoimmune diseases occur when a specific adaptive immune response is mounted against self-antigens. A combination of genetic and environmental factors plays a role in the pathogenesis of autoimmune disorders. The development of autoimmunity specifically in the liver, a tolerogenic organ, is a great paradox. The reasons for that are largely obscured currently and hopefully will be uncovered utilizing novel scientific methods in the near future.
Studies have shown that experimental allergic encephalomyelitis, a model for Multiple Sclerosis, can be inhibited by repeated administration of anti-inflammatory cytokines or cytokine neutralising agents. Given the short half-life of many biological agents, administration must be frequent to maintain a therapeutic effect. Gene therapy allows long-term in vivo delivery of these biological therapeutic agents. This study has investigated the use of viral and non-viral vectors administered systemically and locally to the central nervous system, to deliver the anti-inflammatory cytokines IL-4, IL-10 TGF-β and IFN-γ with the aim of ameliorating EAE. In addition, neutralisation of TNF using soluble TNF receptors and blockade of T-cell costimulation using CTLA4-Ig fusion proteins and anti-B7 antibodies were also studied. This study demonstrates that EAE can be successfully inhibited using gene delivered biological agents by a variety of vectors, either during the priming stage of disease or after disease onset. Local administration of the vector to the CNS increases the efficacy of therapeutic agents even at lower doses than used systemically, and may reduce the systemic side effects seen with standard high-dose protein therapy. The study of currently available vectors under the same conditions and in the same model allows a useful comparison and highlights both advantages and disadvantages of each method. This study provides data on a variety of successful approaches of gene delivery to treat an ongoing CNS autoimmune disease, which hopefully will provide a basis for using this method in humans.
Background and objectives
The possibility of manipulating the immune response in lambs to the gastrointestinal nematode Trichostrongylus colubriformis to reduce production losses associated with infection was investigated. In a series of four experiments attempts to immunise sheep via the mucosal route to modify the immune response and induce mucosal tolerance are outlined. Initially, a proof of concept study was conducted with lambs being injected with multiple doses of a somatic T. colubriformis antigen without an adjuvant in the rectal submucosa and subsequently challenged with T. colubriformis L3 larvae. This was followed by a dose response study comparing different antigen doses to identify the optimum dose of the nematode antigen for successful induction of mucosal tolerance. The final two studies were conducted to determine the larval stage specificity of the parasite antigen and the most suitable site of delivery required to stimulate mucosal tolerance.
Methods
In the proof of concept study lambs either received repeated injections in the rectal sub‐mucosa at 3 x weekly intervals with 15 µg of L3, 11 µg of L4 and 21 µg of immature adult (L5) somatic T. colubriformis antigens (ANT) or not (INF) prior to infection with T. colubriformis . In the dose rate study antigen dose rates of 100%, 50%, 10%, 1% or 0% of the antigen concentration used in the proof of concept study were compared while the larval stage study compared antigen from either L3, L4, L5 stages or combination of all (COMB) and the route of administration study compared antigen delivery into either the rectal submucosa (RE) or sub‐ cutaneous injection (SC).
Results
During infection lamb growth was improved by antigen treatment between days 21 and 42 in the proof of concept study (P=0.009), for groups 10%, 50% and 100% in the dose rate study (P<0.05 for all) and in RE in the route of administration study with no improvement observed in the larval stage study. No differences in faecal egg counts were observed (P>0.05 for all). Parasite specific IgA and IgE showed a dose response (the dose rate study), were not affected by larval stage (the larval stage study) and were greater in RE than SC (the route of administration study). IL‐4 production following lymphocyte stimulation was greatest in COMB ( the larval stage study) and RE (the route of administration study).
Conclusions
Although antigen treatment improved performance, this was inconsistent and appeared to stimulate immunity rather than induce tolerance. Combined larval stages were more efficient than individual stages and intra‐rectal administration was more effective than sub‐cutaneous.
In pregnancy, the semi-allogeneic fetus needs to be tolerated by the mother’s immune system. Regulatory T cells (Tregs) play a prominent role in this process. Novel technologies allow for in-depth phenotyping of previously unidentified immune cell subsets, which has resulted in the appreciation of a vast heterogeneity of Treg subsets. Similar to other immunological events, there appears to be great diversity within the Treg population during pregnancy, both at the maternal-fetal interface as in the peripheral blood. Different Treg subsets have distinct phenotypes and various ways of functioning. Furthermore, the frequency of individual Treg subsets varies throughout gestation and is altered in aberrant pregnancies. This suggests that distinct Treg subsets play a role at different time points of gestation and that their role in maintaining healthy pregnancy is crucial, as reflected for instance by their reduced frequency in women with recurrent pregnancy loss. Since pregnancy is essential for the existence of mankind, multiple immune regulatory mechanisms and cell types are likely at play to assure successful pregnancy. Therefore, it is important to understand the complete microenvironment of the decidua, preferably in the context of the whole immune cell repertoire of the pregnant woman. So far, most studies have focused on a single mechanism or cell type, which often is the FoxP3 positive regulatory T cell when studying immune regulation. In this review, we instead focus on the contribution of FoxP3 negative Treg subsets to the decidual microenvironment and their possible role in pregnancy complications. Their phenotype, function, and effect in pregnancy are discussed.
Introduction:
Oral tolerance is defined as the suppression of the immune response to antigens that have been previously administered orally. The purpose of inducing oral tolerance is to avoid using immunosuppressive drugs since, considering that they are not antigen-specific, they make the host more susceptible to acquire infections and develop neoplasms.
Objective:
To carry out a literature review on the most relevant theoretical references regarding oral tolerance induction in organ and tissue transplantation to prove that oral tolerance is a viable alternative therapy in transplant patients.
Materials and methods:
A literature review was conducted in the PubMed, MEDLINE, LILACS and Embase databases using the following search strategy: publication time: no limits; publication language: English and Spanish; type of studies: case-control studies, literature and systematic reviews; search terms: "T-Lymphocytes, Regulatory", "Autoimmunity", Immunosuppression", "Immune system" and "Immune Tolerance", and their equivalents in Spanish.
Results:
After the initial search was completed, 719 records were retrieved; however, only 99 addressed oral tolerance induction. Once duplicate records and articles without full-text access were removed, 75 studies were included for analysis.
Conclusions:
Oral administration of antigens is an effective way to induce immune tolerance in transplant patients (murine models) as it eliminates the adverse effects associated with immunosuppressive therapy, which currently is the standard therapy to treat these patients worldwide.
CDR3 of the functional rearranged T-cell receptor variable β region (TCR-Vβ) transcript was sequenced in order to demonstrate for the first time the identity between a long-term cultured T-cell line derived from a cutaneous T-cell lymphoma (CTCL) patient and the malignant T-cell clone present in the blood. The patient's peripheral blood lymphocyte-derived cultured T-cell line had a CD3+Vβ22+CD4+CD8+CD25−phenotype. It was named Pno and had been cultured for more than 1 year. Both fresh and long-term–cultured tumor cells proliferated highly in response to interleukin-7 (IL-7), and exogeneous IL-7 prevented Pno lymphocytes from apoptosis and maintained high levels of Bcl-2 expression. This unique malignant cloned lymphocyte line was further used to carry out functional studies. The results indicated that the CD3/TCR structures expressed by the Pno lymphocytes were functional because an immobilized anti-CD3 monoclonal antibody (mAb) or the combination of a soluble anti-CD3 mAb with submitogenic doses of phorbol 12 β-myristate 13 -acetate induced a proliferative response. Further, the CD2 and CD28 coreceptors were functional because they were able to induce a strong proliferative response upon their specific stimulation. Finally, the Pno T cell line had a Th3-type cytokine profile because it produced high amounts of the immunosuppressor cytokine tumor growth factor–β1 (TGF-β1). This high production of TGF-β1 may inhibit antitumor specific responses in CTCL.
Chronic graft versus host disease (cGVHD) is a major complication that can develop after bone marrow transplantation. It involves an immune-mediated attack by transplanted donor lymphocytes, and often results in inflammatory damage of host target organs. Immune hyporesponsiveness induced by oral antigen administration has been recently shown to prevent the development of cGVHD in a murine model. The aim of this study was to evaluate whether tolerance induction in bone marrow transplant (BMT) recipients after transplantation, toward their pretransplant antigens, can alleviate preexisting cGVHD in a mouse model. cGVHD was generated by infusing 2.5 × 107splenocytes from B10.D2 donor mice, to sublethally irradiated (6 Gy) BALB/c recipient mice, which differ by minor histocompatibility antigens. Transplantation resulted in cGVHD, with characteristic scleroderma-like cutaneous fibrosis, increased skin collagen content, decreased body weight, and hepatic and small bowel inflammation. Oral tolerance was induced by feeding recipient BALB/c mice with proteins extracted from BALB/c splenocytes for 11 days after B10.D2 splenocyte transplantation. Tolerance induction was evidenced by a significant reduction in mixed lymphocyte response of effector splenocytes from tolerant BALB/c mice transplanted with B10.D2 splenocytes against BALB/c target splenocytes. Oral tolerance decreased skin collagen deposits. Reduction of collagen 1(I) gene expression and skin collagen were shown by in situ hybridization and histochemistry, respectively. Liver and bowel biopsy specimens revealed less inflammation. Serum IL-10 levels were higher in tolerant mice than in controls, whereas IFNγ was significantly reduced. Oral tolerance of BMT recipients toward their pretransplant antigens after splenocyte transplantation down-regulated the immune attack by transplanted cells, thus ameliorating cGVHD.
Peanuts are a source of proteins and fats but they are also considered a harmful food for individuals who are allergic to them due to their ability to trigger severe and life-threatening reactions. Strict avoidance of peanuts is the most effective means to prevent the development of an allergic reaction. Physical or chemical strategies employing autoclaving can represent a valid alternative to produce a final food with a decreased allergenic power as in the case of peanuts. Thermal processing might induce protein modifications in foods and affect protein digestibility or absorption of nutrients across the intestinal mucosa. Besides, the type of processing could also alter food protein allergenicity thus influencing the interplay with the biological system at the gut level. In this paper, we investigated the influence of autoclaving based treatments on the proliferation of epithelial cells at the intestinal level. Extractable proteins of raw and autoclaved peanuts were analysed by SDS-PAGE and untargeted LC-high resolution-MS/MS to investigate the peptide composition. Our findings show that when raw peanuts were assayed on Caco 2 cell lines, an antiproliferative effect was observed. By contrast, peanuts subjected to hydration and autoclaving did not show an inhibition of proliferation on Caco-2 cells. In parallel, extensive fragmentation induced by autoclaving treatments on the original peanut proteins was also recorded by LC-MS/MS analysis with a consequent increase in the number of peptides detected. These results indicate that the processing applied to peanuts can have an influence on both the nutritional and allergological sides, and more investigations will be required on this issue to understand the alteration of inflammatory mediators induced by the treatment applied.
Background
Insulin is usually the first target of the autoimmune response in children developing type 1 diabetes and is therefore considered a promising candidate for an antigen-specific immunomodulation for the preventive treatment of the disease.
Study results
To date, several hundred of children worldwide have been treated with oral insulin in clinical studies, in various doses and without adverse reactions. Two large studies could not achieve a preventive effect with 7.5 mg oral insulin daily in β‑cell autoantibody-positive individuals overall, although delayed diabetes development was observed in subgroups. In a pilot study, 67.5 mg oral insulin daily induced a regulatory immune response against insulin.
Ongoing studies with high-dose oral insulin
The preventive effect of 67.5 mg oral insulin daily on diabetes development is currently under investigation: in β‑cell-autoantibody-positive children with an asymptomatic early stage of type 1 diabetes (Fr1da-Insulin-Intervention study) and in autoantibody-negative children with genetic risk for type 1 diabetes (POInT study [POInT: Primary Oral Insulin Trial]).
For many years, the intestine was one of the poor relations of the immunology world, being a realm inhabited mostly by specialists and those interested in unusual phenomena. However, this has changed dramatically in recent years with the realization of how important the microbiota is in shaping immune function throughout the body, and almost every major immunology institution now includes the intestine as an area of interest. One of the most important aspects of the intestinal immune system is how it discriminates carefully between harmless and harmful antigens, in particular, its ability to generate active tolerance to materials such as commensal bacteria and food proteins. This phenomenon has been recognized for more than 100 years, and it is essential for preventing inflammatory disease in the intestine, but its basis remains enigmatic. Here, I discuss the progress that has been made in understanding oral tolerance during my 40 years in the field and highlight the topics that will be the focus of future research.
Experimental autoimmune encephalomyelitis (EAE) in the Lewis rat is a self-limited inflammatory process localized to the central nervous system that is induced by the injection of myelin basic protein (MBP) in adjuvant. Oral administration of MBP suppresses EAE, and this suppression is mediated by CD8+ T cells that adoptively transfer protection and suppress both in vitro and in vivo by the release of transforming growth factor (TGF) beta after antigen-specific triggering. Furthermore, oral tolerance to MBP is enhanced by the concomitant oral administration of lipopolysaccharide (LPS). The present study was undertaken to determine whether the disease course in EAE and its suppression by oral tolerization to MBP is associated with distinct patterns of cytokine expression in the target organ. Detailed immunohistology of the brain was performed at the peak of clinical disease (day 14 after immunization) and after recovery (day 18) in control (ovalbumin [OVA]-fed), MBP-fed, and MBP plus LPS-fed animals. Brains from OVA-fed animals at the peak of disease showed perivascular infiltration with activated mononuclear cells which secreted the inflammatory cytokines interleukins (IL) 1, 2, 6, 8, TNF-alpha, and interferon gamma. The inhibitory cytokines TGF-beta and IL-4, and prostaglandin E2 (PGE2) were absent. In MBP orally tolerized animals there was a marked reduction of the perivascular infiltrate and downregulation of all inflammatory cytokines. In addition, there was upregulation of the inhibitory cytokine TGF-beta. In MBP plus LPS orally tolerized animals, in addition to upregulation of TGF-beta and reduction of inflammatory cytokines, there was enhanced expression of IL-4 and PGE2, presumably secondary to activation of an additional population of immunoregulatory cells. In OVA-fed animals that had recovered (day 18), staining for inflammatory cytokines diminished, and there was the appearance of TGF-beta and IL-4. These results suggest that suppression of EAE, either induced by oral tolerization or that which occurs during natural recovery is related to the secretion of inhibitory cytokines or factors that actively suppress the inflammatory process in the target organ.
Cellular immune hyporesponsiveness can be induced by the presentation of soluble protein an-tigens to mucosal surfaces. Most studies of mucosa-mediated tolerance have used the oral route of antigen delivery and few have examined autoantigens in natural models of autoimmune disease. Insulin is an autoantigen in humans and nonobese diabetic (NOD) mice with insulin-dependent diabetes mellitus (IDDM). When we administered insulin aerosol to NOD mice after the onset of subclinical disease, pancreatic islet pathology and diabetes incidence were both significantly reduced. Insulin-treated mice had increased circulating antibodies to insulin, absent splenocyte proliferation to the major epitope, insulin B chain amino acids 9–23, which was associated with increased IL-4 and particularly IL-10 secretion, and reduced proliferation to glutamic acid decarboxylase, another islet autoantigen. The ability of splenocytes from insulin treated mice to suppress the adoptive transfer of diabetes to nondiabetic mice by T cells of diabetic mice was shown to be caused by small numbers of CD8 T cells. These findings reveal a novel mechanism for suppressing cell-mediated autoimmune disease. Induction of regulatory CD8 T cells by aerosol insulin is a therapeutic strategy with implications for the prevention of human IDDM.
We established conditions for inducing antigen-specific tolerance in Th2 lymphocytes by means of oral tolerance. Mice were continuously exposed to ovalbumin in their drinking water for a minimal
period of 20 days and then immunized against antigen in either complete Freund's adjuvant or Al(OH)3. This feeding regimen tolerized both Th2 and Th1 responses as shown by diminished proliferation, cytokine secretion (IL-4, IL-2 and IFN-γ) and specific cytokine mRNA expression
(IL-4, IL-2 and IFN-γ) in vitro, as well as by absence of specific antibody production (lgG1, lgG2a, gG2b and IgE) in vivo. Conditions for generating Th2 lymphocyte tolerance were different from those required to generate tolerance in Th1 lymphocytes: these included extended, continuous exposure to high dosages of antigen, rather than a single or intermittent
feeding regimen which was sufficient to induce tolerance in Th1 lymphocytes. These findings suggest that continuous oral exposure to a tolerogen may be a biologically relevant strategy
to tolerize both Th1- and Th-dependent responses, and extend the potential clinical use of oral tolerance to ailments mediated by Th2 lymphocytes.
We have developed a new model of autoimmune arthritis in DBA/1 mice by feeding chick type II collagen (CII) for 2–3 week intervals
over a 15 week period. Clinically evident arthritis occurred in 87/10 mice receiving native CII (nCII; 100 μg/mouse) alone
at 9–l13 weeks. Arthritis was aggravated by the further ingestion of CII, while remission occurred after withdrawal of the
CII. Heat-denatured CII (dCII; 200 μg/mouse) was also arthritogenic if co-administered with ovoinhibitor (OVI; 2 mg/mouse),
a proteinase inhibitor. Co-oral administration of lipopolysaccharide (LPS; 10μg/mouse) with CII enhanced the antibody production
and T-cell responses to CII, and induced a more chronic arthritis that progressed spontaneously without further administration
of CII or LPS. Long-term oral administration of LPS alone also induced a mild arthritis characterized by destruction of bone
rather than cartilage. These observations suggest that abnormal gastrointestinal absorption of dietary mimic antigens and
intestinal bacterial toxins can potentially disrupt self-tolerance mechanisms, thereby precipitating or exacerbating autoimmune
disease in genetically susceptible individuals.
Summary In previous studies we showed that a chronic colitis associated with a Thl T cell response can be induced by the rectal administration of the haptenizing reagent 2,4,6-trinitrobenzene sul- fonic acid (TNBS). We report here that oral administration of haptenized colonic proteins (HCP) before rectal administration of TNBS effectively suppresses the ability of the latter to in- duce colitis. This suppression (oral tolerance) appears to be due to the generation ofmucosal T cells producing TGF-(3 and Th2-type cytokines after oral HCP administration. Peyer's patch and lamina propria CD4+ T cells from HCP-fed animals stimulated with anti-CD3/anti- CD28 had a 5-10-fold increase in their production of TGF-(3 and secreted increased amounts oflL-4 and IL-10 but lower levels oflFN-~ in comparison to T cells from ovalbumin-fed con- trol animals. In addition, the colons of HCP-fed mice showed strikingly increased TGF-(3 but decreased IL-12 expression by immunohistochemicat studies and isolated mononuclear cells from HCP-fed animals secreted less IL-12 heterodimer. Finally, and most importantly, the sup- pressive effect of orally administered HCP was abrogated by the concomitant systemic adminis- tration ofanti-TGF-(3 or rlL-12 suggesting a reciprocal relationship between IL-12 and TGF-(3 on tolerance induction in TNBS-induced colitis. In parallel studies we demonstrated that TNBS-induced colitis can be transferred to naive recipient animals with purified CD4+ T cells from the colon of TNBS-treated animals and that such animals develop lethal pancolitis when exposed to very low doses of TNBS. Feeding of HCP suppressed this sensitivity to TNBS, in- dicating that oral feeding can suppress the response of pre-committed T cells in vivo. These studies suggest for the first time that TGF-(3 production can abrogate experimental granuloma- tous colitis even after such colitis is established, and thus, that regulation of TGF-(3 levels may have relevance to the treatment of human inflammatory bowel disease.
Mucosally induced immunological tolerance is an attractive strategy for preventing or treating illnesses resulting from untoward inflammatory immune reactions against self- or non-self-antigens. Oral administration of relevant autoantigens and allergens has been reported to delay or suppress onset of clinical disease in a number of experimental autoimmune and allergic disorders. However, the approach often requires repeated feeding of large amounts of tolerogens over long periods and is only partly effective in animals already systemically sensitized to the ingested antigen such as in animals already harboring autoreactive T cells, and thus presumably also in humans with an autoimmune disease. We have recently shown that oral administration of microgram amounts of antigen coupled to cholera toxin B subunit (CTB), can effectively suppress systemic T cell reactivity in naive as well as in immune animals. We now report that feeding small amounts (2–20 μg) of human insulin conjugated to CTB can effectively suppress beta cell destruction and clinical diabetes in adult nonobese diabetic (NOD) mice. The protective effect could be transferred by T cells from CTB-insulin-treated animals and was associated with reduced lesions of insulitis. Furthermore, adoptive co-transfer experiments involving injection of Thy-1,2 recipients with diabetogenic T cells from syngeneic mice and T cells from congenic Thy-1,1 mice fed with CTB-insulin demonstrated a selective recruitment of Thy-1,1 donor cells in the peripancreatic lymph nodes concomitant with reduced islet cell infiltration. These results suggest that protection against autoimmune diabetes can be achieved by feeding minute amounts of a pancreas islet cell autoantigen linked to CTB and appears to involve the selective migration and retention of protective T cells into lymphoid tissues draining the site of organ injury.
Experimental autoimmune encephalomyelitis (EAE) in the Lewis rat is a self-limited inflammatory process localized to the central nervous system that is induced by the injection of myelin basic protein (MBP) in adjuvant. Oral administration of MBP suppresses EAE, and this suppression is mediated by CD8+ T cells that adoptively transfer protection and suppress both in vitro and in vivo by the release of transforming growth factor (TGF) beta after antigen-specific triggering. Furthermore, oral tolerance to MBP is enhanced by the concomitant oral administration of lipopolysaccharide (LPS). The present study was undertaken to determine whether the disease course in EAE and its suppression by oral tolerization to MBP is associated with distinct patterns of cytokine expression in the target organ. Detailed immunohistology of the brain was performed at the peak of clinical disease (day 14 after immunization) and after recovery (day 18) in control (ovalbumin [OVA]-fed), MBP-fed, and MBP plus LPS-fed animals. Brains from OVA-fed animals at the peak of disease showed perivascular infiltration with activated mononuclear cells which secreted the inflammatory cytokines interleukins (IL) 1, 2, 6, 8, TNF-alpha, and interferon gamma. The inhibitory cytokines TGF-beta and IL-4, and prostaglandin E2 (PGE2) were absent. In MBP orally tolerized animals there was a marked reduction of the perivascular infiltrate and downregulation of all inflammatory cytokines. In addition, there was upregulation of the inhibitory cytokine TGF-beta. In MBP plus LPS orally tolerized animals, in addition to upregulation of TGF-beta and reduction of inflammatory cytokines, there was enhanced expression of IL-4 and PGE2, presumably secondary to activation of an additional population of immunoregulatory cells. In OVA-fed animals that had recovered (day 18), staining for inflammatory cytokines diminished, and there was the appearance of TGF-beta and IL-4. These results suggest that suppression of EAE, either induced by oral tolerization or that which occurs during natural recovery is related to the secretion of inhibitory cytokines or factors that actively suppress the inflammatory process in the target organ.
Suppression of experimental autoimmune encephalomyelitis (EAE) in Lewis rats by the oral administration of myelin basic protein (MBP) is mediated by CD8+ T cells that can be isolated from the spleens of MBP-fed animals. These cells adoptively transfer protection to naive animals subsequently immunized with MBP and complete Freund's adjuvant (CFA) and suppress in vitro MBP proliferative responses. Using a transwell system in which the modulator spleen cells from MBP-fed animals are separated by a semipermeable membrane from responder cells, MBP, or OVA-specific T cell lines, we have found that cell contact is not required for in vitro suppression to occur. In vitro suppression is dependent, however, upon antigen-specific triggering of modulator T cells. Once antigen-specific triggering occurs, suppression across the transwell is mediated by an antigen-nonspecific soluble factor that equally suppresses an MBP line or an ovalbumin (OVA) line. This phenomenon of antigen-driven bystander suppression was also demonstrated in vivo. Specifically, Lewis rats fed OVA which were then immunized with MBP/CFA plus OVA given separately subcutaneously were protected from EAE. Animals fed OVA and then immunized with MBP/CFA without OVA given subcutaneously were not protected. The protective effect of feeding OVA could be adoptively transferred by CD8+ T cells from OVA-fed animals into MBP/CFA plus OVA-injected animals. Feeding bovine serum albumin (BSA) or keyhole limpet hemocyanin did not suppress EAE in animals immunized with MBP/CFA plus OVA. EAE was suppressed, however, if BSA was fed and animals then immunized with MBP/CFA plus BSA given subcutaneously. Antigen-driven bystander suppression appears to be an important mechanism by which antigen-driven peripheral tolerance after oral administration of antigen is mediated, and presumably occurs in the microenvironment accounting for the antigen specificity of suppression generated by oral tolerization to antigens.
We investigated the capacity of murine T lymphocytes, isolated from various lymphoid organs of normal or antigen-primed donors, to produce IL-2 or IL-4 after activation with anti-CD3 or specific antigen. Our results established that T cells resident within lymphoid organs being drained by nonmucosal tissue sites (e.g., axillary, inguinal, brachial lymph nodes, or spleen) produced IL-2 as the predominant T cell growth factor (TCGF) after activation. Conversely, activated T cells from lymphoid organs being drained by mucosal tissues (Peyer's patches, and cervical, periaortic, and parathymic lymph nodes) produced IL-4 as the major species of TCGF. Analysis of the lymphoid tissues obtained from adoptive recipients of antigen-primed lymphocytes provided by syngeneic donors provided evidence that direct influences were being exerted on T cells during their residence within defined lymphoid compartments. These lymphoid tissue influences appeared to be responsible for altering the potential of resident T cells to produce distinct species of TCGF. Steroid hormones, known transcriptional enhancers and repressors of specific cellular genes, were implicated in the controlling mechanisms over TCGF production. Glucocorticoids (GCs) were found to exert a systemic effect on all recirculating T cells, evidenced by a marked dominance in IL-4 production by T cells obtained from all lymphoid organs of GC-treated mice, or after a direct exposure of normal lymphoid cells to GCs in vitro before cellular activation with T cell mitogens. Further, the androgen steroid DHEA appeared to be responsible for providing an epigenetic influence to T cells trafficking through peripheral lymphoid organs. This steroid influence resulted in an enhanced potential for IL-2 secretion after activation. Anatomic compartmentalization of the DHEA-facilitated influence appears to be mediated by differential levels of DHEA-sulfatase in lymphoid tissues. DHEA-sulfatase is an enzyme capable of converting DHEA-sulfate (inactive) to the active hormone DHEA. We find very high activities of this enzyme isolated in murine macrophages. The implications of our findings to immunobiology are very great, and indicate that T cells, while clonally restricted for antigen peptide recognition, also appear to exhibit an extreme flexibility with regards to the species of lymphokines they produce after activation. Regulation of this highly conservative mechanism appears to be partially, if not exclusively, controlled by cellular influences being exerted by distinct species of steroid hormones, supplied in an endocrine or a paracrine manner where they mediate either systemic or tissue-localized influences, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)
The role of murine epithelial cells from small intestine (IEC) in orally induced immune unresponsiveness has been investigated. Murine IEC were shown to express a relatively low level of class II MHC Ag and were incapable of presenting BSA to Ag-specific Th cells. IFN-gamma treatment of the animals dramatically enhanced expression of class II MHC Ag by IEC enabling Ag presentation to Th cells. Ag-presenting function of IEC after IFN-gamma treatment was not affected by IEC panning on anti-IgG or anti-MAC-1-coated plates but it was blocked by a class II MHC-specific mAb. IFN-gamma-treated mice were responsive to parenteral BSA challenge after oral administration of the Ag, whereas untreated mice were immunosuppressed. Our data support the concept that poor expression of class II MHC Ag on IEC contributes to immunosuppression induced by oral administration of a protein Ag.
We report that experimental autoimmune encephalomyelitis, a T cell-mediated autoimmune disease studied as a model for multiple sclerosis, can be suppressed in Lewis rats by the oral administration of myelin basic protein (MBP). Both the clinical and histopathologic manifestations of the disease were suppressed in a dose-dependent manner. In addition, proliferative responses to MBP and, to a lesser extent, serum levels of anti-MBP antibody were suppressed by feeding MBP. Suppression of clinical and histologic disease was observed whether animals were fed MBP before or after disease induction, although suppression was more complete when rats were fed before immunization. Disease was also suppressed by the oral administration of either encephalitogenic or nonencephalitogenic fragments and decapeptides of the MBP molecule, with more complete suppression observed when nonencephalitogenic fragments were fed, suggesting that suppressor determinants exist in the MBP molecule distinct from the encephalitogenic region. The oral administration of a non-disease-inducing portion of an autoantigen represents an antigen-specific method by which an experimental autoimmune disease can be immunoregulated.
Nickel (Ni) is the most frequent cause of allergic contact dermatitis, mainly in female patients. In animals, oral tolerance to Ni sensitization can be obtained by feeding with Ni sulfate (NiSO4). The aim of the present study was to compare the specific proliferative responses and frequencies of Ni-responding T cells from peripheral blood of patients before and after a protocol of Ni ingestion. Ten patients with chronic disseminated eczema and patch test-proved contact allergy to Ni gave informed consent for the study. All were nonpregnant female patients between the ages of 21 and 40 years (mean 35 years). They ingested 22.4 mg NiSO4 (5 mg Ni) once a week for 8 weeks. All patients experienced an exacerbation of pruritus 12 to 24 hours after ingesting the first capsules, resolving within 24 hours. Two patients had a major flare-up of their eczema. Eight patients completed the whole study, and presented a progressive improvement of their cutaneous lesions. In these patients, the sums of individual epicutaneous test scores were decreased after 8 weeks (p < 0.02). Peripheral blood lymphocytes (PBL) were isolated and stored before and after Ni ingestion. Proliferation assays were performed with 5 x 10-5 mol/l NiSO4. Stimulation indexes were decreased after 8 weeks when compared to preingestion PBL (p < 0.02). A limiting dilution assay was developed to quantify Ni-specific T cells from peripheral blood. The frequencies of responding T-cells were decreased after 8 weeks (1/49 061 to 1/2 517 920), when compared to preingestion PBL (1/14 547 to 1/128 682). Our results confirm that oral hyposensitization may decrease the degree of contact hypersensitivity as measured by epicutaneous tests and induce clinical improvement of cutaneous lesions. In addition, oral nickel intake can decrease Ni-specific proliferations and the numbers of Ni-responding T cells in peripheral blood. These data open a new area of investigation for the treatment of Ni allergy.
We investigated the capacity of murine T lymphocytes, isolated from various lymphoid organs of normal or antigen-primed donors, to produce IL-2 or IL-4 after activation with anti-CD3 or specific antigen. Our results established that T cells resident within lymphoid organs being drained by nonmucosal tissue sites (e.g., axillary, inguinal, brachial lymph nodes, or spleen) produced IL-2 as the predominant T cell growth factor (TCGF) after activation. Conversely, activated T cells from lymphoid organs being drained by mucosal tissues (Peyer's patches, and cervical, periaortic, and parathymic lymph nodes) produced IL-4 as the major species of TCGF. Analysis of the lymphoid tissues obtained from adoptive recipients of antigen-primed lymphocytes provided by syngeneic donors provided evidence that direct influences were being exerted on T cells during their residence within defined lymphoid compartments. These lymphoid tissue influences appeared to be responsible for altering the potential of resident T cells to produce distinct species of TCGF. Steroid hormones, known transcriptional enhancers and repressors of specific cellular genes, were implicated in the controlling mechanisms over TCGF production. Glucocorticoids (GCs) were found to exert a systemic effect on all recirculating T cells, evidenced by a marked dominance in IL-4 production by T cells obtained from all lymphoid organs of GC-treated mice, or after a direct exposure of normal lymphoid cells to GCs in vitro before cellular activation with T cell mitogens. Further, the androgen steroid DHEA appeared to be responsible for providing an epigenetic influence to T cells trafficking through peripheral lymphoid organs. This steroid influence resulted in an enhanced potential for IL-2 secretion after activation. Anatomic compartmentalization of the DHEA-facilitated influence appears to be mediated by differential levels of DHEA-sulfatase in lymphoid tissues. DHEA-sulfatase is an enzyme capable of converting DHEA-sulfate (inactive) to the active hormone DHEA. We find very high activities of this enzyme isolated in murine macrophages. The implications of our findings to immunobiology are very great, and indicate that T cells, while clonally restricted for antigen peptide recognition, also appear to exhibit an extreme flexibility with regards to the species of lymphokines they produce after activation. Regulation of this highly conservative mechanism appears to be partially, if not exclusively, controlled by cellular influences being exerted by distinct species of steroid hormones, supplied in an endocrine or a paracrine manner where they mediate either systemic or tissue-localized influences, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)
We previously demonstrated that a spontaneous Th1 response against glutamate decarboxylase (GAD65) arises in NOD mice at four weeks in age and subsequently T cell autoimmunity spreads both intramolecularly and intermolecularly. Induction of passive tolerance to GAD65, through inactivation of reactive T cells before the onset of autoimmunity, prevented determinant spreading and the development of insulin-dependent diabetes mellitus (IDDM). Here, we examined whether an alternative strategy, designed to induce active tolerance via the engagement of Th2 immune responses to GAD65, before the spontaneous onset of autoimmunity, could inhibit the cascade of Th1 responses that lead to IDDM. We observed that a single intranasal administration of GAD65 peptides to 2-3-wk-old NOD mice induced high levels of IgG1 antibodies to GAD65. GAD65 peptide treated mice displayed greatly reduced IFN gamma responses and increased IL-5 responses to GAD65, confirming the diversion of the spontaneous GAD65 Th1 response toward a Th2 phenotype. Consistent with the induction of an active tolerance mechanism, splenic CD4+ (but not CD8+) T cells from GAD65 peptide-treated mice, inhibited the adoptive transfer of IDDM to NOD-scid/scid mice. This active mechanism not only inhibited the development of proliferative T cell responses to GAD65, it also limited the expansion of autoreactive T cell responses to other beta cell antigens (i.e., determinant spreading). Finally, GAD65 peptide treatment reduced insulitis and long-term IDDM incidence. Collectively, these data suggest that the nasal administration of GAD65 peptides induces a Th2 cell response that inhibits the spontaneous development of autoreactive Th1 responses and the progression of beta cell autoimmunity in NOD mice.
1 Department of Microbiology and Immunology Northwestern University Medical School 303 Chicago Avenue, 6-721 Tarry Building Chicago, Illinois 60611
Mucosal tissues of mice are enriched in T cells that express the gamma/delta T cell receptor. Since the function of these cells remains unclear, we have compared mucosal immune responses in gamma/delta T cell receptor-deficient (TCRdelta-/-) mice versus control mice of the same genetic background. The frequency of intestinal immunoglobulin (Ig) A plasma cells as well as IgA levels in serum, bile, saliva, and fecal samples were markedly reduced in TCRdelta-/- mice. The TCRdelta-/- mice produced much lower levels of IgA antibodies when immunized orally with a vaccine of tetanus toxoid plus cholera toxin as adjuvant. Conversely, the antigen-specific IgM and IgG antibody responses were comparable to orally immunized control mice. Direct assessment of the cells forming antibodies against the tetanus toxoid and cholera toxin antigens indicated that significantly lower numbers of IgA antibody-producing cells were present in the intestinal lamina propria and Peyer's patches of TCRdelta-/- mice compared with the orally immunized control mice. The selective reduction of IgA responses to ingested antigens in the absence of gamma/delta T cells suggests a specialized role for gamma/delta cells in mucosal immunity.
A T helper type 1 (Th1)-mediated colitis with similarities to inflammatory bowel disease in humans developed in severe combined immunodeficiency mice reconstituted with CD45RB(high) CD4+ splenic T cells and could be prevented by cotransfer of CD45RB(low) CD4+ T cells. Inhibition of this Th1 response by the CD45RB(low) T cell population could be reversed in vivo by an anti-transforming growth factor (TGF) beta antibody. Interleukin (IL) 4 was not required for either the differentiation of function of protective cells as CD45RB(low) CD4+ cells from IL-4-deficient mice were fully effective. These results identify a subpopulation of peripheral CD4+ cells and TGF-beta as critical components of the natural immune regulatory mechanism, which prevents the development of pathogenic Th1 responses in the gut, and suggests that this immunoregulatory population is distinct from Th2 cells.
In previous studies we showed that a chronic colitis associated with a Th1 T cell response can be induced by the rectal administration of the haptenizing reagent 2,4,6-trinitrobenzene sulfonic acid (TNBS). We report here that oral administration of haptenized colonic proteins (HCP) before rectal administration of TNBS effectively suppresses the ability of the latter to induce colitis. This suppression (oral tolerance) appears to be due to the generation of mucosal T cells producing TGF-beta and Th2-type cytokines after oral HCP administration. Peyer's patch and lamina propria CD4+ T cells from HCP-fed animals stimulated with anti-CD3/anti-CD28 had a 5-10-fold increase in their production of TGF-beta and secreted increased amounts of IL-4 and IL-10 but lower levels of IFN-gamma in comparison to T cells from ovalbumin-fed control animals. In addition, the colons of HCP-fed mice showed strikingly increased TGF-beta but decreased IL-12 expression by immunohistochemical studies and isolated mononuclear cells from HCP-fed animals secreted less IL-12 heterodimer. Finally, and most importantly, the suppressive effect of orally administered HCP was abrogated by the concomitant systemic administration of anti-TGF-beta or rIL-12 suggesting a reciprocal relationship between IL-12 and TGF-beta on tolerance induction in TNBS-induced colitis. In parallel studies we demonstrated that TNBS-induced colitis can be transferred to naive recipient animals with purified CD4+ T cells from the colon of TNBS-treated animals and that such animals develop lethal pancolitis when exposed to very low doses of TNBS. Feeding of HCP suppressed this sensitivity to TNBS, indicating that oral feeding can suppress the response of pre-committed T cells in vivo. These studies suggest for the first time that TGF-beta production can abrogate experimental granulomatous colitis even after such colitis is established, and thus, that regulation of TGF-beta levels may have relevance to the treatment of human inflammatory bowel disease.
Equal numbers of CD4+ T cells recognizing myelin basic protein (MBP) and proteolipid protein (PLP) are found in the circulation of normal individuals and multiple sclerosis (MS) patients. We hypothesized that if myelin-reactive T cells are critical for the pathogenesis of MS, they would exist in a different state of activation as compared with myelin-reactive T cells cloned from the blood of normal individuals. This was investigated in a total of 62 subjects with definitive MS. While there were no differences in the frequencies of MBP- and PLP-reactive T cells after primary antigen stimulation, the frequency of MBP or PLP but not tetanus toxoid-reactive T cells generated after primary recombinant interleukin (rIL-2) stimulation was significantly higher in MS patients as compared with control individuals. Primary rIL-2-stimulated MBP-reactive T cell lines were CD4+ and recognized MBP epitopes 84-102 and 143-168 similar to MBP-reactive T cell lines generated with primary MBP stimulation. In the cerebrospinal fluid (CSF) of MS patients, MBP-reactive T cells generated with primary rIL-2 stimulation accounted for 7% of the IL-2-responsive cells, greater than 10-fold higher than paired blood samples, and these T cells also selectively recognized MBP peptides 84-102 and 143-168. In striking contrast, MBP-reactive T cells were not detected in CSF obtained from patients with other neurologic diseases. These results provide definitive in vitro evidence of an absolute difference in the activation state of myelin-reactive T cells in the central nervous system of patients with MS and provide evidence of a pathogenic role of autoreactive T cells in the disease.
Hypersensitivity reactions to food proteins are rare, probably because the intestinal immune system has evolved efficient means of preventing such responses. In this article Allan Mowat reviews the mechanisms underlying the induction of immunological tolerance after feeding proteins and suggests how a breakdown in oral tolerance may lead to potentially harmful hypersensitivity in the intestine.
The present study examined the potential of orally induced tolerance for preventing immunological rejection of corneal allografts. Orthotopic corneal allografts were transplanted from either C3H (MHC + multiple minor H-mismatched) or NZB (multiple minor H-mismatched only) donors to CBGF(1) recipients on day 0. Tissue cultured corneal epithelial and endothelial cells from relevant donor strains were administered orally from day -14 to day -4 on a daily basis. The incidence of graft rejection, graft mean survival time (MST), and alloimmune responses, and the antigen specificity of induced tolerance were studied. Oral immunization induced a remarkable tolerance such that only 55% of the orally immunized hosts rejected their fully allogeneic corneal grafts (MST = 43 days) compared with 100% rejection (MST = 18 days) in normal controls. Likewise, rejection of MHC-matched, multiple minor H-mismatched corneal grafts fell from 80% in untreated controls to 36% in orally immunized hosts. Oral immunization was effective in desensitizing previously immunized hosts. Rejection of MHC-matched, multiple H minor-mismatched corneal allografts fell from 93% in preimmune, unfed hosts to 36% in preimmune, orally tolerized mice. Thus, oral immunization is a safe and effective method for desensitizing high-risk, preimmune hosts and promoting corneal allograft survival.
Oral administration of nicotinic acetylcholine receptor (AChR) to Lewis rats prior to myasthenogenic immunization with Torpedo AChR + complete Freund's adjuvant (CFA) results in the prevention of experimental autoimmune myasthenia gravis (EAMG) and the suppression of AChR-specific B cell responses and counteracts the development of AChR-reactive interferon-γ (IFN-γ) secreting T cells. To study the involvement of the T helper type 1 (Th1) cell-related lymphokine IFN-γ, the Th2 cell-related interleukin-4 (IL-4), and transforming growth factor β (TGF-β) that suppresses the synthesis of IFN-γ and IL-4, we used in situ hybridization with complementary DNA oligonucleotide probes to enumerate mononuclear cells (MNC) expressing mRNA for the cytokines IFN-γ IL-4, and TGF-β. Upon in vivo recognition of AChR, popliteal, inguinal, and mesenteric lymph nodes, spleen and thymus of rats with EAMG contained higher levels of IFN-γ, IL-4, and TGF-β mRNA-expressing cells compared to CFA-injected control rats, implicating the involvement in EAMG of AChR-reactive Th1 and Th2 cells in parallel. TGF-β was also upregulated in EAMG. Oral tolerance to EAMG was characterized by suppression of the levels of MNC expressing IFN-γ and IL-4, but augmentation of cells expressing TGF-β. The results suggest that IFN-γ, IL-4, and TGF-β are involved in the development of EAMG, and that TGF-β is important in the induction of oral tolerance to EAMG.
Experimental autoimmune uveitis (EAU) in Lewis rats is a T-cell dependent disease, which is induced by immunization with retinal S-Antigen (S-Ag) or its unveitogenic peptides. We have recently reported that the oral administration of S-Ag prior to the uveitogenic challenge results in suppression of the disease and of the cellular responses. We examined the effect of oral administration of a recently described uveitogenic peptide (Peptide 35) on the development of EAU induced with the whole protein. The severity of inflammation and retinal destruction, as well as the cellular proliferative responses, were suppressed compared to controls fed with Keyhole Limpet Hemocyanin (KLH). In addition, the levels of antigenspecific antibody isotypes in the serum of gavaged rats were determined. Although (IgA) levels were reduced in rats gavaged with S-Ag or the uveitogenic peptide, IgM and IgG levels were unaltered. Thus, the oral administration of Peptide 35 approaches the state of unresponsiveness seen in S-Ag feeding. In addition, the unresponsiveness can be demonstrated on both the cellular and humoral level.
S-Antigen, a photoreceptor cell protein, is highly efficient in inducing experimental autoimmune uveoretinitis (EAU), a severe inflammation of the uveal tract and retina of the eye. S-Antigen and six synthetic peptides, all of which correspond to known T-cell or B-cell recognition sites, were tested for their ability to induce oral tolerance to EAU in LEW rats. Feeding three 1-mg doses of native S-Antigen or three doses of one synthetic peptide, designated BSA(343-362) (200 micrograms per dose), reduced the incidence and severity of EAU induced by immunization with 50 micrograms of S-Antigen. Another peptide, BSA(270-289), was able to inhibit EAU only when a low dose (10 micrograms) of the uveitogenic S-Antigen was used to induce EAU. Animals which received 200 micrograms doses of four other immunologically active peptides, BSA(31-51), BSA(143-162), BSA(303-327) and BSA(333-352), were not significantly protected. Furthermore, animals fed BSA(343-362) were significantly less susceptible to EAU induced by adoptive transfer (tEAU) of the uveitogenic R9 T-cell lines. Con A-activated lymphocytes purified from spleens of rats fed peptide BSA(343-362) transferred partial resistance to tEAU induced by adoptive transfer of R9 line cells. The resistance of orally tolerized rats to induction of EAU by adoptive transfer of an activated, pathogenic T-cell line, and the ability of lymphocytes from orally-tolerized animals to transfer resistance to tEAU shows that effector mechanisms can be inhibited by oral feeding as well as the afferent mechanisms reported here and elsewhere. Circulating levels of IgG specific for S-Antigen were not affected by feeding any of the peptides.
Immunization with myelin basic protein (MBP) induces experimental allergic encephalomyelitis (EAE), a prototype of CD4+ T-cell mediated autoimmune disease. In rodents, MBP-reactive T-cell clones are specific for a single, dominant determinant on MBP and use a highly restricted number of T-cell receptor genes. Accordingly, EAE has been prevented by various receptor-specific treatments, suggesting similar strategies may be useful for therapy of human autoimmune disease. Here we report that in (SJL x B10.PL)F1 mice, immune dominance of a single determinant, MBP:Ac1-11, is confined to the inductive phase of EAE. In mice with chronic EAE, several additional determinants of MBP in peptides 35-47, 81-100 and 121-140 recall proliferative responses. Most importantly, reactivity to the latter determinants was also detected after induction of EAE with MBP peptide Ac1-11 alone; this demonstrates priming by endogenous MBP determinants. Thus, determinants of MBP that are cryptic after primary immunization can become immunogenic in the course of EAE. Diversification of the autoreactive T-cell repertoire due to 'determinant spreading' has major implications for the pathogenesis of, and the therapeutic approach to, T-cell driven autoimmune disease.
We have been investigating the suppression of experimental autoimmune encephalomyelitis (EAE) by oral tolerization to autoantigens. In the present study the tolerizing effect of orally administered myelin basic protein (MBP) from different species was examined in the Lewis rat, Hartley guinea pig, and SJL/J mouse model of EAE. Animals were fed guinea pig, rat, bovine, human or mouse-MBP and then immunized with the homologous species of MBP or myelin: Lewis rats were immunized with rat MBP, Hartley guinea pigs with guinea pig-MBP, and SJL/J mice with mouse myelin. Clinical expression of EAE and delayed-type hypersensitivity (DTH) responses to MBP were assessed. In each species, suppression of disease and DTH responses were most pronounced by tolerization with the homologous species of MBP. In addition, cross-species tolerization was observed in each species and in general was less suppressive than homologous MBP although in some instances MBP from a heterologous species was as effective as tolerization with the homologous species. We also studied guinea pig-MBP induced EAE in the Lewis rat because it is a widely studied model of EAE and found that oral tolerization with guinea pig MBP was as suppressive as rat MBP. Of note is that oral tolerization with mouse MBP suppressed myelin-induced EAE in the SJL mouse in which autoimmunity to proteolipid protein appears to play a primary role, suggesting that antigen-driven bystander suppression following oral tolerization with autoantigens (Miller et al., 1991b) may be an important contributing mechanism for suppression of EAE following oral tolerization with MBP in this model.(ABSTRACT TRUNCATED AT 250 WORDS)
Transforming growth factor-beta 1 (TGF-beta 1) is a multifunctional growth factor that has profound regulatory effects on many developmental and physiological processes. Disruption of the TGF-beta 1 gene by homologous recombination in murine embryonic stem cells enables mice to be generated that carry the disrupted allele. Animals homozygous for the mutated TGF-beta 1 allele show no gross developmental abnormalities, but about 20 days after birth they succumb to a wasting syndrome accompanied by a multifocal, mixed inflammatory cell response and tissue necrosis, leading to organ failure and death. TGF-beta 1-deficient mice may be valuable models for human immune and inflammatory disorders, including autoimmune diseases, transplant rejection and graft versus host reactions.
A major goal of research into IDDM has been the identification of the 'causative antigen'. As described in this article by Len Harrison, this reductionist aim is confounded by the fact that numerous candidate islet cell antigens have been described. He scrutinizes the credentials of these candidates and discusses the problem of using autoantibodies to identify causative antigens in a T-cell-mediated disease.
We studied the immunogenicity and tolerogenicity of class II major histocompatibility complex (MHC) allopeptides in the rat. Inbred LEW (RT1l) rats, used as responders, were immunized in the foot pad with a mixture of eight synthetic class II MHC allopeptides emulsified in complete Freund's adjuvant. These sequences represent the full-length second domain of RT1.Bu and RT1.Du (WF) beta chains. In vitro, responder lymphocytes harvested from popliteal and inguinal lymph nodes of immunized animals exhibited significant proliferation to the MHC allopeptide mixture. In addition, these responder lymphocytes had significantly increased proliferation to allogeneic WF (RT1u) stimulator cells, when compared to naive controls in the standard one-way mixed lymphocyte response. In vivo, peptide-immunized LEW animals were challenged in the ear 2 weeks after immunization with the allopeptide mixture, the individual allopeptide sequences, or allogeneic WF splenocytes. When compared to controls, these animals had significant delayed-type hypersensitivity responses to the allopeptide mixture, to the beta-pleated sheet allopeptide sequences, and to allogeneic WF splenocytes but not to the alpha-helix allopeptide sequences, to syngeneic LEW splenocytes, or to third party allogeneic BN splenocytes. Oral administration of the allopeptide mixture to LEW responder rats daily for 5 days before immunization effected significant reduction of delayed-type hypersensitivity responses both to the allopeptide mixture and to allogeneic splenocytes. This reduction was antigen-specific, since there was no reduction of delayed-type hypersensitivity responses to mycobacterium tuberculosis. These data demonstrate that lymphocytes from animals immunized with polymorphic class II MHC allopeptides can recognize and proliferate to the same amino acid sequences on allogeneic cell surface MHC molecules. In addition, oral administration of these peptides down-regulates the systemic cell-mediated immune response in a specific fashion. Synthetic MHC allopeptides should allow the study of alloimmunity in vivo, including induction of immune tolerance.
Nylon wool adherent, CD4+ T cells from the spleens of rats that have recovered from experimental autoimmune encephalomyelitis (EAE) inhibit the in vitro production of IFN-gamma, but not IL-2, by effector cells of EAE when cocultured in the presence of myelin basic protein Ag. When anti-transforming growth factor-beta (TGF-beta) antibodies are added to the co-cultures, IFN-gamma production is restored to normal levels. Irrelevant control antibodies have no effect. The same pattern of response was obtained with cells incubated in serum-free medium. In other experiments, purified TGF-beta was added to cultures of effector cells in the presence of antigen. TGF-beta inhibited the production of IFN-gamma by these cells in a dose-dependent manner, but had no apparent inhibitory effect on IL-2 production. Finally, supernatants from cultures containing effector cells and CD4+ suppressor cells plus Ag contained measurable amounts of TGF-beta, whereas supernatants from cultures of effector cells plus Ag contained no measurable amounts of TGF-beta. These results suggest that CD4+ Ts cells of EAE regulate effector cells of EAE through a mechanism that involves the secretion of TGF-beta and that the inhibitory function of this cytokine can be reversed with neutralizing antibodies directed against TGF-beta.
We have recently reported that the oral administration of myelin basic protein (MBP) prior to encephalitogenic challenge results in suppression of experimental autoimmune encephalomyelitis (EAE). We examined the serum and salivary antibody responses to MBP in orally tolerant rats using an avidin-biotin enzyme-linked immunosorbent assay. Serum anti-MBP IgA and IgG, but not IgM levels are suppressed in orally tolerant versus control rats. This suppression is time dependent and is confined to the period when animals would otherwise be manifesting EAE clinical signs. In contrast, there is an increase in salivary anti-MBP IgA levels in MBP-fed rats relative to vehicle-fed controls. Thus, MBP-induced unresponsiveness is demonstrable at the humoral level, and moreover, a discrete compartmentalization between the serum and salivary anti-MBP responses exists.
In order to assess T cell antigen specificities and class II restriction requirements during the course of chronic relapsing experimental allergic encephalomyelitis (CREAE), (SJL x PL)F1 mice were used as a model. EAE can be passively transferred in these mice by F1 T cells incubated with Ia-positive antigen-presenting cells (APC) from either parent SJL (H-2s) or PL (H-2u) and MBP fragments 89-169 or 1-37, respectively. T cells purified from F1 mice immunized with MBP fragment 1-37 were positively selected for I-Au-supported proliferation by culture in the presence of irradiated Iau-positive PL spleen cells as APC. I-As-supported proliferation and proliferation to residues 89-169 were not detected following selection. Adoptive transfer of this T cell line induced CREAE in naive recipient F1 mice and 2 weeks after the second attack of EAE recipient proliferative responses were measured. Recipient T cells proliferated to both fragment 1-37 and fragment 89-169. Moreover, proliferation was supported by I-As-positive as well as I-Au-positive macrophages. These findings demonstrate that T cells with novel epitope specificities and class II restriction requirements can be generated during the course of CREAE and suggest the possibility that such cells may be involved in the pathogenesis of this chronic autoimmune illness.
Oral administration of myelin basic protein (MBP) suppresses experimental autoimmune encephalomyelitis (EAE) in Lewis rats immunized with MBP in Freund's adjuvant. The immunomodulator bacterial lipopolysaccharide (LPS) when given orally in conjunction with MBP enhances the protective effects of MBP feeding in EAE. This synergy was achieved only following oral administration of LPS but not following subcutaneous injection. In contrast, subcutaneous administration of LPS abrogated oral tolerance. A synergism between oral LPS and MBP was also demonstrated for antigen-specific suppression of delayed type hypersensitivity (DTH) responses. Antibody responses to MBP were suppressed by oral administration of MBP but not by MBP plus LPS. The lipid A moeity of LPS mimicked the effects of LPS on disease protection and DTH suppression. These data demonstrate that adjuvants can enhance the induction of antigen-specific oral tolerance for suppression of cell-mediated experimental autoimmune responses.
Oral administration of proteins is a long-recognized method of inducing antigen-specific peripheral immune tolerance. We previously showed that oral administration of myelin basic protein suppresses monophasic experimental autoimmune encephalomyelitis in the Lewis rat when it is given in association with immunization and prior to disease onset. As a potential therapy for human autoimmune disease, it is crucial to determine whether oral tolerance can ameliorate an ongoing immune response. We therefore asked whether oral administration of myelin antigens, after sensitization and disease expression has occurred, could affect immunological, clinical, or pathological features of experimental autoimmune encephalomyelitis. Chronic relapsing experimental autoimmune encephalomyelitis was induced in the Lewis rat and strain 13 guinea pig by immunization with whole guinea pig cord homogenate, complete Freund's adjuvant, and Mycobacterium tuberculosis. Following recovery from the first attack, animals were orally given bovine myelin, guinea pig myelin, or guinea pig myelin basic protein three times per week for up to 3 months. Animals receiving myelin products orally had decreased severity and frequency of clinical relapses, decreased delayed-type hypersensitivity responses to myelin antigens, diminished inflammation in the central nervous system (CNS), and decreased areas of CNS demyelination. In the rat, guinea pig myelin basic protein was as effective as guinea pig myelin in ameliorating the disease and also resulted in decreased serum anti-myelin basic protein antibody levels. No exacerbation of disease or worsening of pathological findings occurred in the animals given myelin products. These results demonstrate that oral administration of myelin antigens can suppress chronic relapsing experimental autoimmune encephalomyelitis and have direct relevance to therapy of human demyelinating disorders such as multiple sclerosis.
We have recently reported that experimental autoimmune encephalomyelitis (EAE) can be suppressed by the oral administration of myelin basic protein (MBP). The oral introduction of 20 mg MBP together with a trypsin inhibitor results in inhibition of EAE clinical signs, decreased CNS histopathologic changes and dramatically reduced MBP-specific proliferative responses in fed and challenged Lewis rats. In the present study, we have investigated the mechanism underlying MBP-induced oral tolerance in EAE. Neither lymphoid cells (lymph node cells, spleen cells, Peyer's patch lymphocytes, thymocytes) nor humoral elements derived from tolerant donors were capable of transferring the tolerance to naive recipients. Moreover, lymphoid cells obtained from orally tolerant donors exhibited a marked decrease in their capacity to transfer EAE to naive recipient rats, even after in vitro activation with MBP or Con A. We observed that EAE could be readily transferred into orally tolerant rats using MBP-specific encephalitogenic T cell lines. In vitro cell mixing studies showed that the proliferation of lymphocytes from MBP-sensitized donors was not inhibited by the addition of lymphoid cells from tolerant donors, arguing against the role of a suppressor cell. Investigation of MBP-stimulated lymphokine production showed that both IL-2 and IFN-gamma levels were substantially decreased in spleen and lymph node cell cultures from MBP-fed rats compared to vehicle-fed control animals. Furthermore, limiting dilution analyses revealed that MBP-fed rats exhibited a profound decrease in MBP-reactive, IL-2-secreting lymphocytes relative to control animals. Thus, because lymphocytes from MBP-fed rats neither proliferate nor secrete IL-2 or IFN-gamma in response to MBP and we can find no compelling evidence for the role of suppressor cells, we propose that the oral administration of MBP results in a state of clonal anergy.
The oral administration of S-antigen fragment (a synthetic peptide designated as peptide M and known to be uveitopathogenic for rat, guinea pig, and monkey) to Lewis rats prior to challenge with an emulsion of peptide M and CFA resulted in either a total or partial suppression of experimental autoimmune uveitis (EAU), a T cell-mediated autoimmune disease studied as a model for human uveitis and experimental autoimmune pinealitis (EPA). Both the clinical and histopathologic manifestations of the disease were suppressed in a dose-dependent manner. Pinealitis associated with EAU was also suppressed by the oral administration of peptide M. Additionally, ingestion of a fragment of baker's yeast (Saccharomyces cerevisiae) histone H3, which has five consecutive amino acids identical to peptide M and which has been found to be uveitopathogenic in Lewis rats, induced tolerance to either peptide M or synthetic histone H3 peptide. In addition, the proliferative response to peptide M was inhibited in peptide M-fed rats. The suppression of EAU and in vitro lymphocyte proliferative responses to peptide M were observed to be antigen specific, since oral feeding of a control protein (BSA) exerted no suppressive effect. Furthermore, the T cells isolated from the spleen and lymph nodes of animals rendered tolerant by oral administration of peptide M can transfer protection against EAU adoptively. These results demonstrate that the oral administration of an autoantigen or its homologous peptide initiates an antigen-specific cellular mechanism which may ameliorate EAU.
The effects of oral administration of major histocompatibility antigens on the alloimmune response have not been investigated. Lymphocytes from inbred LEW (RT1u) rats that were pre-fed allogeneic WF (RT1l) splenocytes exhibited significant antigen specific reduction of the mixed lymphocyte response in vitro and delayed-type hypersensitivity response in vivo, when compared with unfed controls. In an accelerated allograft rejection model, LEW rats were presensitized with BN (RT1n) skin allografts 7 days before challenging them with (LEW x BN)F1 or BN vascularized cardiac allografts. While sensitized control animals hyperacutely reject their cardiac allografts within 2 days, animals prefed with BN splenocytes maintained cardiac allograft survival to 7 days, a time similar to that observed in unsensitized control recipients. This phenomenon was antigen-specific, as third-party WF grafts were rejected within 2 days. Immunohistologic examination of cardiac allografts harvested on day 2 from the fed animals had markedly reduced deposition of IgG, IgM, C3, and fibrin. In addition, there were significantly fewer cellular infiltrates of total white blood cells, neutrophils, macrophages, T cells, IL-2 receptor-positive T cells, and mononuclear cells with positive staining for the activation cytokines IL-2 and IFN-g. On day 6 posttransplant, the grafts from fed animals showed immunohistologic changes typical of acute cellular rejection usually seen in unsensitized rejecting controls. Feeding allogeneic splenocytes prevents sensitization by skin grafts and transforms accelerated rejection of vascularized cardiac allografts to an acute form typical of unsensitized recipients. Oral administration of alloantigen provides a novel approach to down-regulate the specific systemic alloimmune response against histocompatibility antigens.
The adhesion of lymphocytes to endothelial cells lining the postcapillary high endothelial venules (HEV) is the first step in their emigration from the bloodstream into lymph nodes and Peyer's patches (PP). We have recently shown that the adhesiveness of cultured rat lymph node and PP HEV cells for thoracic duct lymphocytes can be increased significantly by pretreatment with TNF-alpha, IFN-gamma, and IL-4. In the present study we investigated the role of transforming growth factor-beta 1 (TGF-beta) on the adhesiveness of nonstimulated and cytokine-stimulated PP HEV cells for rat lymphocytes. The results indicated that at picomolar concentrations, TGF-beta significantly (p less than 0.001) decreased the ability of PP HEV cells to adhere 51Cr-labeled rat lymphocytes. Maximal inhibition was observed with a TGF-beta dose of 0.5 ng/ml and an incubation time of 6 to 12 h. TGF-beta did not affect the morphology of HEV cells and had no adverse effect on their viability. Moreover, the decrease in HEV adhesiveness by TGF-beta was reversible, with lymphocyte binding returning to control level 24 h after removal of the cytokine. The specificity of TGF-beta was confirmed by the ability of neutralizing anti-TGF-beta 1 antibody, but not control serum, to abolish the inhibitory properties of the cytokine. In addition, TGF-beta completely abrogated the increased adhesiveness of PP HEV cells normally induced by TNF-alpha or IFN-gamma. In contrast, TGF-beta had no effect on the stimulating effects of IL-4. Moreover, preincubation of PP HEV cells with TGF-beta did not alter the ability of these cells to respond to IL-4. Importantly, the adhesion of rat lymphocytes to IL-4-stimulated PP HEV cells can be blocked by pretreatment of lymphocytes with the PP-homing receptor-specific 1B.2.6 antibody whereas pretreatment of human mononuclear cells with anti-very late activation antigen-4 alpha antibody inhibited only partially the binding of these cells to the IL-4-stimulated PP HEV monolayers. Taken together, these findings strongly suggest that TGF-beta and IL-4 play important regulatory roles in lymphocyte-HEV adhesion and that the stimulatory effect of IL-4 is mediated at least in part through the increased expression of organ-specific ligands on HEV cells.
The in vitro production of a variety of cytokines by lymphocytes isolated from spleen mesenteric lymph node (MLN), Peyer's patches (PP) and lamina propria (LP) was measured, after oral immunization with keyhole limpet hemocyanin using cholera toxin as an adjuvant. LP responses were characterized by very high levels of interleukin (IL) 4, IL 5 and IL 6 with lower levels of IL 2 and interferon-gamma (IFN-gamma). The PP had lower levels of IL 4, IL 5 and IL 6 than LP but higher levels of IL 2, IFN-gamma was only present at very low levels in this organ. The MLN had a pattern of cytokine production similar to the PP but did produce IFN-gamma. The spleen produced all cytokines measured except IL 5. Antibody production was characterized by IgA in the LP and PP but IgG was the dominant isotype in the spleen, the MLN was a poor source of antibody-producing cells. We interpret the results to show that (a) the LP response to cholera toxin/keyhole limpet hemocyanin is dominated by Th2-type cytokines compared to a lower production of Th1 type and (b) that the PP has responses typical of an organ with a high proportion of resting lymphocytes which develop mainly into Th2-types cells. The spleen is less dominated by Th2-type cytokines than the mucosal sites and this difference is paralleled by IgA antibody production at the mucosal sites and IgG antibody dominance in the spleen.
Adjuvant arthritis is induced by intradermal injection of Mycobacterium tuberculosis (MT) in oil. The role of immunity to type II collagen (CII) in adjuvant arthritis (AA) has not been well defined. We found that oral administration of chicken CII given 3 micrograms per feeding on days -7, -5, and -2 before disease induction consistently suppressed the development of AA. A decrease in delayed-type hypersensitivity responses to CII was also observed that correlated with suppression of AA. AA was optimally suppressed by 3 and 30 micrograms of collagen type II variably by 300 micrograms, and not by 0.3 microgram or 1 mg. Oral administration of collagen type I also suppressed AA; only minimal effects were seen with collagen type III. Suppression was Ag specific: feeding CII did not suppress experimental autoimmune encephalomyelitis; feeding myelin basic protein suppressed experimental autoimmune encephalomyelitis, but not AA. Suppression of AA could not be consistently obtained by feeding MT. Suppression of AA could be adoptively transferred by T cells from CII fed animals and could be obtained when CII was fed after disease onset. Our results suggest that autoimmunity to CII has a pathogenic role in AA and raise the possibility that cross-reactive epitopes exist between CII and MT. Alternatively, the pathogenesis of AA may be dependent on developing immunity to CII. These results further demonstrate the effectiveness of oral tolerance as a means to suppress experimental autoimmune diseases.
Our earlier studies have demonstrated that intragastric administration of the hapten trinitrochlorobenzene (TNCB) 2 to 3 weeks prior to attempting sensitization with epidermally applied hapten can abrogate development of systemic contact sensitivity (CS). In this paper, we have examined whether onset of tolerance following intragastric administration of the hapten is preceded by development of hapten-specific CS. Indeed, CS was found to be present 5 days after feeding TNCB and in most experiments the response decreased significantly by Days 10 to 12. The kinetics of development of CS by the oral and epidermal routes were strikingly similar except that the magnitude of reactivity (up to 5 days) in orally sensitized mice was somewhat less than that of epidermally sensitized mice. With the exception of Peyer's patches (PP), effector cells of CS were recovered from such gut-associated lymphoid tissues as mesenteric lymph nodes (MLN), lamina propria, and lymphocytes that are present in the intraepithelial compartment of the intestinal wall. These cells as well as spleen cells of TNCB-fed mice were able to adoptively transfer CS to naive mice. The capacity of MLN and spleen cells of TNCB-fed mice to confer CS adoptively was abrogated after treating cells with anti-Thy 1.2 and anti-Lyt 1.1 antibodies plus complement thereby identifying them as T lymphocytes. Although CS decreased by 10-12 days after feeding TNCB, the decline was reversed by pretreating mice with cyclophosphamide (CY) 2 days before giving the hapten. Whereas spleen cells from animals fed hapten 5 days earlier transferred CS readily, those from mice fed hapten 12 days earlier did not. However, when 12-day spleen cells were depleted of Lyt 2+ cells their ability to adoptively transfer CS was restored. These observations indicate that feeding TNCB to mice initially produces CS, mediated by Thy 1.2+, Lyt 1.1+ lymphocytes. CS is subsequently down-regulated by activation of Lyt 2+ suppressor cells, precursors of which are sensitive to CY.
The ability to prevent the expression of retinal SAg induced experimental autoimmune uveitis (EAU) in Lewis rats by oral administration of the SAg and SAg fragments was investigated. Oral administration of the SAg molecule prevented or markedly diminished the clinical appearance of SAg-induced disease as measured by ocular inflammation. Furthermore, oral administration of the SAg also markedly diminished uveitis induced by the uveitogenic M and N fragments of the SAg. M and N fragments were not effective in preventing SAg-induced EAU, although feeding the M fragment prevented disease induced by the M fragment. Oral administration of the SAg did not prevent myelin basic protein induced experimental autoimmune encephalomyelitis, whereas feeding myelin basic protein did. In vitro studies demonstrated a significant decrease in proliferative responses to the SAg in lymph node cells draining the site of immunization from fed vs nonfed animals. Furthermore, the addition of splenocytes from SAg-fed animals to cultures of a CD4+ SAg-specific cell line profoundly suppressed the cell line's response to the SAg, whereas these splenocytes had no effect on a purified protein derivative-specific cell line. The Ag-specific in vitro suppression was blocked by anti-CD8 antibody (OX-8) demonstrating that this suppression is dependent on CD8+ T-cells. These experiments demonstrate that Ag-specific immunomanipulation can be achieved in the EAU model by oral administration of the SAg and raise the possibility that such an approach may have practical clinical implications in uveitis as well as other human autoimmune diseases.
The role of murine epithelial cells from small intestine (IEC) in orally induced immune unresponsiveness has been investigated. Murine IEC were shown to express a relatively low level of class II MHC Ag and were incapable of presenting BSA to Ag-specific Th cells. IFN-gamma treatment of the animals dramatically enhanced expression of class II MHC Ag by IEC enabling Ag presentation to Th cells. Ag-presenting function of IEC after IFN-gamma treatment was not affected by IEC panning on anti-IgG or anti-MAC-1-coated plates but it was blocked by a class II MHC-specific mAb. IFN-gamma-treated mice were responsive to parenteral BSA challenge after oral administration of the Ag, whereas untreated mice were immunosuppressed. Our data support the concept that poor expression of class II MHC Ag on IEC contributes to immunosuppression induced by oral administration of a protein Ag.
Transforming growth factor-beta 1 (TGF-beta 1) belongs to a family of polypeptides involved in the regulation of cell growth and differentiation. We have examined the ability of TGF-beta 1 to regulate isotype specific Ig secretion by murine spleen B cells. TGF-beta 1, in the presence of rIL-2, induced a synergistic 10-fold or greater increase in IgA secretion by LPS-stimulated spleen B cells. TGF-beta 1 alone had little to no effect on IgA secretion. In contrast, TGF-beta 1, with or without rIL-2, markedly inhibited IgG1 and IgM secretion under the same conditions. The costimulatory activity of TGF-beta 1 and rIL-2 on IgA secretion was seen in cultures of surface IgA negative B cells and was inhibited by anti-TGF-beta 1 antibody in a dose dependent manner. Vicia villosa agglutinin non-adherent Peyer's patch T cells, which secrete IL-2, also synergized with TGF-beta 1 and could substitute for the activity of LPS and rIL-2 on the IgA response. Finally, IL-5 added after 2 days of culture, but not at the beginning of culture, synergized with TGF-beta 1 on the IgA response. These studies indicate that TGF-beta 1 can interact with other lymphokines and selectively modulate the IgA response.
We report that experimental autoimmune encephalomyelitis, a T cell-mediated autoimmune disease studied as a model for multiple sclerosis, can be suppressed in Lewis rats by the oral administration of myelin basic protein (MBP). Both the clinical and histopathologic manifestations of the disease were suppressed in a dose-dependent manner. In addition, proliferative responses to MBP and, to a lesser extent, serum levels of anti-MBP antibody were suppressed by feeding MBP. Suppression of clinical and histologic disease was observed whether animals were fed MBP before or after disease induction, although suppression was more complete when rats were fed before immunization. Disease was also suppressed by the oral administration of either encephalitogenic or nonencephalitogenic fragments and decapeptides of the MBP molecule, with more complete suppression observed when nonencephalitogenic fragments were fed, suggesting that suppressor determinants exist in the MBP molecule distinct from the encephalitogenic region. The oral administration of a non-disease-inducing portion of an autoantigen represents an antigen-specific method by which an experimental autoimmune disease can be immunoregulated.
The oral administration of myelin basic protein (MBP) to Lewis rats prior to an encephalitogenic challenge resulted in total inhibition or a significant delay in the onset of experimental autoimmune encephalomyelitis (EAE). In vitro lymphocyte proliferative responses to MBP were significantly decreased in MBP-fed rats when compared with vehicle-fed controls. Suppression of EAE and in vitro proliferative responses to MBP were observed to be antigen specific, since oral feeding of a control protein exerted no suppressive effect. Moreover, the specificity of MBP-induced oral tolerance was shown to be species specific, since feeding guinea pig MBP (GPMBP) or human MBP (HuMBP) induced protection only against a GPMBP or HuMBP challenge, respectively. Conversely, Lewis rats could not be orally tolerized to the self antigen rat MBP.
We have previously demonstrated that the oral administration of guinea pig myelin basic protein (MBP) protects Lewis rats against the induction of experimental autoimmune encephalomyelitis (EAE) when subsequently immunized with guinea pig MBP in CFA. In addition, animals made orally tolerant to MBP also have diminished proliferative and antibody responses to MBP, but not to other Ag. Nonetheless, the mechanism of oral tolerance to MBP in the EAE model remains undefined. In the present study, we report that T cells isolated from the spleen and mesenteric lymph nodes of MBP orally tolerized animals can adoptively transfer protection against EAE. Furthermore, these T cells are of the CD8+ subclass. In addition, CD8+ T cells from MBP orally tolerized animals also suppress in vitro proliferative responses and antibody responses to MBP in an Ag-specific fashion. These results demonstrate that active cellular mechanisms are initiated after oral administration of an autoantigen that can down-regulate an experimental autoimmune disease and provide the basis for the isolation and characterization of the cells mediating both in vivo and in vitro suppression.