RESUMO
We have demonstrated recently that the glycoinositolphospholipid (GIPL) molecule from the protozoan Trypanosoma cruzi is a TLR4 agonist with proinflammatory effects. Here, we show that GIPL-induced neutrophil recruitment into the peritoneal cavity is mediated by at least two pathways: one, where IL-1beta acts downstream of TNF-alpha, and a second, which is IL-1beta- and TNFRI-independent. Moreover, NKT cells participate in this proinflammatory cascade, as in GIPL-treated CD1d(-/-) mice, TNF-alpha and MIP-2 levels are reduced significantly. As a consequence of this inflammatory response, spleen and lymph nodes of GIPL-treated mice have an increase in the percentage of T and B cells expressing the CD69 activation marker. Cell-transfer experiments demonstrate that T and B cell activation by GIPL is an indirect effect, which relies on the expression of TLR4 by other cell types. Moreover, although signaling through TNFRI contributes to the activation of B and gammadelta+ T cells, it is not required for increasing CD69 expression on alphabeta+ T lymphocytes. It is interesting that T cells are also functionally affected by GIPL treatment, as spleen cells from GIPL-injected mice show enhanced production of IL-4 following in vitro stimulation by anti-CD3. Together, these results contribute to the understanding of the inflammatory properties of the GIPL molecule, pointing to its potential role as a parasite-derived modulator of the immune response during T. cruzi infection.
Assuntos
Glicolipídeos/fisiologia , Mediadores da Inflamação/fisiologia , Fosfolipídeos/fisiologia , Receptor 4 Toll-Like/metabolismo , Trypanosoma cruzi/imunologia , Animais , Antígenos CD1/genética , Antígenos CD1/fisiologia , Antígenos CD1d , Quimiocina CXCL2 , Quimiocinas/metabolismo , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Glicolipídeos/administração & dosagem , Glicolipídeos/farmacologia , Imunidade Inata/genética , Interleucina-1beta/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/genética , Infiltração de Neutrófilos/imunologia , Fosfolipídeos/administração & dosagem , Fosfolipídeos/farmacologia , RNA Mensageiro/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores Tipo I de Fatores de Necrose Tumoral/fisiologia , Linfócitos T/metabolismo , Receptor 4 Toll-Like/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
TLRs function as pattern recognition receptors in mammals and play an essential role in the recognition of microbial components. We found that the injection of glycoinositolphospholipids (GIPLs) from Trypanosoma cruzi into the peritoneal cavity of mice induced neutrophil recruitment in a TLR4-dependent manner: the injection of GIPL in the TLR4-deficient strain of mice (C57BL/10ScCr) caused no inflammatory response. In contrast, in TLR2 knockout mice, neutrophil chemoattraction did not differ significantly from that seen in wild-type controls. GIPL-induced neutrophil attraction and MIP-2 production were also severely affected in TLR4-mutant C3H/HeJ mice. The role of TLR4 was confirmed in vitro by testing genetically engineered mutants derived from TLR2-deficient Chinese hamster ovary (CHO)-K1 fibroblasts that were transfected with CD14 (CHO/CD14). Wild-type CHO/CD14 cells express the hamster TLR4 molecule and the mutant line, in addition, expresses a nonfunctional form of MD-2. In comparison to wild-type cells, mutant CHO/CD14 cells failed to respond to GIPLs, indicating a necessity for a functional TLR4/MD-2 complex in GIPL-induced NF-kappaB activation. Finally, we found that TLR4-mutant mice were hypersusceptible to T. cruzi infection, as evidenced by a higher parasitemia and earlier mortality. These results demonstrate that natural resistance to T. cruzi is TLR4 dependent, most likely due to TLR4 recognition of their GIPLs.