Regulatory Cells Generated by Testicular Tolerization to Retinal S-Antigen: Possible Involvement of IL-4, IL-10, and TGF-β in the Suppression of Experimental Autoimmune Uveoretinitis

doi:10.1006/cimm.1997.1229

Cellular Immunology

Volume 182, Issue 2, 15 December 1997, Pages 89-98

https://doi.org/10.1006/cimm.1997.1229 Get rights and content

Abstract

Intratesticular injection of a retinal protein (S-antigen) into Lewis rats induces systemic immunotolerance (designated orchidic tolerance) and renders animals refractory to experimental autoimmune uveoretinitis (EAU) produced by S-antigen immunization. We demonstrated in this work that the immunotolerance could be transferred to syngeneic naive rats by both CD4⁺and CD8⁺regulatory cells. Attempts were then made to characterize the cytokines involved in the immunosuppressive activity of these regulatory cells. Using thein vitrolymphoproliferation assay, the inhibitory effect of CD4⁺cells on effector cells was found to be reversed by antibodies against IL-4 and IL-10 but not by anti-TGF-β antibody. IL-4 (IC₅₀= 1.6 ng/10⁶cells) and IL-10 (IC₅₀= 0.6 ng/10⁶cells) added to the assay medium were potent inhibitors of effector cell proliferation. Increased immunoreactivity and mRNA expression for IL-4 and IL-10 was observed for CD4⁺regulatory cells. The inhibitory effect of CD8⁺regulatory cells was reversed by anti-TGF-β antibody but not by antibodies against IL-4 and IL-10. Compared with control CD8⁺cells, CD8⁺cells from tolerized rats demonstrated higher immunoreactivity for TGF-β but did not show an enhanced expression of mRNA for TGF-β. TGF-β1 and TGF-β2 added to effector cells showed dichotomous effects; both isoforms stimulated cell proliferation at 2.5 ng/10⁶cells and inhibited at lower or higher concentrations. These results led us to conclude that IL-4 and IL-10 are important cytokines for the immunosuppressive effect of CD4⁺regulatory cells generated in orchidic tolerance. TGF-β is an important immunosuppressive cytokine for CD8⁺regulatory cells but further studies will determine whether other cytokines are also involved.

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The importance of transforming growth factor-beta-1 (TGF-β1) in immunoregulation and tolerance has been increasingly recognized. It is now proposed that there are populations of regulatory T cells (T-reg), some designated T-helper type 3 (Th3), that exert their action primarily by secreting this cytokine. Here, we emphasize the following concepts: (1) TGF-β1 has multiple suppressive actions on T cells, B cells, macrophages, and other cells, and increased TGF-β1 production correlates with protection and/or recovery from autoimmune diseases; (2) TGF-β1 and CTLA-4 are molecules that work together to terminate immune responses; (3) Th0, Th1 and Th2 clones can all secrete TGF-β1 upon cross-linking of CTLA-4 (the functional significance of this in autoimmune diseases has not been reported, but TGF-β1-producing regulatory T-cell clones can produce type 1 inflammatory cytokines); (4) TGF-β1 may play a role in the passage from effector to memory T cells; (5) TGF-β1 acts with some other inhibitory molecules to maintain a state of tolerance, which is most evident in immunologically privileged sites, but may also be important in other organs; (6) TGF-β1 is produced by many cell types, is always present in the plasma (in its latent form) and permeates all organs, binding to matrix components and creating a reservoir of this immunosuppressive molecule; and (7) TGF-β1 downregulates adhesion molecules and inhibits adhesion of leukocytes to endothelial cells. We propose that rather than being passive targets of autoimmunity, tissues and organs actively suppress autoreactive lymphocytes. We review the beneficial effects of administering TGF-β1 in several autoimmune diseases, and show that it can be effectively administered by a somatic gene therapy approach, which results in depressed inflammatory cytokine production and increased endogenous regulatory cytokine production.
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Mechanisms of mucosal tolerance induction, including anergy/deletion and active suppression are frequently described as mutually exclusive; dependent upon nature, dose and route of antigen administration. We have previously described induction of low-dose tolerance with administration of retinal autoantigens via the nasorespiratory tract which is antigen-specific, suppresses both cell mediated immunity and ultimately tissue destruction in experimental autoimmune uveoretinitis (EAU) and is mediated by splenic-derived regulatory cells. The present data further shows that splenocytes or fractionated splenic T cells, which secrete IL-4 and IL-10 when stimulated with retinal antigen in vitro , and not regional drainage lymph node cells transfer tolerance to naı̈ve animals. Analysis of apparent mechanistic differences shows that during intranasal antigen administration, the proportion of CD4⁺T cells within drainage lymph nodes increases, concurrent with a burst of IFN-γ Following subsequent antigen challenge, T cells downregulate αβTCR expression and undergo apoptosis in regional drainage lymph nodes. An increase in functional Th2 cytokine activity was noted in both Con-A and retinal antigen stimulated lymph node cultures in tolerised animals. T cells from tolerized animals secreted IL-4, whereas IL-10 was secreted predominantly by the non-T cell population present equally in control and tolerized animals. Therefore, spleen derived regulatory cells which suppress Th1 responses and T cell deletion/apoptosis in regional drainage lymph nodes are mechanisms which co-exist in tolerant rats. Th2 cytokine production after immunization appears consequential to tolerance-induced Th1 suppression.

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Regular ArticleRegulatory Cells Generated by Testicular Tolerization to Retinal S-Antigen: Possible Involvement of IL-4, IL-10, and TGF-β in the Suppression of Experimental Autoimmune Uveoretinitis

Abstract

J. Neuroimmunol.

Clin. Immunol. Immunopathol.

J. Autoimmunity

Immunol. Lett.

J. Immunol.

Annu. Rev. Immunol.

Invest. Ophthalmol. Vis. Sci.

Immunol.

Ocular Immunol. Inflam.

Reg. Immunol.

FASEB J.

Autoimmunity

J. Immunol.

Intl. Immunol.

Regular Article
Regulatory Cells Generated by Testicular Tolerization to Retinal S-Antigen: Possible Involvement of IL-4, IL-10, and TGF-β in the Suppression of Experimental Autoimmune Uveoretinitis