REGULATORY CELLS GENERATED BY TESTICULAR TOLERIZATION TO RETINAL S-ANTIGEN - POSSIBLE INVOLVEMENT OF IL-4, IL-10, AND TGF-BETA IN THE SUPPRESSION OF EXPERIMENTAL AUTOIMMUNE UVEORETINITIS

Intratesticular injection of a retinal protein (S-antigen) into Lewis rats induces systemic immunotolerance (designated orchidic tolerance) and renders animals refractory to experimental autoimmune uveoretiniti s (EAU) produced by S-antigen immunization. We demonstrated in this wo rk that the immunotolerance could be transferred to syngeneic naive ra ts by both CD4(+) and CD8(+) regulatory cells. Attempts were then made to characterize the cytokines involved in the immunosuppressive activ ity of these regulatory cells. Using the in vitro lymphoproliferation assay, the inhibitory effect of CD4(+) cells on effector cells was fou nd to be reversed by antibodies against IL-4 and IL-10 but not by anti -TGF-beta antibody. IL-4 (IC50 = 1.6 ng/10(6) cells) and IL-10 (IC50 = 0.6 ng/10(6) cells) added to the assay medium were potent inhibitors of effector cell proliferation. Increased immunoreactivity and mRNA ex pression for IL-4 and IL-10 was observed for CD4(+) regulatory cells. The inhibitory effect of CD8(+) regulatory cells was reversed by anti- TGF-beta antibody but not by antibodies against IL-4 and IL-10. Compar ed with control CD8(+) cells, CD8(+) cells from tolerized rats demonst rated higher immunoreactivity for TGF-beta but did not show an enhance d expression of mRNA for TGF-beta. TGF-beta 1 and TGF-beta 2 added to effector cells showed dichotomous effects; both isoforms stimulated ce ll proliferation at 2.5 ng/10(6) cells and inhibited at lower or highe r concentrations. These results led us to conclude that IL-4 and IL-10 are important cytokines for the immunosuppressive effect of CD4(+) re gulatory cells generated in orchidic tolerance. TGF-beta is an importa nt immunosuppressive cytokine for CD8(+) regulatory cells but further studies will determine whether other cytokines are also involved. (C) 1997 Academic Press.