RESUMO
BACKGROUND: Although eosinophils co-express multiple integrin receptors, the contributions of integrins to eosinophil development have not been explored. We previously described extensive aggregation and cytological immaturity in eosinophils developing in bone-marrow (BM) cultures exposed to dexamethasone. Here we examined the relationship of alpha 4 integrins with these effects of dexamethasone. OBJECTIVES: We evaluated: (a) the effects of exposure to dexamethasone in BM culture on eosinophil expression of alpha 4 integrin receptors and ligands; (b) the contribution of alpha 4 integrins to eosinophil aggregation and maturation. METHODS: Cultures were established with IL-5 (alone or with dexamethasone) for up to 7 days, and eosinophil production, alpha 4 integrin receptor/ligand expression, aggregation and morphology were evaluated before and after targeting alpha 4 integrin-dependent adhesions. Because prostaglandin E2 (PGE2) modifies the effects of dexamethasone on eosinophilopoiesis, PGE2 effects on alpha 4 integrin expression and function were also evaluated. RESULTS: Dexamethasone increased the yield of eosinophils up to day 7. The frequency of eosinophils expressing alpha 4, beta1 and beta 7 integrin receptors at day 7 was also increased by dexamethasone. Eosinophils also expressed the alpha 4 beta 1 ligand, VCAM-1. Dexamethasone increased the expression of alpha 4 integrin and VCAM-1 in aggregates containing eosinophils as early as day 3. PGE2, added up to day 3, modified the effects of dexamethasone to suppress the expression of alpha 4 integrin, decrease aggregation and promote cytological maturation of eosinophils recovered at day 7. Dissociation of immature eosinophils from clusters present at day 3 by reagents targeting alpha 4 or beta1 integrins or VCAM-1 also induced cytological maturation. The concordant effects of targeting alpha 4 integrins with drugs and antibodies support a relationship between alpha 4-mediated aggregation and maturational arrest. CONCLUSIONS: These observations support a novel role for alpha 4 integrin receptors and ligands in eosinophilopoiesis. In addition, increased alpha 4 expression following glucocorticoid exposure may contribute to the retention and accumulation of eosinophils in haemopoietic tissue.
Assuntos
Células da Medula Óssea/efeitos dos fármacos , Dexametasona/farmacologia , Eosinófilos/efeitos dos fármacos , Eosinófilos/imunologia , Integrina alfa4/imunologia , Animais , Células da Medula Óssea/imunologia , Células Cultivadas , Eosinófilos/citologia , Integrina alfa4/efeitos dos fármacos , Integrina alfa4beta1/biossíntese , Integrina alfa4beta1/efeitos dos fármacos , Interleucina-5/farmacologia , Ligantes , Camundongos , Camundongos Endogâmicos BALB C , Receptores Imunológicos/biossíntese , Receptores Imunológicos/efeitos dos fármacos , Molécula 1 de Adesão de Célula Vascular/biossíntese , Molécula 1 de Adesão de Célula Vascular/efeitos dos fármacosRESUMO
The comprehension of the pathogenesis of Trypanosoma cruzi-elicited myocarditis is crucial to delineate new therapeutic strategies aiming to ameliorate the inflammation that leads to heart dysfunction, without hampering parasite control. The augmented expression of CCL5/RANTES and CCL3/MIP-1alpha, and their receptor CCR5, in the heart of T. cruzi-infected mice suggests a role for CC-chemokines and their receptors in the pathogenesis of T. cruzi-elicited myocarditis. Herein, we discuss our recent results using a CC-chemokine receptor inhibitor (Met-RANTES), showing the participation of CC-chemokines in T. cruzi infection and unraveling CC-chemokine receptors as an attractive therapeutic target for further evaluation in Chagas disease.
Assuntos
Cardiomiopatia Chagásica/tratamento farmacológico , Quimiocina CCL5/análogos & derivados , Quimiocinas CC/metabolismo , Miocardite/tratamento farmacológico , Receptores de Quimiocinas/antagonistas & inibidores , Trypanosoma cruzi , Animais , Relação CD4-CD8 , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Cardiomiopatia Chagásica/imunologia , Cardiomiopatia Chagásica/metabolismo , Quimiocina CCL5/uso terapêutico , Quimiotaxia de Leucócito/imunologia , Camundongos , Camundongos Endogâmicos C3H , Miocardite/imunologia , Miocardite/metabolismo , Miocardite/parasitologia , Trypanosoma cruzi/imunologiaRESUMO
The comprehension of the pathogenesis of Trypanosoma cruzi-elicited myocarditis is crucial to delineate new therapeutic strategies aiming to ameliorate the inflammation that leads to heart dysfunction, without hampering parasite control. The augmented expression of CCL5/RANTES and CCL3/MIP-1alpha, and their receptor CCR5, in the heart of T. cruzi-infected mice suggests a role for CC-chemokines and their receptors in the pathogenesis of T. cruzi-elicited myocarditis. Herein, we discuss our recent results using a CC-chemokine receptor inhibitor (Met-RANTES), showing the participation of CC-chemokines in T. cruzi infection and unraveling CC-chemokine receptors as an attractive therapeutic target for further evaluation in Chagas disease.
Assuntos
Animais , Camundongos , Cardiomiopatia Chagásica/tratamento farmacológico , /análogos & derivados , Quimiocinas CC/metabolismo , Miocardite/tratamento farmacológico , Receptores de Quimiocinas/antagonistas & inibidores , Trypanosoma cruzi , /imunologia , /imunologia , Cardiomiopatia Chagásica/imunologia , Cardiomiopatia Chagásica/metabolismo , /uso terapêutico , Quimiotaxia de Leucócito/imunologia , Miocardite/imunologia , Miocardite/metabolismo , Miocardite/parasitologia , Trypanosoma cruzi/imunologiaRESUMO
Several lines of evidence have shown that Trypanosoma cruzi interacts with host extracellular matrix (ECM) components producing breakdown products that play an important role in parasite mobilization and infectivity. Parasite-released antigens also modulate ECM expression that could participate in cell-cell and/or cell-parasite interactions. Increased expression of ECM components has been described in the cardiac tissue of chronic chagasic patients and diverse target tissues including heart, thymus, central nervous system and skeletal muscle of experimentally T. cruzi-infected mice. ECM components may adsorb parasite antigens and cytokines that could contribute to the establishment and perpetuation of inflammation. Furthermore, T. cruzi-infected mammalian cells produce cytokines and chemokines that not only participate in the control of parasitism but also contribute to the establishment of chronic inflammatory lesions in several target tissues and most frequently lead to severe myocarditis. T. cruzi-driven cytokines and chemokines may also modulate VCAM-1 and ICAM-1 adhesion molecules on endothelial cells of target tissues and play a key role in cell recruitment, especially of activated VLA-4+LFA-1+CD8+ T lymphocytes, resulting in a predominance of this cell population in the inflamed heart, central nervous system and skeletal muscle. The VLA-4+-invading cells are surrounded by a fine network of fibronectin that could contribute to cell anchorage, activation and effector functions. Since persistent "danger signals" triggered by the parasite and its antigens are required for the establishment of inflammation and ECM alterations, therapeutic interventions that control parasitism and selectively modulate cell migration improve ECM abnormalities, paving the way for the development of new therapeutic strategies improving the prognosis of T. cruzi-infected individuals
Assuntos
Animais , Humanos , Camundongos , Moléculas de Adesão Celular , Cardiomiopatia Chagásica , Matriz Extracelular , Proteínas Quimioatraentes de Monócitos , Cardiomiopatia Chagásica , Doença Crônica , Matriz Extracelular , Interações Hospedeiro-Parasita , Índice de Gravidade de DoençaRESUMO
Several lines of evidence have shown that Trypanosoma cruzi interacts with host extracellular matrix (ECM) components producing breakdown products that play an important role in parasite mobilization and infectivity. Parasite-released antigens also modulate ECM expression that could participate in cell-cell and/or cell-parasite interactions. Increased expression of ECM components has been described in the cardiac tissue of chronic chagasic patients and diverse target tissues including heart, thymus, central nervous system and skeletal muscle of experimentally T. cruzi-infected mice. ECM components may adsorb parasite antigens and cytokines that could contribute to the establishment and perpetuation of inflammation. Furthermore, T. cruzi-infected mammalian cells produce cytokines and chemokines that not only participate in the control of parasitism but also contribute to the establishment of chronic inflammatory lesions in several target tissues and most frequently lead to severe myocarditis. T. cruzi-driven cytokines and chemokines may also modulate VCAM-1 and ICAM-1 adhesion molecules on endothelial cells of target tissues and play a key role in cell recruitment, especially of activated VLA-4+LFA-1+CD8+ T lymphocytes, resulting in a predominance of this cell population in the inflamed heart, central nervous system and skeletal muscle. The VLA-4+-invading cells are surrounded by a fine network of fibronectin that could contribute to cell anchorage, activation and effector functions. Since persistent "danger signals" triggered by the parasite and its antigens are required for the establishment of inflammation and ECM alterations, therapeutic interventions that control parasitism and selectively modulate cell migration improve ECM abnormalities, paving the way for the development of new therapeutic strategies improving the prognosis of T. cruzi-infected individuals.
Assuntos
Moléculas de Adesão Celular/fisiologia , Cardiomiopatia Chagásica/parasitologia , Fatores Quimiotáticos/fisiologia , Matriz Extracelular/parasitologia , Trypanosoma cruzi/fisiologia , Animais , Cardiomiopatia Chagásica/patologia , Doença Crônica , Proteínas da Matriz Extracelular/metabolismo , Interações Hospedeiro-Parasita , Humanos , Inflamação/metabolismo , Inflamação/parasitologia , Camundongos , Índice de Gravidade de DoençaRESUMO
It has been proposed that antigens released by Trypanosoma cruzi sensitize vertebrate cells leading to their destruction by the immune response raised against the parasite. Here, we characterized antigens released by trypomastigotes of T. cruzi that bind to non-infected cells and investigated biological consequences of this adsorption. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of antigens released by [(35)S]-methionine-labeled parasites revealed the presence of polypeptides mainly ranging from 85 to 170 kDa that were specifically recognized by sera from chronically T. cruzi infected rabbits. Polypeptides of 85-110 and 160-170 kDa bound to non-infected epithelial, fibroblast and muscle mammalian cell lines, which thus became targets for anti-T. cruzi antibody binding. Cysteine-proteinase, but not trans-sialidase, was detected among the cell-bound antigens, and purified cysteine-proteinase was adsorbed to non-infected cells. Immunoelectron microscopic studies showed that parasite antigens were mainly released as membrane vesicles that adhered to membrane microvilli and were internalized by mammalian cells. We provide evidence that adsorption of parasite antigens induced an increase in expression of extracellular matrix (ECM) components (fibronectin, laminin and type I collagen) by sensitized cells. Thus, our data reinforce the idea that in vivo T. cruzi released antigens might be involved in the establishment of inflammation, sensitizing non-infected host cells and triggering an immune response against parasite antigens. Further, our data showed that antigen sensitization modulates biological cell functions as ECM expression that could mediate cell-cell or parasite-host cell interactions, contributing to the establishment of inflammation.
Assuntos
Antígenos de Protozoários/imunologia , Antígenos de Protozoários/metabolismo , Antígenos de Superfície/imunologia , Matriz Extracelular/metabolismo , Trypanosoma cruzi/imunologia , Glicoproteínas Variantes de Superfície de Trypanosoma , Adsorção , Animais , Células Cultivadas , Eletroforese em Gel de Poliacrilamida , Matriz Extracelular/imunologiaRESUMO
The role of cytokines in the control of tissue parasitism and pathogenesis of experimental Chagas' disease was investigated. Wild-type and different cytokine as well as inducible nitric oxide synthase (iNOS) knockout mice were infected with the Colombian strain of Trypanosoma cruzi, and the kinetics of tissue parasitism, inflammatory reaction, parasitemia, and mortality were determined. We demonstrate the pivotal role of the interleukin (IL)-12/interferon (IFN)-gamma/iNOS axis and the antagonistic effect of IL-4 in controlling heart tissue parasitism, inflammation, and host resistance to acute infection with T. cruzi. Further, the heart and central nervous system were shown the main sites of reactivation of T. cruzi infection in mice lacking functional genes for IFN-gamma and IL-12, respectively. Our results also show that in contrast to IFN-gamma knockout (KO) mice, splenocytes from IL-12 KO mice infected with T. cruzi produced low levels of IFN-gamma upon stimulation with antigen. Consistently, high levels of anti-T. cruzi IgG2a antibodies were detected in the sera from IL-12 KO, but not from IFN-gamma KO mice, infected with the Colombian strain of T. cruzi. Thus, our results suggest that the level of IFN-gamma deficiency is a major determinant of the site of reactivation of T. cruzi infection in immunocompromised host.
Assuntos
Doenças do Sistema Nervoso Central/patologia , Doenças do Sistema Nervoso Central/parasitologia , Cardiomiopatia Chagásica/patologia , Cardiomiopatia Chagásica/parasitologia , Doença de Chagas/patologia , Doença de Chagas/parasitologia , Interferon gama/fisiologia , Interleucina-12/fisiologia , Animais , Sistema Nervoso Central/parasitologia , Sistema Nervoso Central/patologia , Suscetibilidade a Doenças , Feminino , Coração/parasitologia , Interferon gama/deficiência , Interferon gama/genética , Interleucina-12/deficiência , Interleucina-12/genética , Interleucina-4/deficiência , Interleucina-4/genética , Interleucina-4/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout/genética , Miocardite/parasitologia , Miocardite/patologia , Miocárdio/patologia , Óxido Nítrico Sintase/deficiência , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase/fisiologia , Óxido Nítrico Sintase Tipo II , RecidivaRESUMO
The determinants of the prevalence of CD8(+) T cells in the inflamed myocardium of Trypanosoma cruzi-infected patients and experimental animals are undefined. Using C3H/He mice infected with the Colombiana strain of T. cruzi, we found that the distribution of CD4(+)/CD8(-) and CD4(-)/CD8(+) T cells in the myocardium mirrors the frequency of cells expressing the CD62L(Low)LFA-1(High)VLA-4(High) activation phenotype among CD4(+)/CD8(-) and CD4(-)/CD8(+ )peripheral blood T cells. Consistently, vascular cell adhesion molecule-1-positive endothelial cells and a fine fibronectin network surrounding VLA-4(+) mononuclear cells were found in the inflamed myocardium. Further, interferon gamma (IFN-gamma) and IFN-gamma-induced chemokines (RANTES, MIG and CRG-2/IP-10), as well as JE/MCP-1 and MIP1-alpha, were found to be the dominant cytokines expressed in situ during acute and chronic myocarditis elicited by T. cruzi. In contrast, interleukin 4 mRNA was only detected during the chronic phase. Altogether, the results indicate that the distribution of T-cell subsets in the myocardium of T. cruzi-infected mice reflects the particular profile of adhesion molecules acquired by most peripheral CD8(+) T lymphocytes and point to the possibility that multiple IFN-gamma-inducible molecules present in the inflamed tissue contribute to the establishment and maintenance of T. cruzi-induced myocarditis.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Cardiomiopatia Chagásica/imunologia , Integrinas/análise , Interferon gama/farmacologia , Selectina L/análise , Antígeno-1 Associado à Função Linfocitária/análise , Receptores de Antígenos de Linfócitos T alfa-beta/análise , Receptores de Retorno de Linfócitos/análise , Animais , Moléculas de Adesão Celular/biossíntese , Cardiomiopatia Chagásica/parasitologia , Cardiomiopatia Chagásica/patologia , Quimiocinas/biossíntese , Citocinas/biossíntese , Feminino , Imunofenotipagem , Integrina alfa4beta1 , Camundongos , Camundongos Endogâmicos C3H , Miocárdio/patologia , Parasitemia/mortalidadeRESUMO
Infection with Trypanosoma cruzi causes a strong inflammatory reaction at the inoculation site and, later, in the myocardium. The present study investigates the role of cytokines as modulators of T. cruzi-induced chemokine expression in vivo and in vitro. In macrophage cultures, although the stimulation with interferon (IFN)-gamma increases the expression of IP-10, it blocks KC expression. Tumor necrosis factor (TNF)-alpha, on the other hand, potentiates KC, IP-10, macrophage inflammatory protein-1alpha, and JE/monocyte chemotatic protein-1 expression. Interleukin-10 and transforming growth factor-beta inhibited almost all chemokines tested. The role of IFN-gamma and TNF-alpha in chemokine modulation during infection was investigated in T. cruzi-infected IFN-gamma-deficient (GKO) or TNF-R1/p55-deficient (p55-/-) mice. The expression of chemokines detected in the inoculation site correlated with the infiltrating cell type observed. Although GKO mice had a delayed and intense neutrophilic infiltrate correlating with the expression of KC and macrophage inflammatory protein-2, none of the above was observed in p55-/- mice. The detection of infiltrating T cells, Mig, and IP-10 in the myocardium was observed in wild-type and p55-/-, but not in GKO mice. Together, these results suggest that the regulatory roles of IFN-gamma and TNF-alpha on chemokine expression may play a crucial role in the modulation of the inflammatory response during T. cruzi infection and mediate resistance to infection.
Assuntos
Doença de Chagas/metabolismo , Quimiocinas/biossíntese , Interferon gama/deficiência , Peritonite/parasitologia , Receptores do Fator de Necrose Tumoral/deficiência , Animais , Antígenos CD , Movimento Celular , Doença de Chagas/patologia , Quimiocina CXCL10 , Quimiocina CXCL9 , Quimiocinas/genética , Quimiocinas CXC/metabolismo , Feminino , Imunofenotipagem , Interferon gama/fisiologia , Interleucina-10/fisiologia , Linfócitos/fisiologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Miocárdio/metabolismo , RNA Mensageiro/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral , Fator de Crescimento Transformador beta/fisiologia , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
We investigated the kinetics of parasite replication, leukocyte migration, and cytokine/chemokine mRNA expression in the heart tissue from animals infected with the Colombiana strain of Trypanosoma cruzi. Cardiac tissue parasitism was noticeable at 15 days, peaked around 30 days and was dramatically reduced at 120 days postinfection (p.i.). Kinetic studies showed that the inflammatory infiltrate was dominated by the presence of alphabetaT CD3(+ )CD4(+ )CD8(-), alphabetaT CD3(+ )CD4(-)CD8(+ )lymphocytes and macrophages. The mRNA expression of the monokines IL-1beta and IL-12(p40) was elevated at 15 days p.i. and controlled at later time points. In contrast, TNF-alpha mRNA was expressed throughout the infection. Interestingly, we found that at 15 and 30 days p.i. cytokine expression was dominated by the presence of IFN-gamma mRNA, whereas at 60 days or later time points the balance of type 1 and type 2 cytokines was switched in favor of IL-4 and IL-10 mRNAs. The chemokine mRNAs encoding JE, MIP-1alpha, MIP-1beta, KC, and MIP-2 were all mainly expressed at 15 and/or 30 days p.i. and diminished thereafter. In contrast, the expression of RANTES, MIG and IP-10 mRNAs was augmented at 15 days p.i. and persisted at high levels up to 120 days p.i. Taken together, our results indicate that regulation of IFN-gamma and chemokine expression, associated with decreased tissue parasitism, may be largely responsible for the control of inflammation and immunopathology observed in the cardiac tissue of animals infected with T. cruzi.