RESUMO
Trypanosome infections are marked by severe pathological features, including anemia, splenomegaly, and suppression of T-cell proliferation. We have used lymphotoxin-alpha-deficient (LT-alpha(-/-)) mice, as well as LT-alpha-tumor necrosis factor-double-deficient (LT-alpha(-/-) TNF(-/-)) mice, to analyze the contributions of these related cytokines in both induction of trypanosomosis-associated immunopathology and infection control. Moreover, as the cytokine-deficient mice used have no detectable lymph nodes and lack germinal-center formation upon immune stimulation, we have analyzed the functional importance of both the lymph nodes and spleen during experimental Trypanosoma brucei infections. First, we show that the absence of LT-alpha does not significantly alter early trypanosomosis development or pathology but does result in better control of late-stage parasitemia levels and slightly prolonged survival. This increased survival of infected LT-alpha(-/-) mice coincides with the appearance of increased chronic-stage anti-trypanosome immunoglobulin M (IgM)-IgG2a serum titers that are generated in the absence of functional peripheral lymphoid tissue and do not require germinal-center formation. Second, we show that splenectomized mice control their parasitemia to the same extent as fully immune-competent littermates. Finally, using LT-alpha(-/-) TNF(-/-) double-deficient mice, we show that in these mice T. brucei infections are very well controlled during the chronic infection stage and that infection-induced pathology is minimized. Together, these findings indicate that while increased IgM-IgG2a anti-trypanosome antibody titers (generated in the absence of LT-alpha, peripheral lymph nodes, and germinal-center formation) coincide with improved parasitemia control, it is TNF that has a major impact on trypanosomosis-associated immunopathology.
Assuntos
Linfotoxina-alfa/biossíntese , Trypanosoma brucei brucei/imunologia , Tripanossomíase Africana/imunologia , Animais , Anticorpos Antiprotozoários/sangue , Feminino , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Interferon gama/sangue , Linfonodos/patologia , Linfotoxina-alfa/genética , Camundongos , Camundongos Mutantes , Parasitemia , Baço/imunologia , Esplenectomia , Tripanossomíase Africana/mortalidade , Tripanossomíase Africana/patologia , Fator de Necrose Tumoral alfa/análise , Fator de Necrose Tumoral alfa/genéticaRESUMO
Experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein peptide 35-55 (MOG) leads to a chronic form of disease characterized by demyelination, inflammation and gliosis in the central nervous system (CNS). Recently IL-6 and LT alpha were found to be required for induction of the disease. The main features associated with EAE resistance of IL-6(-/-) and LT alpha(-/-) mice were reduced T cell proliferation and endothelial activation. As shown here treatment of MOG-immunized IL-6(-/-) mice with staphylococcal enterotoxin B (SEB)reversed their resistance to MOG-induced EAE. SEB failed to restore susceptibility to EAE in LT alpha(-/-) mice. The effect of SEB to induce EAE in IL-6(-/-) mice depends on TNF receptor type 1 (TNFR1) signaling because IL-6/TNF/LT alpha(-/-) and IL-6/TNFR1(-/-) are refractory to SEB. TNFR1 is involved in SEB induced trafficking of T cells into the CNS as evidenced by the failure to up-regulate VCAM-1 on CNS endothelium and lack of accumulation of V beta 8(+) T cells in the CNS of IL-6/TNFR1(-/-) mice upon immunization with MOG and treatment with SEB. The course of SEB triggered EAE in MOG immunized IL-6(-/-) mice was characterized by reduced severity and duration of clinical manifestations, which were associated with a significant drop of CNS infiltrating neutrophils and MIP-2 expression after peak disease. Taken collectively the effect of SEB to overcome EAE resistance points to a transient IL-6 independent but TNFR1 dependent proinflamatory pathway in EAE pathogenesis and suggests a crucial function for IL-6 in disease perpetuation.
Assuntos
Antígenos CD/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Interleucina-6/deficiência , Glicoproteína Associada a Mielina/imunologia , Receptores do Fator de Necrose Tumoral/metabolismo , Superantígenos/imunologia , Animais , Apresentação de Antígeno/imunologia , Apoptose , Divisão Celular , Sistema Nervoso Central/imunologia , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/patologia , Citocinas/biossíntese , Citocinas/genética , Encefalomielite Autoimune Experimental/induzido quimicamente , Enterotoxinas/imunologia , Inflamação/imunologia , Inflamação/patologia , Interleucina-6/genética , Interleucina-6/imunologia , Ativação Linfocitária , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos , Camundongos Knockout , Proteínas da Mielina , Glicoproteína Mielina-Oligodendrócito , Neutrófilos/imunologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
Signaling via TNF receptor type 1 (TNFR1) was shown to be crucial in host defense against the intracellular pathogens L. monocytogenes, M. tuberculosis and M. bovis. To investigate the function of TNF and LTalpha in host defense against M. bovis, mice double deficient for TNF and LTalpha (TNF / LTalpha (- / -)), TNF / LTalpha (- / -) mice complemented with a murine LTalpha transgene (TNF(- / -)) and LTalpha (- / -) mice were infected with BCG and the ensuing pathology was investigated. Control mice showed a normal host defense with early clearance of bacteria. The granulomatous reaction in the liver was accompanied by recruitment of activated macrophages characterized by their acid phosphatase positivity and differentiation into epithelioid cells as well as a coordinated expression of proinflammatory transcripts. In contrast, TNF / LTalpha (- / -) mice showed no comparable recruitment of activated macrophages in the liver. Furthermore, these mice showed extensive necrotic pulmonary lesions with massive growth of acid fast bacilli. Reintroduction of LTalpha as a transgene into TNF / LTalpha (- / -) mice prolonged survival but did not restore resistance to BCG. This, at least partially protective role of LTalpha was further supported by data demonstrating that LTalpha -deficient mice as well were susceptible to BCG infection. In contrast to the deleterious effect of TNF / LTalpha deficiency in BCG infection, BCG-infected TNF / LTalpha (- / -) mice were tolerant to LPS-induced shock. These results demonstrate that TNF as well as LTalpha are involved in murine host defense against BCG and that absence of TNF / LTalpha protects BCG-infected mice from LPS mediated shock.
Assuntos
Linfotoxina-alfa/imunologia , Mycobacterium bovis/imunologia , Fator de Necrose Tumoral alfa/imunologia , Animais , Células Cultivadas , Quimiocinas/genética , Citocinas/genética , Expressão Gênica , Granuloma/imunologia , Imunocompetência/imunologia , Linfotoxina-alfa/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Monócitos/imunologia , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase Tipo II , RNA Mensageiro , Baço/citologia , Baço/imunologia , Tuberculose/imunologia , Fator de Necrose Tumoral alfa/genéticaRESUMO
The expression of the chemokines macrophage inflammatory protein (MIP)-2 and MIP-1alpha and of their receptors CXCR2 and CCR5 was assessed in wild type (WT) and TNF/lymphotoxin-alpha knockout (TNF/LT-alpha-/-) mice subjected to closed head injury (CHI). At 4 h after trauma intracerebral MIP-2 and MIP-1alpha levels were increased in both groups with MIP-2 concentrations being significantly higher in WT than in TNF/LT-alpha-/- animals (p < 0.05). Thereafter, MIP-2 production declined rapidly, whereas MIP-1alpha remained elevated for 7 days. Expression of CXCR2 was confined to astrocytes and increased dramatically within 24 h in both mouse types. Contrarily, CCR5 expression remained constitutively low and was mainly localized to microglia. These results show that after CHI, chemokines and their receptors are regulated differentially and with independent kinetics.
Assuntos
Córtex Cerebral/metabolismo , Quimiocinas/metabolismo , Encefalite/metabolismo , Traumatismos Cranianos Fechados/metabolismo , Receptores de Quimiocinas/metabolismo , Animais , Astrócitos/metabolismo , Córtex Cerebral/fisiopatologia , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocina CXCL2 , Encefalite/fisiopatologia , Regulação da Expressão Gênica/fisiologia , Traumatismos Cranianos Fechados/fisiopatologia , Linfotoxina-alfa/genética , Linfotoxina-alfa/metabolismo , Proteínas Inflamatórias de Macrófagos/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/metabolismo , Monocinas/metabolismo , Receptores CCR5/metabolismo , Receptores de Interleucina-8B/metabolismo , Fator de Necrose Tumoral alfa/deficiência , Fator de Necrose Tumoral alfa/genética , Regulação para Cima/genéticaRESUMO
Systemic hyporesponsiveness occurs following oral administration of antigen (oral tolerance) and involves the uptake and processing of antigen by the gut-associated lymphoid tissue (GALT), which includes Peyer's patches (PP) lamina propria lymphocytes and mesenteric lymph nodes (MLN). Animals with targeted mutations of genes in the tumor necrosis factor (TNF) family have differential defects in the development of peripheral lymphoid organs including PP and MLN, and provide a unique opportunity to investigate the role of GALT structures in the induction of oral tolerance. Oral tolerance could not be induced in TNF/lymphotoxin (LT) alpha-/- mice, which are devoid of both PP and MLN, although these animals could be tolerized by intraperitoneal administration of antigen, demonstrating the requirement for GALT for oral tolerance induction. LTbeta-/- mice and LTalpha/LTbeta+/- animals do not have PP but could be orally tolerized, as measured by IFN-gamma production and delayed-type hypersensitivity responses by administration of both low or high doses of ovalbumin. To further investigate the requirement for PP, we tested the progeny of LTbeta-receptor-IgG-fusion-protein (LTbetaRigG)-treated mice, which do not form PP but have an otherwise intact immune system. Although these animals had decreased fecal IgA production, they could be orally tolerized. Our results demonstrate that PP are not an absolute requirement for the induction of either high- or low-dose oral tolerance, although oral tolerance could not be induced in animals devoid of both PP and MLN.
Assuntos
Tolerância Imunológica/imunologia , Imunidade nas Mucosas/imunologia , Nódulos Linfáticos Agregados/anormalidades , Nódulos Linfáticos Agregados/imunologia , Administração Oral , Animais , Anticorpos/imunologia , Anticorpos/farmacologia , Relação Dose-Resposta Imunológica , Ensaio de Imunoadsorção Enzimática , Fezes/química , Deleção de Genes , Hipersensibilidade Tardia/imunologia , Tolerância Imunológica/efeitos dos fármacos , Imunidade nas Mucosas/efeitos dos fármacos , Imunoglobulina A/imunologia , Interferon gama/biossíntese , Interferon gama/metabolismo , Interleucina-2/metabolismo , Linfonodos/anormalidades , Linfonodos/efeitos dos fármacos , Linfonodos/imunologia , Linfotoxina-alfa/antagonistas & inibidores , Linfotoxina-alfa/genética , Linfotoxina-alfa/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina/administração & dosagem , Ovalbumina/imunologia , Nódulos Linfáticos Agregados/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
In this review we summarize the essential findings about the function of tumour necrosis factor (TNF) and its cognate receptors TNFR1 and TNFR2, and lymphotoxin alpha (LT-alpha) ligands in immune-mediated CNS inflammation and demyelination. The advent of homologous recombination technology in rodents provides a new method which has been used during the last 5 years and has led to insights into the pathophysiology of experimental autoimmune encephalomyelitis (EAE) in an unprecedented way. Studies with knockout mice in which genes of the TNF ligand/receptor superfamily are not expressed and studies with transgenic mice overexpressing TNF and TNFR reveal the critical role of the TNFR1 signalling pathway in the control of CNS demyelination and inflammation. These studies provide novel findings and at the same time shed light on the complex pathophysiology of EAE. Together, these findings may contribute to better understanding of EAE and open new avenues in experimental therapies for multiple sclerosis.
Assuntos
Antígenos CD/imunologia , Doenças Desmielinizantes/fisiopatologia , Encefalomielite Autoimune Experimental/fisiopatologia , Receptores do Fator de Necrose Tumoral/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Animais , Doenças Desmielinizantes/imunologia , Encefalomielite Autoimune Experimental/imunologia , Esclerose Múltipla/imunologia , Esclerose Múltipla/fisiopatologia , Receptores Tipo I de Fatores de Necrose TumoralRESUMO
Cytokines are important mediators of intracranial inflammation following traumatic brain injury (TBI). In the present study, the neurological impairment and mortality, blood-brain barrier (BBB) function, intracranial polymorphonuclear leukocyte (PMN) accumulation, and posttraumatic neuronal cell death were monitored in mice lacking the genes for tumor necrosis factor (TNF)/lymphotoxin-alpha (LT-alpha) (TNF/LT-alpha-/-) and interleukin-6 (IL-6) and in wild-type (WT) littermates subjected to experimental closed head injury (total n = 107). The posttraumatic mortality was significantly increased in TNF/LT-alpha-/- mice (40%; P < 0.02) compared with WT animals (10%). The IL-6-/- mice also showed a higher mortality (17%) than their WT littermates (5.6%), but the difference was not statistically significant (P > 0.05). The neurological severity score was similar among all groups from 1 to 72 hours after trauma, whereas at 7 days, the TNF/LT-alpha-/- mice showed a tendency toward better neurological recovery than their WT littermates. Interestingly, neither the degree of BBB dysfunction nor the number of infiltrating PMNs in the injured hemisphere was different between WT and cytokine-deficient mice. Furthermore, the analysis of brain sections by in situ DNA nick end labeling (TUNEL histochemistry) at 24 hours and 7 days after head injury revealed a similar extent of posttraumatic intracranial cell death in all animals. These results show that the pathophysiological sequelae of TBI are not significantly altered in mice lacking the genes for the proinflammatory cytokines TNF, LT-alpha, and IL-6. Nevertheless, the increased posttraumatic mortality in TNF/LT-alpha-deficient mice suggests a protective effect of these cytokines by mechanisms that have not been elucidated yet.
Assuntos
Barreira Hematoencefálica/fisiologia , Citocinas/genética , Traumatismos Cranianos Fechados/imunologia , Traumatismos Cranianos Fechados/fisiopatologia , Neutrófilos/imunologia , Animais , Morte Celular , Modelos Animais de Doenças , Traumatismos Cranianos Fechados/mortalidade , Marcação In Situ das Extremidades Cortadas , Interleucina-6/genética , Linfotoxina-alfa/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Exame Neurológico , Neurônios/citologia , Fator de Necrose Tumoral alfa/genéticaRESUMO
Tumor necrosis factor-alpha (TNFalpha) exists in two biologically active forms, a 26-kDa transmembrane form and a proteolytically cleaved and secreted form. We sequentially inactivated all three known cleavage sites of mouse TNFalpha by mutating the corresponding DNA sequences. A murine T cell hybridoma transfected with the nonsecretable mutant TNFalpha efficiently lysed L929 target cells in a cell contact-dependent manner and induced expression of vascular cell adhesion molecule-1 on mouse endothelioma cells. A genomic mouse TNFalpha clone encoding this mutant was subsequently introduced as a transgene into TNFalpha(-/-) lymphotoxin-alpha(-/-) mice. The 3' AU-rich regulatory elements of the TNF locus were maintained in the transgene to assure adequate gene regulation. Transmembrane TNFalpha transgenic mice were fully protected from endotoxic shock, and no TNFalpha bioactivity was detectable in the serum after stimulation with lipopolysaccharide. Activated CD4 T cells from these animals, however, lysed L929 cells in a cell contact-dependent way. After administration of lipopolysaccharide, transmembrane TNFalpha transgenic mice produced significantly higher levels of interleukin-12 than wild-type mice or TNF-deficient mice. This indicates that transmembrane TNFalpha may greatly affect the course of a cellular immune responses in vivo and exerts quantitatively and qualitatively distinct functions from secreted TNFalpha in vitro and in vivo.
Assuntos
Proteínas de Membrana/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Células 3T3 , Sequência de Aminoácidos , Animais , Técnicas de Cocultura , DNA Complementar , Interleucina-12/sangue , Lipopolissacarídeos/farmacologia , Proteínas de Membrana/genética , Camundongos , Dados de Sequência Molecular , Choque Séptico/genética , Choque Séptico/prevenção & controle , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/genética , Molécula 1 de Adesão de Célula Vascular/biossínteseRESUMO
In this study, we addressed the role of tumor necrosis factor (TNF)-alpha and lymphotoxin (LT)-alpha in the development of colitis and defined the cellular sources (T cells versus non-T cells) of TNF (TNF-alpha and LT-alpha) relevant to disease development. After adoptive transfer of TNF(+/+) CD4(+)CD45RB(hi) splenocytes into TNF(+/+) recombination activating gene (RAG)2(-/-) mice, the recipients develop massive inflammation of the large intestinal mucosa concurrent with massive weight loss. In contrast, clinical signs of disease are completely absent in TNF(-/-)RAG2(-/-) recipients of TNF(-/-) CD4(+)CD45RB(hi) T cells, although elevated numbers of interferon-gamma-producing cells are present in the colonic mucosa. Surprisingly, upon transfer of TNF(-/-)CD4(+)CD45RB(hi) T cells into TNF(+/+)RAG2(-/-) recipients, colitis develops with kinetics similar to those upon transfer of TNF(+/+)CD4(+)CD45RB(hi) donor cells. In contrast, no clinical signs of colitis are observed in TNF(-/-)RAG2(-/-) recipients of TNF(+/+)CD4(+)CD45RB(hi) T cells. This protection from colitis is not a consequence of the absence of LT-alpha, as TNF-alpha(-/-)RAG2(-/-) recipients of TNF-alpha(-/-) CD4(+)CD45RB(hi) T cells are also protected from colitis induction. These results demonstrate the importance of TNF production by non-T cells of the colonic mucosa in the pathogenesis of colitis and provide direct evidence for a nonredundant role of TNF-alpha in this mouse model of colitis.
Assuntos
Transferência Adotiva , Linfócitos T CD4-Positivos/fisiologia , Proteínas de Ligação a DNA/fisiologia , Doenças Inflamatórias Intestinais/etiologia , Antígenos Comuns de Leucócito/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Animais , Proteínas de Ligação a DNA/genética , Interferon gama/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/análise , Fator de Necrose Tumoral alfa/genéticaRESUMO
Mice deficient for TNF ligand and receptor type 1 have demonstrated the importance of TNF in the host defense against Listeria monocytogenes. To investigate the particular deficiency of macrophages derived from TNF/lymphotoxin (LT)-alpha(-/-) mice in antilisterial growth control, bone marrow-derived macrophages (BMDM) were used for in vitro infection experiments. After the combined treatment with IFN-gamma and lipopolysaccharide (LPS), production of NO by wild-type (wt) and TNF/LT-alpha(-/-) BMDM was induced to comparable levels, but only wt BMDM controlled L. monocytogenes growth efficiently. Nevertheless, inhibition of NO production led to a remarkable loss of antilisterial activity. This suggests that presence of NO is necessary but not sufficient for L. monocytogenes killing and that elimination of L. monocytogenes requires additional effector molecules. The LPS-inducible superoxide production of TNF/LT-alpha(-/-) BMDM was impaired. Accordingly both scavenging of superoxide and peroxynitrite led to reduced L. monocytogenes killing by wt BMDM. In addition, peroxynitrite was able to kill L. monocytogenes in vitro. Together these findings suggest that the defective host defense of TNF/LT-alpha-deficient mice against L. monocytogenes partially stems from reduced superoxide production of macrophages due to the absence of TNF and imply a function for peroxynitrite, the reaction product of NO and superoxide, in the intracellular killing of L. monocytogenes.
Assuntos
Listeria monocytogenes/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Superóxidos/metabolismo , Fator de Necrose Tumoral alfa/deficiência , Animais , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Células da Medula Óssea/microbiologia , Sequestradores de Radicais Livres/imunologia , Sequestradores de Radicais Livres/metabolismo , Síndromes de Imunodeficiência/metabolismo , Síndromes de Imunodeficiência/microbiologia , Listeria monocytogenes/crescimento & desenvolvimento , Macrófagos/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Nitratos/metabolismo , Óxido Nítrico/biossíntese , Óxido Nítrico/imunologia , Oxidantes/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/imunologiaRESUMO
T-cell apoptosis in inflammatory demyelinating lesions of chronic myelin oligodendrocyte glycoprotein peptide35-55 induced autoimmune encephalomyelitis was studied in several different gene knockout mice as well as their wild-type counterparts. The gene deletions included tumor necrosis factor (TNF) alpha, lymphotoxin, TNF receptor 1 or 2, Fas-L, inducible nitric oxide synthase, perforin, and interleukin1beta-converting enzyme. Impairment of the TNF receptor 1 pathway led to a 50% reduction of T-cell apoptosis in the central nervous system lesions, whereas the other genetic deletions showed no significant effect. Our study thus identified the TNF receptor 1 signaling pathway as one mechanism responsible for the removal of T lymphocytes from inflammatory demyelinating lesions of the central nervous system.
Assuntos
Apoptose/fisiologia , Encefalomielite Autoimune Experimental/fisiopatologia , Glicoproteína Associada a Mielina/toxicidade , Fragmentos de Peptídeos/toxicidade , Receptores do Fator de Necrose Tumoral/fisiologia , Transdução de Sinais/fisiologia , Linfócitos T/patologia , Animais , Caspase 10 , Caspases/metabolismo , Encefalomielite Autoimune Experimental/induzido quimicamente , Proteína Ligante Fas , Glicoproteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Proteínas da Mielina , Glicoproteína Associada a Mielina/química , Glicoproteína Mielina-Oligodendrócito , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II , Perforina , Proteínas Citotóxicas Formadoras de Poros , Receptor fasRESUMO
The individual role of tumor necrosis factor receptor 1 (TNFR1) and TNFR2 signaling in experimental autoimmune encephalomeylitis (EAE) was investigated using mice lacking TNFR1 (TNFR1-/-), TNFR2 (TNFR2-/-) as well as double receptor (TNFR1/2-/-) and double ligand (TNF/LT alpha-/-) knockout mice. In wild-type (wt) mice immunized with myelin oligodendrocyte glycoprotein (MOG) peptide 35-55 the clinical course is characterized by an acute disease onset with peak disease scores and a consecutive chronic phase lasting up to 60 days. Compared to control mice, TNF/LT alpha-deficient mice showed a significant delay in disease onset and a remarkable reduction in demyelination which was, however, associated with increased inflammation. In TNFR1-/- and TNFR1/2-/- mice, the disease course was comparable to TNF/LT alpha-deficient mice but rather monophasic and less severe at late time points. Likewise only minimal spinal cord demyelination became apparent. In contrast, the course of EAE in TNFR2-/- mice was severe and associated with remarkable demyelination. Taken together these findings define TNFR1 as crucial mediator in MOG-induced EAE and suggest a protective role for TNFR2 signaling in the clinical course of EAE.
Assuntos
Encefalomielite Autoimune Experimental/imunologia , Receptores do Fator de Necrose Tumoral/imunologia , Animais , Encefalomielite Autoimune Experimental/fisiopatologia , Camundongos , Camundongos Knockout , Proteínas da Mielina , Bainha de Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Receptores do Fator de Necrose Tumoral/genéticaRESUMO
The constellation of changes known as the acute phase response (APR) is a cytokine-driven process initiated by tissue inflammation. The proinflammatory cytokines, TNF, IL-1 and IL-6, are considered to be the primary mediators of the APR. IL-6 and IL-1beta gene-deleted mice (Fattori et al., J. Exp. Med. 1994. 180: 1243-1250; Kopf et al., Nature 1994. 368: 339-342; Fantuzzi et al., J. Immunol. 1996. 157: 291-296, respectively), exhibit impaired APR to turpentine injection but only a slight reduction in plasma acute phase protein levels in response to lipopolysaccharide (LPS). This infers an important role for TNF in the LPS-induced APR, however, in the present study, normal APR to both turpentine and LPS were observed in TNF/LTalpha-deficient mice. A striking absence of elevated major acute phase proteins, SAP and SAA, was observed in mice deficient in TNF/LTalpha and IL-6, suggesting that TNF-alpha or LTalpha do indeed exert important nonredundant synergism in the IL-1/IL-6 primary response. The regulation of other parameters typically altered in an APR, body weight, blood glucose and haptoglobin, was normal in LPS-dosed TNF/LTalpha-deficient and wild-type mice. The observed transcriptional response for SAA and SAP in these TNF/LTalpha/IL-6-deficient mice, in lieu of elevated plasma levels, suggests that SAA and SAP expression is possibly posttranscriptionally regulated.
Assuntos
Reação de Fase Aguda/imunologia , Interleucina-6/imunologia , Lipopolissacarídeos/toxicidade , Fator de Necrose Tumoral alfa/imunologia , Reação de Fase Aguda/induzido quimicamente , Animais , Interleucina-1/deficiência , Interleucina-1/genética , Interleucina-1/imunologia , Interleucina-6/deficiência , Interleucina-6/genética , Linfotoxina-alfa/genética , Linfotoxina-alfa/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Fator de Necrose Tumoral alfa/deficiência , Fator de Necrose Tumoral alfa/genéticaRESUMO
To evaluate the importance of the proinflammatory cytokines TNF and lymphotoxin-alpha (LT alpha) in an experimental model of Staphylococcus aureus sepsis and arthritis, we used TNF/LT alpha-double-deficient mice raised on the C57BL/6 background. Mice were i.v. inoculated with a toxic shock syndrome toxin-1 (TSST-1)-producing S. aureus strain, LS-1. Intravenous inoculation of a high dose of bacteria (1 x 10(7)/mouse) resulted in 67% mortality in TNF/LT alpha-deficient mice, whereas none of the controls died (p = 0.009). Those results correlated to a significantly decreased phagocytosis in vitro and inefficient bacterial clearance in vivo in mice lacking capacity to produce TNF/LT alpha. Thus, at day 6 after inoculation, S. aureus could not be found in the bloodstream of controls, but bacteremia developed in all TNF/LT alpha-deficient mice examined (p = 0.02). Interestingly, upon infection with a lower dose of staphylococci (3 x 10(6)/mouse) the mortality was overall low, but the frequency of arthritis was clearly higher in the wild-type group as compared with the TNF/LT alpha-deficient mice (40% vs 13%). Histopathologic examination revealed a lower frequency of synovitis (38% vs 90%, p < 0.05) and erosivity (25% vs 60%, NS) in TNF/LT alpha-deficient mice as compared with wild-type counterparts. Our results show the importance of TNF/LT alpha in defense against systemic S. aureus infections and point out the detrimental role of these cytokines as mediators of inflammatory response in S. aureus arthritis.
Assuntos
Artrite Infecciosa/imunologia , Linfotoxina-alfa/genética , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/imunologia , Fator de Necrose Tumoral alfa/genética , Animais , Especificidade de Anticorpos , Artrite Infecciosa/genética , Artrite Infecciosa/microbiologia , Artrite Infecciosa/mortalidade , Linfócitos B/imunologia , Linfócitos B/metabolismo , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Epitopos de Linfócito B/imunologia , Imunoglobulina G/biossíntese , Interferon gama/biossíntese , Ativação Linfocitária/genética , Linfotoxina-alfa/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NZB , Camundongos Knockout , Fagocitose , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/mortalidade , Staphylococcus aureus/crescimento & desenvolvimento , Staphylococcus aureus/isolamento & purificação , Análise de Sobrevida , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Mice with inactivated TNF-LT alpha genes have profound abnormalities of the immune system with hypoimmunoglobulinaemia, lack of lymph nodes, undifferentiated spleen, and defective Ig class switch. Transplantation of bone marrow cells from wild-type mice restored the synthesis of TNF, corrected the splenic microarchitecture, repopulated the lamina propria with IgA-producing plasma cells, and normalized the serum immunoglobulin levels of TNF-LT alpha deficient mice. Furthermore, the formation of germinal centers in the spleen and the defective Ig class switch in response to a T-cell-dependent antigen is corrected. These data demonstrate that most TNF-producing cells are bone-marrow-derived, and that the immunodeficiency due to TNF-LT alpha deletion can be corrected to a large extent by normal bone marrow, cell transplantation.
Assuntos
Células da Medula Óssea/imunologia , Transplante de Medula Óssea , Linfotoxina-alfa/biossíntese , Fator de Necrose Tumoral alfa/biossíntese , Animais , Imunofluorescência , Centro Germinativo/imunologia , Imunoglobulina A/biossíntese , Imunoglobulina A/sangue , Switching de Imunoglobulina , Imunoglobulina G/sangue , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/imunologia , Linfotoxina-alfa/genética , Camundongos , Camundongos Knockout , Plasmócitos/imunologia , Baço/imunologia , Fator de Necrose Tumoral alfa/deficiência , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Experimental autoimmune encephalomyelitis (EAE) is induced by immunization with myelin components including myelin oligodendrocyte glycoprotein (MOG). Myelin-specific Th1 cells enter the central nervous system (CNS) via binding of very late antigen 4 (VLA-4) to the endothelial vascular cell adhesion molecule 1 (VCAM-1). In the present study, mice with a homologous disruption of the gene encoding IL-6 are found to be resistant to MOG-induced EAE as evidenced by absence of clinical symptoms, minimal infiltration of CD3+ T cells and monocytes into the CNS and lack of demyelination. The failure to induce EAE in IL-6-/- mice is not due to the absence of priming, since lymphocytes of immunized IL-6-/- mice proliferate in response to MOG and produce pro-inflammatory cytokines including IL-2 and IFN-gamma. However, in MOG-immunized IL-6-/- mice, serum anti-MOG antibody titers were found to be drastically reduced. This observation is unlikely to be responsible for resistance to EAE, because B cell-deficient (microMT) mice proved to be fully susceptible to the disease. A striking difference between MOG-immunized wild-type (wt) and IL-6-/- mice was the expression of endothelial VCAM-1 and ICAM-1, which were dramatically up-regulated in the CNS in wt but not in IL-6-/- mice. Taking into account recent studies on the role of VCAM-1 in the entry of Th1 cells into the CNS, the absence of VCAM-1 on endothelial cells in IL-6-/- mice may explain their resistance to EAE.
Assuntos
Encefalomielite Autoimune Experimental/etiologia , Interleucina-6/fisiologia , Glicoproteína Associada a Mielina/imunologia , Animais , Suscetibilidade a Doenças , Encefalomielite Autoimune Experimental/imunologia , Endotélio Vascular/metabolismo , Feminino , Molécula 1 de Adesão Intercelular/biossíntese , Interleucina-6/deficiência , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas da Mielina , Glicoproteína Mielina-Oligodendrócito , Linfócitos T/imunologia , Regulação para Cima , Molécula 1 de Adesão de Célula Vascular/biossínteseRESUMO
Leukocyte extravasation is governed by the endothelium, expressing a defined pattern of adhesion molecules in response to inflammatory stimuli. Among them, E-selectin, which is expressed in response to interleukin 1 (IL-1) or tumour necrosis factor alpha (TNF-alpha), provides rolling adhesion of the circulating leukocytes, a transient and reversible interaction that initiates leukocyte extravasation. In our experiments, E-selectin expression culminated after 4 to 6 h and declined thereafter. After 24 h a considerable amount of E-selectin was presented, reflecting its continuous expression in different inflammatory skin disorders. After preincubation of the endothelial cells with TNF-alpha together with IL-4 or IL-13, E-selectin mRNA transcription and protein expression were markedly reduced at 8 h and almost abolished at 20 h. In contrast, early E-selectin expression between 2 to 6 h was not significantly impaired. In a rotating adherence assay that mimics physiological shear forces in circulation, preincubation of the endothelial cells with TNF-alpha for 4 and 20 h induced similar adherence of neutrophils, which was largely E-selectin dependent. According to the modified expression kinetics of E-selectin in the presence of IL-4 of IL-13 rolling adhesion was unimpaired at 4 h but significantly diminished after 20 h of preincubation. Similar results were obtained with clones of the cutaneous T cell lymphoma cell line HUT78 highly expressing the E-selectin ligand cutaneous lymphocyte-associated Ag. Together, these data suggest that IL-4 and IL-13 control the rolling adhesion by limiting the period of the induced E-selectin expression and may thereby confine the acute phase of leukocyte emigration.
Assuntos
Selectina E/metabolismo , Endotélio Vascular/metabolismo , Interleucina-13/farmacologia , Interleucina-4/farmacologia , Neutrófilos/metabolismo , Anticorpos Bloqueadores/farmacologia , Anticorpos Monoclonais/farmacologia , Antígenos de Diferenciação de Linfócitos T , Antígenos de Neoplasias , Adesão Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo , Selectina E/imunologia , Endotélio Vascular/efeitos dos fármacos , Citometria de Fluxo , Humanos , Interleucina-1/farmacologia , Glicoproteínas de Membrana/metabolismo , Reação em Cadeia da Polimerase , Receptores de Retorno de Linfócitos/metabolismo , Fatores de Tempo , Transcrição Gênica/efeitos dos fármacos , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/farmacologia , Molécula 1 de Adesão de Célula Vascular/metabolismoRESUMO
Recent evidence suggests that macrophages and/or other nonparenchymal cells may release important mediators contributing to the hepatic necrosis induced by high doses of acetaminophen (APAP). The nature and causative role of these mediators has remained elusive, however. To investigate the role of the proinflammatory cytokine, tumor necrosis factor (TNF) in the initiation and early propagation of APAP-induced liver injury, we have used mice deficient in both TNF and the closely related lymphotoxin-alpha (LT-alpha). Male TNF/LT-alpha knockout mice and C57BL/6 wild-type mice were treated with a hepatotoxic dose of APAP (400 mg/kg, intraperitoneally), and the development of liver injury was monitored over 8 hours. Both genotypes exhibited similar basal activities of hepatic cytochrome P450 2E1 and 1A2. After APAP administration, both the rate of glutathione consumption and the extent of subsequent selective protein binding did not differ significantly in the knockout and wild-type mice. The TNF/LT-alpha-deficient mice developed severe centrilobular necrosis and exhibited highly increased levels of serum alanine aminotransferase and aspartate aminotransferase, the extent of which was not significantly different from that in wild-type mice. In C57BL/6 mice exposed to APAP, no increases in hepatic transcripts of TNF or LT-alpha were found by reverse transcription-polymerase chain reaction, nor was immunoreactive serum TNF detected by enzyme-linked immunosorbent assay over 8 hours posttreatment. These data indicate that, in the absence of the genes encoding for TNF and LT-alpha, APAP bioactivation was not altered and mice still developed severe hepatic necrosis. Thus, TNF is unlikely to be a key mediator in the early pathogenesis of APAP-induced hepatotoxicity.
Assuntos
Acetaminofen/toxicidade , Analgésicos não Narcóticos/toxicidade , Fígado/efeitos dos fármacos , Linfotoxina-alfa/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Acetaminofen/farmacocinética , Animais , Citocromo P-450 CYP1A2/metabolismo , Citocromo P-450 CYP2E1/metabolismo , Linfotoxina-alfa/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/análise , Coelhos , Fator de Necrose Tumoral alfa/genéticaRESUMO
Complement has been shown to contribute to intrathecal inflammation in bacterial meningitis. However, the cellular source of complement in the infected central nervous system has not been determined. In this study, we analyzed protein and mRNA expression of two alternative pathway complement activation proteins, C3 and factor B, in the brains of mice with Listeria monocytogenes meningitis. Complement protein levels were found elevated in the cerebrospinal fluid of infected mice, compared with mock-infected animals. In the course of the disease, enhanced C3 and factor B mRNA expression was detected on pyramidal neurons and Purkinje cells within 6 hours, peaking at 12 hours and then gradually decreasing by 72 hours after infection. In addition, leukocytes infiltrating the subarachnoid space, within 12 to 24 hours, expressed mRNA for C3 and factor B. The cellular infiltration increased dramatically up to 72 hours. Intraperitoneal injection of tumor necrosis factor (TNF)-alpha up-regulated C3 and factor B mRNA expression on neurons in normal mice, suggesting that TNF-alpha may represent one cytokine regulating complement expression in this model of bacterial meningitis. However, additional mediators may be involved in regulation of intrathecal complement expression, as infected mice deficient of TNF/lymphotoxin-alpha genes did not demonstrate attenuated complement expression in the brain.
Assuntos
Encéfalo/metabolismo , Complemento C3/biossíntese , Fator B do Complemento/biossíntese , Via Alternativa do Complemento , Meningites Bacterianas/metabolismo , Animais , Encéfalo/microbiologia , Encéfalo/patologia , Complemento C3/líquido cefalorraquidiano , Complemento C3/genética , Fator B do Complemento/líquido cefalorraquidiano , Fator B do Complemento/genética , Feminino , Regulação da Expressão Gênica , Leucócitos/metabolismo , Listeria monocytogenes , Listeriose , Camundongos , Neurônios/metabolismo , Neurônios/patologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
The anaphylatoxin C5a has been implicated in the pathogenesis of bacterial meningitis as a potent mediator of inflammation in the subarachnoid space. We investigated the expression of the receptor for C5a (C5aR) in brains of mice with experimental Listeria monocytogenes (LM) meningoencephalitis. In the course of the disease, infiltrating cells in the meninges and the ventricles were found to express C5aR mRNA and protein. In the brain parenchyma, very low constitutive C5aR expression was seen on pyramidal neurons and Purkinje cells. However, in LM-infected mice, a dramatic increase in C5aR expression occurred on neurons starting 6 h after infection and was maximal between 24 and 36 h. TNF-alpha was identified as an essential mediator of neuronal C5aR expression, since mice lacking the genes for TNF and lymphotoxin-alpha (TNF/lymphotoxin-alpha -/- mice) showed significantly attenuated C5aR expression after LM infection. Furthermore, i.p. injection of recombinant TNF-alpha induced enhanced C5aR expression in the brains of TNF/lymphotoxin-alpha -/- mice and in normal animals even in the absence of a bacterial infection. We also assessed the levels of anaphylatoxin C5a in the cerebrospinal fluid of patients with infectious meningitis. C5a was detected in all patients with bacterial meningitis (n = 9), in 6 of 18 patients with aseptic meningitis, and in 1 of 66 control patients. The finding of TNF-alpha-mediated C5aR expression on neurons in experimental Listeria meningitis and the detection of the ligand, C5a, in the cerebrospinal fluid of human patients with infectious meningitis present new directions in the investigation of the pathophysiologic sequelae leading to secondary brain damage.