RESUMEN
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.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Leishmania mexicana/inmunología , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/mortalidad , Receptores de Superficie Celular/deficiencia , Receptores de Superficie Celular/inmunología , Animales , Femenino , Interleucina-13/inmunología , Interleucina-4/inmunología , Leishmania mexicana/patogenicidad , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Factores SexualesRESUMEN
In this study we generated a novel dual specific phosphatase 4 (DUSP4) deletion mouse using a targeted deletion strategy in order to examine the role of MAP kinase phosphatase-2 (MKP-2) in immune responses. Lipopolysaccharide (LPS) induced a rapid, time and concentration-dependent increase in MKP-2 protein expression in bone marrow-derived macrophages from MKP-2(+/+) but not from MKP-2(-/-) mice. LPS-induced JNK and p38 MAP kinase phosphorylation was significantly increased and prolonged in MKP-2(-/-) macrophages whilst ERK phosphorylation was unaffected. MKP-2 deletion also potentiated LPS-stimulated induction of the inflammatory cytokines, IL-6, IL-12p40, TNF-α, and also COX-2 derived PGE(2) production. However surprisingly, in MKP-2(-/-) macrophages, there was a marked reduction in LPS or IFNγ-induced iNOS and nitric oxide release and enhanced basal expression of arginase-1, suggesting that MKP-2 may have an additional regulatory function significant in pathogen-mediated immunity. Indeed, following infection with the intracellular parasite Leishmania mexicana, MKP-2(-/-) mice displayed increased lesion size and parasite burden, and a significantly modified Th1/Th2 bias compared with wild-type counterparts. However, there was no intrinsic defect in MKP-2(-/-) T cell function as measured by anti-CD3 induced IFN-γ production. Rather, MKP-2(-/-) bone marrow-derived macrophages were found to be inherently more susceptible to infection with Leishmania mexicana, an effect reversed following treatment with the arginase inhibitor nor-NOHA. These findings show for the first time a role for MKP-2 in vivo and demonstrate that MKP-2 may be essential in orchestrating protection against intracellular infection at the level of the macrophage.
Asunto(s)
Citocinas/metabolismo , Mediadores de Inflamación/metabolismo , Leishmania mexicana/patogenicidad , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/prevención & control , Macrófagos/inmunología , Proteínas Tirosina Fosfatasas/fisiología , Animales , Arginasa/metabolismo , Western Blotting , Médula Ósea/inmunología , Médula Ósea/metabolismo , Médula Ósea/patología , Células Cultivadas , Femenino , Leishmaniasis Cutánea/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/metabolismo , Macrófagos/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Óxido Nítrico/metabolismo , Fosforilación , ARN Mensajero/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células TH1/inmunología , Células TH1/metabolismo , Células Th2/inmunología , Células Th2/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Leishmania mexicana cysteine peptidases (CPs) have been identified as important parasite virulence factors. More recently, a natural inhibitor of CPs (ICP) from L. mexicana has been characterized, and ICP mutants have been created. Infection of BALB/c mice with ICP null mutants or ICP reexpressing mutants resulted in nonhealing, progressively growing lesions albeit slightly attenuated compared with the growth of lesions produced by wild-type parasites. In contrast, BALB/c mice infected with mutants overexpressing ICP were able to significantly control lesion growth or heal. While BALB/c mice infected with wild-type parasites, ICP null mutants, or ICP reexpressing mutants produced significant antibody responses, including immunoglobulin E (IgE), no Th1 response, as indicated by antigen-induced splenocyte gamma interferon (IFN-gamma) production, could be demonstrated. In contrast, BALB/c mice infected with mutants overexpressing ICP produced significantly less antibody, particularly IgE, as well as significantly reduced splenocyte interleukin-4 and enhanced IFN-gamma production. BALB/c mice were able to resolve infection following infection with one ICP overexpressing clone, which was subsequently used for vaccination studies with BALB/c mice. However, no protection was afforded these mice when they were challenged with wild-type parasites. Nevertheless, two other mouse strains susceptible to L. mexicana, C3H and C57BL/6, vaccinated with overexpressing ICP mutants were able to control challenge infection associated with an enhanced Th1 response. This study confirms that L. mexicana CPs are virulence factors and that ICPs have therapeutic potential.
Asunto(s)
Cisteína Endopeptidasas/metabolismo , Inhibidores de Cisteína Proteinasa/biosíntesis , Leishmania mexicana/inmunología , Leishmania mexicana/patogenicidad , Leishmaniasis Cutánea/patología , Células TH1/inmunología , Factores de Virulencia/antagonistas & inhibidores , Animales , Anticuerpos Antiprotozoarios/sangre , Inhibidores de Cisteína Proteinasa/genética , Femenino , Eliminación de Gen , Prueba de Complementación Genética , Inmunoglobulina E/sangre , Interferón gamma/metabolismo , Leishmaniasis Cutánea/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Índice de Severidad de la Enfermedad , Bazo/inmunología , VirulenciaRESUMEN
A comparison of the growth of Leishmania mexicana in IL-4(-/-), IL-4Ralpha(-/-) and wild-type BALB/c mice demonstrated a disease exacerbative role for IL-13 as well as IL-4. Thus, while both IL-4(-/-) and IL-4Ralpha(-/-) mice were more resistant than wild-type controls to infection with L. mexicana, IL-4Ralpha(-/-) mice, which are unresponsive to IL-13 as well as IL-4, were significantly more resistant to parasite growth than their IL-4(-/-) counterparts. Cytokine and antibody analysis revealed a Th1-biased specific response in both infected IL-4(-/-) and IL-4Ralpha(-/-) mice compared with wild-type animals. Reconstituting SCID mice with IL-4(-/-), IL-4Ralpha(-/-) or wild-type splenocytes prior to infection demonstrated that the early onset of lesion growth was dependent on the presence of lymphocytes responding to IL-4 and/or IL-13, as lesions failed to develop in only the SCID IL-4Ralpha(-/-) reconstituted mice. An independent role for IL-13 in L. mexicana infection was demonstrated by comparing disease progression in IL-13(-/-), IL-4(-/-)/IL-13(-/-) and wild-type B6/129 mice. In contrast to IL-4(-/-)/IL-13(-/-) mice, which were resistant, IL-13(-/-) mice developed lesions similar in size to wild-type animals up to week 8 post-infection. However, in contrast to wild-type mice in which disease continued to progress, lesions eventually healed in IL-13(-/-) mice, in association with the development of a Th1 response. Collectively our results suggest that IL-4 plays a critical role in early lesion development, and that IL-13 plays a crucial part in maintaining a chronic non-healing infection.