Oral Administration Of Myelin Basic Protein Research Articles

The Thymus Plays a Role in Oral Tolerance in Experimental Autoimmune Encephalomyelitis

The oral administration of myelin proteins has been used for the successful prevention and treatment of experimental autoimmune encephalomyelitis (EAE). We questioned whether the thymus was involved in oral tolerance. In this study, euthymic myelin basic protein (MBP) TCR transgenic mice are protected from EAE when fed MBP but are not protected when thymectomized. Similarly, in a cell transfer system, T cell responses to OVA measured in vivo were suppressed significantly only in the OVA-fed euthymic mice but not in the thymectomized mice. We observed that the absence of the thymus dramatically enhanced the Th1 response. We explored three alternatives to determine the role of the thymus in oral tolerance: 1) as a site for the induction of regulatory T cells; 2) a site for deletion of autoreactive T cells; or 3) a site for the dissemination of naive T cells. We found that Foxp3(+)CD4(+)CD25(+) T cells are increased in the periphery but not in the thymus after Ag feeding. These CD4(+)CD25(+) T cells also express glucocorticoid-induced TNFR and intracellular CTLA4 and suppress Ag-specific proliferation of CD4(+)CD25(-) cells in vitro. The thymus also plays a role in deletion of autoreactive T cells in the periphery following orally administered MBP. However, thymectomy does not result in homeostatic proliferation and the generation of memory cells in this system. Overall, the oral administration of MBP has a profound effect on systemic immune responses, mediated largely by the generation of regulatory T cells that act to prevent or suppress EAE.

T-cell activation and receptor downmodulation precede deletion induced by mucosally administered antigen.

The fate of antigen-specific T cells was characterized in myelin basic protein (MBP) T-cell receptor (TCR) transgenic (Tg) mice after oral administration of MBP. Peripheral Th cells are immediately activated in vivo, as indicated by upregulation of CD69 and increased cytokine responses (Th1 and Th2). Concurrently, surface TCR expression diminishes and internal TCR levels increase. When challenged for experimental autoimmune encephalomyelitis during TCR downmodulation, Tg mice are protected from disease. To characterize Th cells at later times after antigen feeding, it was necessary to prevent thymic release of naive Tg cells. Therefore, adult Tg mice were thymectomized before treatment. TCR expression returns in thymectomized Tg mice 3 days after MBP feeding and then ultimately declines in conjunction with MBP-specific proliferation and cytokine responses (Th1-type and Th2-type). The decline correlates with an increase in apoptosis. Collectively, these results demonstrate that a high dose of fed antigen induces early T-cell activation and TCR downmodulation, followed by an intermediate stage of anergy and subsequent deletion.

Suppression of murine chronic relapsing experimental autoimmune encephalomyelitis by the oral administration of myelin basic protein.

Chronic relapsing experimental autoimmune encephalomyelitis (EAE), induced in mice by the injection of myelin basic protein (MBP), is a T cell-mediated autoimmune disease characterized by periods of paralysis and remission. We have shown previously that the oral administration of MBP or MBP peptides renders Lewis rats refractory to EAE. This study was undertaken to examine the conditions necessary to produce oral tolerance in a chronic relapsing model of EAE in B10.PL mice. The optimal tolerizing regimen for the mouse was found to be a single feeding of 20 mg of MBP suspended in PBS. To determine the ability to suppress chronic disease, a range of doses (0.4-100 mg) was administered orally in a single dose before challenge. Larger oral doses (20 or 100 mg) of MBP provided the best protection from EAE, while 0.4 mg exacerbated the clinical course of disease. Secretion of the proinflammatory cytokines, IL-2 and IFN-gamma, were lowest in the group fed 20 mg. A single feeding of MBP before challenge or as late as the first day of clinical signs showed significant protection over the relapsing disease course. Once relapsing EAE was established, multiple oral doses of MBP were required to achieve suppression of clinical signs of disease. These findings suggest that vehicle, dosage, and timing are important considerations in the successful application of oral tolerance strategies for suppression of chronic disease processes.

Antigen-Driven Peripheral Immune Tolerance: Suppression of Experimental Autoimmune Encephalomyelitis and Collagen-Induced Arthritis by Aerosol Administration of Myelin Basic Protein or Type II Collagen

Antigen-driven tolerance is an effective method of suppressing cell-mediated immune responses. We have previously demonstrated that exposure of gut-associated lymphoid tissue to myelin basic protein (MBP) via oral administration suppresses experimental autoimmune encephalomyelitis (EAE). To further study presentation of antigen to the immune system by mucosal surfaces as a method of antigen-driven tolerance, the effect of inhalation of MBP was investigated. MBP was given as an aerosol to Lewis rats on Days −10, −7, −5, and −3 prior to immunization with MBP in Freund's adjuvant and on Days 0, 2, and 4 following immunization. Aerosolization of MBP completely abrogated clinical EAE in 100% of treated rats. Central nervous system inflammation and delayed-type hypersensitivity and antibody responses to MBP were also significantly reduced in aerosol-treated animals. Aerosolization of histone, a basic protein of similar weight and charge as MBP, had no effect. Disease was also suppressed with one aerosol treatment on Day −3 or by administering MBP nasally. Aerosolization was more effective than oral administration of MBP over a wide dose range (0.005–5 mg). Splenic T cells isolated from animals postaerosolization adoptively transferred protection to naive animals immunized with MBP. Aerosolization of MBP to animals with relapsing EAE after recovery from the first attack decreased the severity of a subsequent attack. Aerosol and oral MBP were equally effective at suppressing thein vitroimmune response as measured by proliferation and interferon-γ production. We then tested aerosolization of a different autoantigen in a different disease model and found that aerosolization of type II collagen was effective in suppressing collagen-induced arthritis. Thus, aerosolization of an autoantigen is a potent method to downregulate an experimental T cell-mediated autoimmune disease and suggests that exposure of antigen to lung mucosal surfaces preferentially generates immunologic tolerance.

Induction of oral tolerance to myelin basic protein in CD8-depleted mice: both CD4+ and CD8+ cells mediate active suppression.

We have previously shown that orally administered myelin basic protein (MBP) suppresses experimental autoimmune encephalomyelitis in both the Lewis rat and the SJL mouse. In the Lewis rat fed low doses of MBP, we found that protection can be adoptively transferred by CD8+ cells and that these cells inhibit immune responses via the secretion of TGF-beta after Ag-specific triggering. In the present study, we investigated the cellular requirements for the generation of active suppression following oral administration of MBP in SJL and (PLJ x SJL)F1 mice. We first determined the frequency of MBP cells secreting Th1 (IFN-gamma) and Th2 (IL-4/IL-10) cytokines or TGF-beta after oral administration of MBP. We found that in SJL mice, orally administered MBP (0.5 mg/feeding) led to an increased frequency of TGF-beta-, IL-4-, and IL-10-secreting cells and a decreased frequency of IFN-gamma-producing cells. This pattern was observed in both CD4+ and CD8+ populations; adoptive transfer of either CD4+ or CD8+ cells from orally tolerized mice suppressed autoimmune encephalomyelitis in recipient animals. We then studied the role of CD8+ cells on the generation of oral tolerance to MBP by depleting CD8+ cells in vivo with anti-CD8 mAb. Oral tolerance was successfully induced in such animals, as demonstrated by a decrease in clinical disease and T cell proliferative responses, although there was less TGF-beta production in vitro and less disease protection on days 20 to 22 in CD8-depleted animals. These studies demonstrate that CD4+ cells in the absence of CD8+ cells can mediate the active suppression component of oral tolerance in mice and that there is a reciprocal relationship between Th1- and Th2-type cytokine production associated with oral tolerization.

Orally administered myelin basic protein in neonates primes for immune responses and enhances experimental autoimmune encephalomyelitis in adult animals.

Antigen-driven tolerance is an effective method for suppression of autoimmune diseases. Adult animals can be tolerized against the induction of experimental autoimmune encephalomyelitis (EAE) by both oral and parenteral administration of myelin basic protein (MBP). We have found that in contrast to previous studies of neonatal tolerance in which parenterally administered autoantigens induced tolerance, the oral administration of MBP in neonatal rats did not result in tolerization to MBP, but instead, primed for immunologic responses. Proliferative responses to MBP and its encephalitogenic epitope were present in animals fed with MBP as neonates and co-culture of encephalitogenic T cells with cells from neonatal rats fed with MBP were associated with enhanced MBP responses rather than the suppression observed with cells from adult rats fed with MBP. Furthermore, neonates fed with MBP and immunized 6-8 weeks later with MBP in adjuvant to induce EAE revealed enhancement of disease severity, and were not protected from a second attack upon active reinduction of EAE. Subcutaneous injection of soluble MBP into neonates had no effect on EAE induction as adults, whereas intraperitoneal injection of MBP in neonates was associated with marked suppression of disease in adults. Suppression of EAE began to appear in animals fed with MBP at 4 weeks of age, and was similar to oral tolerance in adult animals when animals were fed at 6 weeks of age. These results suggest that immaturity of the immunoregulatory network associated with oral tolerance and sensitization to autoantigens via the gut in the neonatal period may contribute to the pathogenesis of autoimmune diseases.

Suppression of experimental autoimmune encephalomyelitis by oral administration of myelin basic protein: III. Synergistic effect of lipopolysaccharide

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.