Non-canonical NLRP3 inflammasome activation and IL-1β signaling are necessary to L. amazonensis control mediated by P2X7 receptor and leukotriene B4

Abstract

Leishmaniasis is a neglected tropical disease affecting millions of individuals worldwide. P2X7 receptor has been linked to the elimination of Leishmania amazonensis . Biological responses evoked by P2X7 receptor activation have been well-documented, including apoptosis, phagocytosis, cytokine release, such as IL-1β. It was demonstrated that NLRP3 inflammasome activation and IL-1β signaling participated in resistance against L . amazonensis . Furthermore, our group has shown that L . amazonensis elimination through P2X7 receptor activation depended on leukotriene B 4 (LTB 4 ) production and release. Therefore, we investigated whether L . amazonensis elimination by P2X7 receptor and LTB 4 involved NLRP3 inflammasome activation and IL-1β signaling. We showed that macrophages from NLRP3 -/- , ASC -/- , Casp-1/11 -/- , gp91 phox-/- , and IL-1R -/- mice treated with ATP or LTB 4 did not decrease parasitic load as was observed in WT mice. When ASC -/- macrophages were treated with exogenous IL-1β, parasite killing was noted, however, we did not see parasitic load reduction in IL-1R -/- macrophages. Similarly, macrophages from P2X7 receptor-deficient mice treated with IL-1β also showed decreased parasitic load. In addition, when we infected Casp-11 -/- macrophages, neither ATP nor LTB 4 were able to reduce parasitic load, and Casp-11 -/- mice were more susceptible to L . amazonensis infection than were WT mice. Furthermore, P2X7 -/- L . amazonensis- infected mice locally treated with exogenous LTB 4 showed resistance to infection, characterized by lower parasite load and smaller lesions compared to untreated P2X7 -/- mice. A similar observation was noted when infected P2X7 -/- mice were treated with IL-1β, i.e., lower parasite load and smaller lesions compared to P2X7 -/- mice. These data suggested that L . amazonensis elimination mediated by P2X7 receptor and LTB 4 was dependent on non-canonical NLRP3 inflammasome activation, ROS production, and IL-1β signaling.

Reconciling protective and pathogenic roles of the NLRP3 inflammasome in leishmaniasis

“…The protective role of caspase‐11 is also evident in BMDM, as Casp11 −/− BMDM failed to restrict L major , L braziliensis, and L amazonensis growth in vitro. Another study demonstrated a similar protective role for caspase‐11 following footpad injection with L amazonensis, where Casp11 −/− mice presented with significantly larger footpad lesions and higher parasite burden compared to WT mice 82 …”

“…More recently, ATP‐induced leukotriene B4 (LTB4) has been suggested to activate caspase‐11‐mediated non‐canonical NLRP3 inflammasome during L amazonensis infection 82,83 (Figure 2). It should be noted, however, that Chaves et al used exogenous ATP or LTB4 during L amazonensis infection to activate the non‐canonical NLRP3 inflammasome 82 .…”

“…More recently, ATP‐induced leukotriene B4 (LTB4) has been suggested to activate caspase‐11‐mediated non‐canonical NLRP3 inflammasome during L amazonensis infection 82,83 (Figure 2). It should be noted, however, that Chaves et al used exogenous ATP or LTB4 during L amazonensis infection to activate the non‐canonical NLRP3 inflammasome 82 . Mechanistically, ATP signals via P2X7 receptor to induce LTB4 production in L amazonensis ‐infected peritoneal macrophages in vitro, demonstrating the importance of ATP‐P2X7 receptor signaling in the production of LTB4 83 .…”

“…Mechanistically, ATP signals via P2X7 receptor to induce LTB4 production in L amazonensis ‐infected peritoneal macrophages in vitro, demonstrating the importance of ATP‐P2X7 receptor signaling in the production of LTB4 83 . LTB4 then engages caspase‐11 and promotes activation of the non‐canonical NLRP3 inflammasome via a yet‐unknown mechanism 82 . Given that these studies used exogenous addition of ATP or LTB4 to engage caspase‐11 pathway, whether endogenous ATP and/or LTB4 are generated during L amazonensis infection and how they might contribute to caspase‐11 activation and non‐canonical NLRP3 inflammasome is not clear.…”

“…In reviewing all known articles that show a protective role for the NLRP3 inflammasome, 56,64,76,82,88,89 we noted that these studies exclusively use C57BL/6 mice, and NLRP3 inflammasome‐deficient mice backcrossed to C57BL/6 as mammalian hosts. Regarding Leishmania spp., these studies mostly use new‐world Leishmania spp.…”