RESUMEN
The effects of Leishmania mexicana metacyclic promastigotes upon MAP kinase signalling in mouse bone marrow macrophages and subsequent expression of the disease regulatory proteins iNOS and COX-2 were studied. At a ratio of 5:1, promastigotes caused a marked increase in phosphorylation of the three major MAP kinases, ERK, p38 and JNK. MAP kinase signalling was substantially reduced in TLR-4(-/-) but not TLR-2(-/-) deficient macrophages and completely abolished in double TLR-2/4(-/-) macrophages. A similar outcome was observed using cysteine peptidase B deficient amastigotes. Furthermore, whilst promastigotes had no independent effect on iNOS or COX-2 expression, they prolonged the induction of these proteins stimulated by LPS and enhanced PGE(2) and NO production. Induction of COX-2 and iNOS was also TLR-4 dependent. Blockade of either PGE(2) or NO production with indomethacin or l-NAME reversed promastigote inhibition of LPS induced IL-12 production. Promastigotes also increased macrophage arginase-1 expression and enhanced arginase activity, both of which were substantially reduced in TLR-4 but not TLR-2 deficient macrophages. Surprisingly, arginase inhibition by Nor-NOHA also caused a reversal of promastigote mediated inhibition of macrophage IL-12 production. These data demonstrate for the first time the role of TLR-4 in mediating the effects of L. mexicana promastigotes on MAP kinase activation, up-regulation of COX-2, iNOS as well as arginase-1 expression in macrophages and further shows that PGE(2), NO and arginase activity all contribute substantially to the inhibition of host cell IL-12 production.
Asunto(s)
Activación Enzimática/inmunología , Interleucina-12/biosíntesis , Leishmaniasis/inmunología , Macrófagos/inmunología , Receptor Toll-Like 4/metabolismo , Animales , Arginasa/biosíntesis , Arginasa/inmunología , Western Blotting , Ciclooxigenasa 2/biosíntesis , Ciclooxigenasa 2/inmunología , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Interleucina-12/inmunología , Leishmania mexicana/inmunología , Leishmaniasis/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiología , Ratones , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Óxido Nítrico Sintasa de Tipo II/inmunología , Transducción de Señal/inmunología , Receptor Toll-Like 4/inmunologíaRESUMEN
Immunologically intact BALB/c mice infected with Leishmania mexicana develop non-healing progressively growing lesions associated with a biased Th2 response while similarly infected IL-4Rα-deficient mice fail to develop lesions and develop a robust Th1 response. In order to determine the functional target(s) for IL-4/IL-13 inducing non-healing disease, the course of L. mexicana infection was monitored in mice lacking IL-4Rα expression in specific cellular compartments. A deficiency of IL-4Rα expression on macrophages/neutrophils (in LysM(cre)IL-4Rα(-/lox) animals) had minimal effect on the outcome of L. mexicana infection compared with control (IL-4Rα(-/flox)) mice. In contrast, CD4(+) T cell specific (Lck(cre)IL-4Rα(-/lox)) IL-4Rα(-/-) mice infected with L. mexicana developed small lesions, which subsequently healed in female mice, but persisted in adult male mice. While a strong Th1 response was manifest in both male and female CD4(+) T cell specific IL-4Rα(-/-) mice infected with L. mexicana, induction of IL-4 was manifest in males but not females, independently of CD4(+) T cell IL-4 responsiveness. Similar results were obtained using pan-T cell specific (iLck(cre)IL-4Rα(-/lox)) IL-4Rα(-/-) mice. Collectively these data demonstrate that upon infection with L. mexicana, initial lesion growth in BALB/c mice is dependent on non-T cell population(s) responsive to IL-4/IL-13 while progressive infection is dependent on CD4(+) T cells responsive to IL-4.