ANERGY IN-VIVO - DOWN-REGULATION OF ANTIGEN-SPECIFIC CD4+ T(H)1 BUT NOT T(H)2 CYTOKINE RESPONSES

Efficient immunologic tolerance, defined as antigen-specific unrespons iveness, can be peripherally induced by the i.v. injection of syngenei c splenocytes coupled with antigen using ethylene carbodiimide (ECDI). We have previously reported that unresponsiveness induced via i.v. in jection of syngeneic splenocytes coupled with intact, UV-inactivated T heiler's murine encephalomyelitis virus (TMEV-SP) resulted in 'split t olerance'. Both virus-specific delayed-type hypersensitivity and IgG2a levels were inhibited, whereas IgG1 levels were increased when compar ed with sham tolerized controls. In the present report we demonstrate that tolerance induced by i.v. injection of TMEV-coupled splenocytes r esulted in antigen-specific inhibition of T cell proliferation, as wel l as IL-2 and IFN-gamma production in response to both whole TMEV and the immunodominant viral epitope. Additionally, tolerance induction re sulted in abrogation of T(h)1-derived [IL-2, IFN-gamma and LT/tumor ne crosis factor-beta (TNF-beta)] cytokine mRNA expression in response to in vitro stimulation with UV-inactivated TMEV as determined by revers e transcriptase polymerase chain reaction. In contrast, expression of T(h)2-derived (IL-4, IL-6 and IL-10) cytokine mRNA was not affected in tolerized mice. Tolerance functioned directly at the level of CD4+ T( h)1 cells at both the induction and effector limbs as depletion of CD8 + T cells both prior to in vivo tolerization or in vitro culture had n o effect on inhibition of T(h)1-specific responses. The mechanism of i n vivo tolerance induction appeared to be anergy of CD4+ T(h)1 cell si nce IL-2, IFN-gamma and LT/TNF-beta mRNA expression as well as virus-s pecific proliferative responses could be restored by addition of rIL-2 to in vitro cultures of tolerant, CD4+ T(h)1 populations. These resul ts suggest that in vivo 'split tolerance' induced by i.v. injection of ECDI-fixed, antigen-coupled splenocytes involves anergy of TMEV-speci fic, CD4+ T(h)1 lymphocytes and concomitant priming of T(h)2 cells. Th e induction of antigen-specific, in vivo anergy has important implicat ions in the design of therapeutic strategies for immunopathologic dise ases mediated by T(h)1 lymphocytes, especially T cell-mediated autoimm une disorders.