ABSTRACT
An in vitro priming system of murine naive splenocytes was established to investigate early immune responses to Leishmania chagasi, the agent of visceral leishmaniasis in the New World. Priming of splenocytes from resistant C3H and CBA or susceptible BALB and B10 mice with L. chagasi resulted in blast transformation and in proliferating parasite-specific CD4(+) T cells secreting a differential complement of cytokines (IFN-gamma and low IL-10 levels for resistant T cells; IFN-gamma, IL-4 and high IL-10 levels for susceptible T cells). After priming, intracellular parasite load was much higher in susceptible than in resistant-type splenocyte cultures. On the other hand, infection of purified splenic macrophages from either resistant or susceptible mice with live L. chagasi promastigotes, resulted in comparable parasite loads. Moreover, when early CD4(+) T cell priming in splenocyte cultures was disrupted with anti-CD4 mAb, polarized parasite growth was abolished, becoming comparable in resistant and susceptible cultures. Neutralizing IL-4 activity during splenocyte priming did not affect the final parasite load in susceptible cultures. However, neutralizing IL-10 activity markedly decreased parasite load in susceptible, but not in resistant splenic macrophages. These results suggest that IL-10 plays an important role in L. chagasi infection in susceptible hosts. The results also indicate that innate control of growth of a visceralizing Leishmania in splenic macrophages results from the ability to activate different CD4(+) T cell subsets.
Subject(s)
Leishmania/growth & development , Macrophages/parasitology , T-Lymphocytes, Helper-Inducer/parasitology , Animals , CD4 Antigens/immunology , Cell Division , Enzyme-Linked Immunosorbent Assay , Female , Interferon-gamma/analysis , Interleukin-10/analysis , Interleukin-4/analysis , Leishmania/isolation & purification , Macrophages/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Inbred CBA , Parasite Egg Count , Spleen/cytology , T-Lymphocytes, Helper-Inducer/drug effects , T-Lymphocytes, Helper-Inducer/immunologyABSTRACT
Recent studies indicate important roles for CTLA-4 engagement in T cells, and for TGF-beta production in the immunopathogenesis of murine kalaazar or visceral leishmaniasis, but a functional link between these two pathways in helping intracellular parasite growth is unknown. Here we report that Ag or anti-CD3 activation of splenic CD4+ T cells from visceral leishmaniasis leads to intense CTLA-4-mediated TGF-beta1 production, as assessed either by CTLA-4 blockade or by direct CTLA-4 cross-linkage. Production of TGF-beta1 accounted for the reciprocal regulation of IFN-gamma production by CTLA-4 engagement. Following CD4+ T cell activation, intracellular growth of Leishmania chagasi in cocultured splenic macrophages required both CTLA-4 function and TGF-beta1 secretion. Cross-linkage of CTLA-4 markedly increased L. chagasi replication in cocultures of infected macrophages and activated CD4+ T cells, and parasite growth could be completely blocked with neutralizing anti-TGF-beta1 Ab. Exogenous addition of rTGF-beta1 restored parasite growth in cultures protected from parasitism by CTLA-4 blockade. These results indicate that the negative costimulatory receptor CTLA-4 is critically involved in TGF-beta production and in intracellular parasite replication seen in murine kalaazar.
Subject(s)
Adjuvants, Immunologic/physiology , Antigens, Differentiation/physiology , Immunoconjugates , Immunosuppressive Agents/pharmacology , Leishmaniasis, Visceral/immunology , Transforming Growth Factor beta/physiology , Abatacept , Animals , Antigens, CD , Antigens, Differentiation/immunology , Antigens, Differentiation/metabolism , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/parasitology , CTLA-4 Antigen , Cells, Cultured , Epitopes, T-Lymphocyte/immunology , Female , Immune Sera/pharmacology , Immunity, Cellular/immunology , Leishmania infantum/growth & development , Leishmania infantum/immunology , Leishmaniasis, Visceral/parasitology , Male , Mice , Mice, Inbred BALB C , Transforming Growth Factor beta/metabolismABSTRACT
Infection of BALB/c mice with Leishmania chagasi results in progressive increase of parasite burden in spleen, in spite of extensive T cell activation in situ. Explanted splenic CD4+ T cells showed decreased proliferation to anti-CD3, compared with controls, and no response to L. chagasi recombinant antigen Lcr1. Blockade of the negative costimulatory receptor CTLA-4 restored responses to anti-CD3 and induced vigorous responses to Lcr1. Blockade of B7-1, but not B7-2, also enhanced T cell responsiveness. CTLA-4 blockade completely restored activation-induced interleukin-2 secretion and increased interferon-gamma production. The effect, however, was not restricted to Th1 responses, since CTLA-4 blockade also enhanced antigen-induced interleukin-4 secretion. CTLA-4 blockade induced almost complete elimination of parasite burden in splenocyte cultures activated with anti-CD3 or Lcr1. These results indicate that CTLA-4 engagement by B7-1 plays an important role in maintaining unresponsiveness in CD4+ T cells in this model of chronic visceral leishmaniasis.
