Leishmania donovani Nucleoside Hydrolase (NH36) Domains Induce T-Cell Cytokine Responses in Human Visceral Leishmaniasis

Abstract

Development of immunoprotection against visceral leishmaniasis (VL) focused on the identification of antigens capable of inducing a Th1 immune response. Alternatively, antigens targeting the CD8 and T-regulatory responses are also relevant in VL pathogenesis and worthy of being included in a preventive human vaccine. We assessed in active and cured patients and VL asymptomatic subjects the clinical signs and cytokine responses to the <i>Leishmania donovani</i> nucleoside hydrolase NH36 antigen and its N-(F1), central (F2) and C-terminal (F3) domains. As markers of VL resistance, the F2 induced the highest levels of IFN-γ, IL-1β, and TNF-α and, together with F1, the strongest secretion of IL-17, IL-6, and IL-10 in DTH⁺ and cured subjects. F2 also promoted the highest frequencies of CD3⁺CD4⁺IL-2⁺TNF-α^-IFN-γ^-, CD3⁺CD4⁺IL-2⁺TNF-α⁺IFN-γ^-, CD3⁺CD4⁺IL-2⁺TNF-α^-IFN-γ⁺, and CD3⁺CD4⁺IL-2⁺TNF-α⁺IFN-γ⁺ T cells in cured and asymptomatic subjects. Consistent with this, the IFN-γ increase was correlated with decreased spleen (<i>R</i> = -0.428, <i>P</i> = 0.05) and liver sizes (<i>R</i> = -0.428, <i>P</i> = 0.05) and with increased hematocrit counts (<i>R</i> = 0.532, <i>P</i> = 0.015) in response to F1 domain, and with increased hematocrit (<i>R</i> = 0.512, <i>P</i> 0.02) and hemoglobin counts (<i>R</i> = 0.434, <i>P</i> = 0.05) in response to F2. Additionally, IL-17 increases were associated with decreased spleen and liver sizes in response to F1 (<i>R</i> = -0.595, <i>P</i> = 0.005) and F2 (<i>R</i> = -0.462, <i>P</i> = 0.04). Conversely, F1 and F3 increased the CD3⁺CD8⁺IL-2⁺TNF-α^-IFN-γ^-, CD3⁺CD8⁺IL-2⁺TNF-α⁺IFN-γ^-, and CD3⁺CD8⁺IL-2⁺TNF-α⁺IFN-γ⁺ T cell frequencies of VL patients correlated with increased spleen and liver sizes and decreased hemoglobin and hematocrit values. Therefore, cure and acquired resistance to VL correlate with the CD4⁺-Th1 and Th-17 T-cell responses to F2 and F1 domains. Clinical VL outcomes, by contrast, correlate with CD8⁺ T-cell responses against F3 and F1, potentially involved in control of the early infection. The <i>in silico</i>-predicted NH36 epitopes are conserved and bind to many HL-DR and HLA and B allotypes. No human vaccine against <i>Leishmania</i> is available thus far. In this investigation, we identified the NH36 domains and epitopes that induce CD4⁺ and CD8⁺ T cell responses, which could be used to potentiate a human universal T-epitope vaccine against leishmaniasis.