T lymphocytes and cytokines in the pathogenesis of experimental autoimmune myasthenia gravis

Abstract

Experimental autoimmune myasthenia gravis (EAMG), induced in different animal species and strains by immunization with nicotinic acetylcholine receptor (AChR) and complete Freund's adjuvant (CFA), represents an experimental model of myasthenia gravis (MG) in humans. Both MG and EAMG are autoimmune diseases mediated by antibodies against AChR of neuromuscular junctions. The production of anti-AChR antibodies is dependent on T cell help by cytokines.

To study the role of the CD8+ T cells in the pathogenesis of EAMG, we depleted these cells in Lewis rats using OX8 monoclonal antibody post immunization (p.i). Depletion of CD8+ T cells effectively suppresses clinical EAMG and anti-AChR antibody levels, accompanied with decreased levels of IFN-y and IL-4, but not TGF-beta mRNA expressing cells in lymphoid organs. When CD4-/-, CD8-/- and CD4-8-mutant C57BL/6J mice were immunized with AChR + CFA, clinical EAMG was nearly completely prevented. This was associated with strongly reduced AChR-specific T and B cell responses, and with reduced levels of AChR-reactive IFN-y and IL-4 mRNA expressing cells in lymphoid organs when compared to CD4+8+ wild type mice.

IL-IO has been considered as an immunosuppressive factor in some T cell-mediated autoimmunities. To evaluate its role in antibody-mediated EAMG, we injected rhlL-10 and showed an earlier and more pronounced muscular weakness compared to PBS-injected rats. Levels of anti-AChR antibodies, anti-AChR IgG antibody secreting cells and IL-4 mRNA expressing cells were upregulated, while lymphocyte proliferative responses to AChR and the levels of Th l cytokines IFN-y, IL-1beta, IL-12, and TNFalpha mRNA expressing cells were suppressed.

After immunization with AChR, EAMG-susceptible Lewis rats and -resistant Wistar Furth (WF) rats had similarly elevated concentrations and affinities of serum anti-AChR antibodies and IL-4 mRNA expressing cells. In contrast, T cell responses to AChR measured by proliferation and by enumeration of IFN-y secreting and mRNA expressing cells were lower in WF rats. This strain showed, instead, an upregulation of the anti-inflammatory cytokine TGF-beta. Current immunotherapies of both EAMG and MG are nonspecific and toxic. Nasal administration of myg doses of Torpedo AChR prior to immunization resulted in the prevention of EAMG, the suppression of anti-AChR antibody levels, AChR-specific IgG secreting cells, AChR-reactive IFN-y secreting cells and T cell proliferation. AChR-specific CD4+CD8-T cell clones established from nasally tolerized rats displayed decreased AChR-induced proliferation, decreased expression of leukocyte function-associated antigen-1 (LFA-I) on cell surface, but elevated numbers of TGF-beta mRNA expressing cells compared to CD4+ clones from non-tolerized rats, indicating that the suppression of EAMG by nasal tolerance is associated with upregulation of TGF-beta in suppressor CD4+ cells and downregulation of LFA-I expression on the T cell surface.

We conclude that (1) both CD4+ and CD8+ T cells are essential for development of EAMG, and a collaboration between these cell types may be necessary; (2) CD4+ as well as CD8+ T cells secrete IFN-y and IL-4, and both cytokines are involved in the development of EAMG; (3) besides T cells, other immune cells might also be responsible for help of anti-AChR antibody production, since there still were some anti-AChR antibodies existing in the CD4-8-mice; (4) IFN-y, TNFalpha, IL-1beta, IL-12, IL-4, IL-1O and LFA-I are involved in the development of EAMG, while TGF-beta plays an important role in tolerance induction and in the strain-resistance to this disease; (5) suppressor CD4+ T cells can produce TGF-beta after induction of tolerance.