NOD-Like Receptor P3 Inflammasome Controls Protective Th1/Th17 Immunity against Pulmonary Paracoccidioidomycosis

The NOD-Like Receptor P3 (NLRP3) inflammasome is an intracellular multimeric complex that triggers the activation of inflammatory caspases and the maturation of IL-1β and IL-18, important cytokines in the innate immune response against pathogens. The functional NLRP3 inflammasome complex consists of NLRP3, the adaptor protein ASC and caspase-1. Various molecular mechanisms are associated with NLRP3 activation, including the presence of extracellular ATP, which is recognized by the cell surface P2X7 receptor (P2X7R). Several pattern recognition receptors (PRRs) on innate immune cells recognize Paracoccidioides brasiliensis components, resulting in diverse responses that influence adaptive immunity and disease outcomes. However, the role of the NLRP3 inflammasome in pulmonary paracoccidioidomycosis is not well investigated, which led us to use an intratracheal (i.t.) model of infection to study the influence of this receptor on anti-fungal immunity and on the severity of infection. For in vivo studies, C57BL/6 mice deficient in several NLRP3 inflammasome components (Nlrp3-/-, Casp1/11-/-, Asc-/-), as well as ATP receptor-deficient mice (P2x7r-/-), were infected i.t. with P. brasiliensis, and several parameters of immunity and disease severity were analyzed during the acute and chronic periods of infection. Pulmonary paracoccidioidomycosis was more severe in Nlrp3-/-, Casp1/11-/-, Asc-/- and P2x7r-/- mice, as demonstrated by increased fungal burdens, tissue pathology and mortality rates. The more severe disease developed by NLRP3-, ASC- and Caspase-1/11-deficient mice was associated with decreased production of IL-1β and IL-18 and reduced inflammatory reactions mediated by PMN leukocytes and activated CD4+ and CD8+ T cells. The decrease in T cell immunity occurred concomitantly with the increased expansion of CD4+CD25+Foxp3 regulatory T (Treg) cells. The characterization of intracellular cytokines showed a persistent reduction in CD4+ and CD8+ T cells expressing IFN-y and IL-17, whereas the frequencies of cells producing IL-4 and TGF-β increased. Histopathological studies showed that all deficient mouse strains developed more severe lesions containing elevated numbers of fungal cells, resulting in increased mortality rates. Collectively, these findings led us to conclude that the activation of the NLRP3 inflammasome has a crucial role in immunoprotection against pulmonary paracoccidioidomycosis by promoting the expansion of Th1/Th17 immunity and reducing the suppressive control mediated by Treg cells.