RESUMO
Listeria monocytogenes (L.m) is efficiently controlled by several cells of the innate immunity, including the Mast Cell (MC). MC is activated by L.m inducing its degranulation, cytokine production and microbicidal mechanisms. TLR2 is required for the optimal control of L.m infection by different cells of the immune system. However, little is known about the MC receptors involved in recognizing this bacterium and whether these interactions mediate MC activation. In this study, we analyzed whether TLR2 is involved in mediating different MC activation responses during L.m infection. We found that despite MC were infected with L.m, they were able to clear the bacterial load. In addition, MC degranulated and produced ROS, TNF-α, IL-1ß, IL-6, IL-13 and MCP-1 in response to bacterial infection. Interestingly, L.m induced the activation of signaling proteins: ERK, p38 and NF-κB. When TLR2 was blocked, L.m endocytosis, bactericidal activity, ROS production and mast cell degranulation were not affected. Interestingly, only IL-6 and IL-13 production were affected when TLR2 was inhibited in response to L.m infection. Furthermore, p38 activation depended on TLR2, but not ERK or NF-κB activation. These results indicate that TLR2 mediates only some MC activation pathways during L.m infection, mainly those related to IL-6 and IL-13 production.
Assuntos
Interleucina-13/imunologia , Interleucina-6/imunologia , Listeria monocytogenes/imunologia , Mastócitos/imunologia , Receptor 2 Toll-Like/imunologia , Animais , Degranulação Celular/imunologia , Degranulação Celular/fisiologia , Células Cultivadas , Citocinas/imunologia , Citocinas/metabolismo , Ativação Enzimática/imunologia , Interações Hospedeiro-Patógeno/imunologia , Interleucina-13/metabolismo , Interleucina-6/metabolismo , Listeria monocytogenes/fisiologia , Mastócitos/microbiologia , Mastócitos/fisiologia , Camundongos Endogâmicos C57BL , NF-kappa B/imunologia , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/imunologia , Espécies Reativas de Oxigênio/metabolismo , Receptor 2 Toll-Like/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
The aim of this study was to analyze the 4-carvomenthenol (carvo) oral treatment on the experimental model of the combined allergic rhinitis and asthma syndrome (CARAS). BALB/c mice were OVA-sensitized on day zero and 7th (50 µg/mL OVA in 10 mg/mL Al (OH)3) and OVA-challenged (5 mg/mL, 20 µL/animal) for three weeks. In the last week, the animals were dally challenged with aerosol of OVA and the carvo treatment (12.5, 25 or 50 mg/kg) occurred one hour before each OVA-challenge. Data were analyzed and p < 0.05 was considered significant. Carvo (12.5-50 mg/kg) decreased significantly the eosinophil migration into the nasal (NALF) and bronchoalveolar (BALF) cavities as well as on the nasal and lung tissues of sick animals. The treatment also decreased mucus production on both tissue sections stained with PAS (periodic acid-Schiff satin). In addition, the histological analyzes demonstrated that sick mice presented hyperplasia and hypertrophy of the lung smooth muscle layer followed by increasing of extracellular matrix and carvo (50 mg/kg) inhibited these asthmatic parameters. We analyzed the allergic rhinitis signals as nasal frictions and sneezing and observed that carvo decreased these two signals as well as serum OVA-specific IgE titer, type 2 cytokine synthesis, mainly IL-13, with increasing of IL-10 production. Decreasing of IL-13 production corroborated with decreasing of mucus production and these effects were dependent on p38MAPK/NF-κB(p65) signaling pathway inhibition. Therefore, these data demonstrated that a monoterpene of essential oils presents anti-allergic property on an experimental model of CARAS suggesting a new drug prototype to treat this allergic syndrome.
Assuntos
Antialérgicos/uso terapêutico , Asma/tratamento farmacológico , Mentol/análogos & derivados , Rinite Alérgica/tratamento farmacológico , Alérgenos , Animais , Antialérgicos/farmacologia , Asma/sangue , Asma/imunologia , Asma/patologia , Líquido da Lavagem Broncoalveolar/imunologia , Citocinas/sangue , Citocinas/imunologia , Feminino , Interleucina-13/antagonistas & inibidores , Interleucina-13/imunologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Mentol/farmacologia , Mentol/uso terapêutico , Camundongos Endogâmicos BALB C , Muco/imunologia , NF-kappa B/imunologia , Ovalbumina , Rinite Alérgica/sangue , Rinite Alérgica/imunologia , Rinite Alérgica/patologia , Transdução de Sinais/efeitos dos fármacos , Síndrome , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
The type 1 TNF-α receptor (TNFR1) has a central role in initiating both pro-inflammatory and pro-apoptotic signaling cascades in neutrophils. Considering that TNFR1 signals Staphylococcus aureus protein A (SpA), the aim of this study was to explore the interaction of this bacterial surface protein with neutrophils and keratinocytes to underscore the signaling pathways that may determine the fate of these innate immune cells in the infected tissue during staphylococcal skin infections. Using human neutrophils cultured in vitro and isogenic staphylococcal strains expressing or not protein A, we demonstrated that SpA is a potent inducer of IL-8 in neutrophils and that the induction of this chemokine is dependent on the SpA-TNFR1 interaction and p38 activation. In addition to IL-8, protein A induced the expression of TNF-α and MIP-1α highlighting the importance of SpA in the amplification of the inflammatory response. Protein A contributed to reduce neutrophil mortality prolonging their lifespan upon the encounter with S. aureus. Signaling initiated by SpA modulated the type of neutrophil cell death in vitro and during skin and soft tissue infections (SSTI) in vivo triggering the apoptotic pathway instead of necrosis. Moreover, SpA induced pro-inflammatory cytokines in keratinocytes, modulating their survival in vitro and preventing the exacerbated necrosis and ulceration of the epithelium during SSTI in vivo. Taken together, these results highlight the importance of the inflammatory signaling induced by protein A in neutrophils and skin epithelial cells. The ability of protein A to modulate the neutrophil/epithelial cell death program in the skin is of clinical relevance considering that lysis of neutrophils and epithelial cells will promote an intense inflammatory response and contribute to tissue damage, a non-desirable feature of complicated SSTI.
Assuntos
Queratinócitos/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Neutrófilos/imunologia , Proteína Estafilocócica A/imunologia , Staphylococcus aureus/imunologia , Citocinas/imunologia , Humanos , Queratinócitos/microbiologia , Neutrófilos/microbiologia , Receptores Tipo I de Fatores de Necrose Tumoral/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
INTRODUCTION: This study investigated the mechanism of action of a synthetic tetrahydroisoquinoline alkaloid, MHTP, in an experimental model of acute lung injury (ALI) in two distinct moments: 72 h and 10 days. METHODOLOGY: To realize this study, 2.5 mg/kg of lipopolysaccharide (LPS) was intranasally administered in BALB/c mice, and nasal instillation of MHTP (1.25; 2.5; 5.0; 10 or 20 mg/kg) was administrated at 1, 24, and 48 h after LPS challenge. The data were statistically analyzed and p < 0.05 was considered statistically significant. RESULTS: MHTP treatment (2.5, 5.0, 10 or 20 mg/kg) significantly decreased neutrophil migration into the bronchoalveolar lavage fluid (BALF), tissue inflammatory cell infiltration, edema, and hemorrhage as well as collagen fiber deposition on the perialveolar regions at both moments. TNF-α and IL-6 levels were significantly diminished in the MHTP-treated animals at 72 h and maintained them, at a basal level, at 10-day observation. These effects of MHTP are due to downregulating p38MAPkinese/p65NFκB signaling pathway-TLR4 dependent. Also, the MHTP treatment promoted a survival rate at 100% and improved their body weights during the 10-day observation. Unlike, the LPS group (non-treated LPS challenged animals) presented less than 50% of surviving rate at 72 h and the animals that survived did not improve their physiological state at 10-day observation. CONCLUSIONS: These data showed for the first time the beneficial and effective activity of a nasal treatment with a synthetic tetrahydroisoquinoline alkaloid on an experimental model of ALI and pointed out the molecular mechanism related to it.
Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Tetra-Hidroisoquinolinas/uso terapêutico , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/patologia , Administração Intranasal , Animais , Líquido da Lavagem Broncoalveolar/citologia , Líquido da Lavagem Broncoalveolar/imunologia , Interleucina-6/imunologia , Lipopolissacarídeos , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Masculino , Camundongos Endogâmicos BALB C , Tetra-Hidroisoquinolinas/farmacologia , Fator de Transcrição RelA/imunologia , Fator de Necrose Tumoral alfa/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
N-Formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF) induce similar intracellular signalling profiles; but only fMLP induces interleukin-8 (IL-8) release and nicotinamide adenine dinucleotide phosphate reduced (NADPH) oxidase activity in neutrophils. Because the role of ROS on IL-8 release in neutrophils is until now controversial, we assessed if NADPH oxidase is involved in the IL-8 secretions and PI3K/Akt, MAPK, and NF-κB pathways activity induced by fMLP. Neutrophils were obtained from healthy volunteers. IL-8 was measured by ELISA, IL-8 mRNA by qPCR, and ROS production by luminol-amplified chemiluminescence, reduction of ferricytochrome c, and FACS. Intracellular pH changes were detected by spectrofluorescence. ERK1/2, p38 MAPK, and Akt phosphorylation were analysed by immunoblotting and NF-κB was analysed by immunocytochemistry. Hydroxy-3-methoxyaceto-phenone (HMAP), diphenyleneiodonium (DPI), and siRNA Nox2 reduced the ROS and IL-8 release in neutrophils treated with fMLP. HMAP, DPI, and amiloride (a Na(+)/H(+) exchanger inhibitor) inhibited the Akt phosphorylation and did not affect the p38 MAPK and ERK1/2 activity. DPI and HMAP reduced NF-κB translocation induced by fMLP. We showed that IL-8 release induced by fMLP is dependent on NADPH oxidase, and ROS could play a redundant role in cell signalling, ultimately activating the PI3K/Akt and NF-κB pathways in neutrophils.
Assuntos
Interleucina-8/metabolismo , Glicoproteínas de Membrana/genética , N-Formilmetionina Leucil-Fenilalanina/farmacologia , NADPH Oxidases/genética , Neutrófilos/efeitos dos fármacos , Espécies Reativas de Oxigênio/imunologia , Transdução de Sinais/efeitos dos fármacos , Acetofenonas/farmacologia , Amilorida/farmacologia , Bloqueadores do Canal de Sódio Epitelial/farmacologia , Regulação da Expressão Gênica , Humanos , Interleucina-8/genética , Interleucina-8/imunologia , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/imunologia , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/imunologia , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/imunologia , NADPH Oxidase 2 , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Neutrófilos/citologia , Neutrófilos/imunologia , Oniocompostos/farmacologia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/imunologia , Fosforilação/efeitos dos fármacos , Cultura Primária de Células , Transporte Proteico/efeitos dos fármacos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Leishmanioses are chronic parasitic diseases and host responses are associated with pro- or anti-inflammatory cytokines involved, respectively, in the control or exacerbation of infection. The relevance of other inflammatory mediators and stress markers has not been widely studied and there is a need to search for biomarkers to leishmaniasis. In this work, the stress and inflammatory molecules p38 mitogen-activated protein kinase, cyclooxygenase-2, migration inhibitory factor, macrophage inflammatory protein 2, heat shock protein 70 kDa, vascular endothelial factor (VEGF), hypoxia-inducible factors (HIF-1α and HIF-2α), heme oxygenase and galectin-3 expression were assessed immunohistochemically in self-controlled lesions in C57BL/6 mice and severe lesions in Balb/c mice infected with Leishmania amazonensis. The results indicated that the majority of molecules were expressed in the cutaneous lesions of both C57BL/6 and Balb/c mice during various phases of infection, suggesting no obvious correlation between the stress and inflammatory molecule expression and the control/exacerbation of leishmanial lesions. However, the cytokine VEGF was only detected in C57BL/6 footpad lesions and small lesions in Balb/c mice treated with antimonial pentavalent. These findings suggest that VEGF expression could be a predictive factor for murine leishmanial control, a hypothesis that should be tested in human leishmaniosis.
Assuntos
Ciclo-Oxigenase 2/imunologia , Citocinas/imunologia , Inflamação/imunologia , Leishmaniose Cutânea/imunologia , Fator A de Crescimento do Endotélio Vascular/imunologia , Animais , Biomarcadores/análise , Ciclo-Oxigenase 2/biossíntese , Modelos Animais de Doenças , Imuno-Histoquímica , Leishmania/imunologia , Leishmaniose Cutânea/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Óxido Nítrico/biossíntese , Nitroimidazóis/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/biossíntese , Proteínas Quinases p38 Ativadas por Mitógeno/biossíntese , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
The observation that Th17 infiltration in ovarian cancer correlates with markedly improved survival has prompted the question of whether ovarian tumor antigen-specific Th17 responses could be stimulated by tumor vaccination. Dendritic cells (DCs) treated with IL-15 and an inhibitor of p38 MAPK signaling (DC(IL-15/p38inhib)) bias T-cell responses toward a Th1/Th17 phenotype, raising the prospect of therapeutic vaccination; however, significant barriers remain. Tumor vaccines, including DC vaccination, usually stimulate immune responses, but the lack of clinical responses in cancer patients has been disappointing. Possible reasons may include an inability of antitumor T cells to migrate into the tumor microenvironment, and an inability of T cells to retain effector function in the face of tumor-associated immune suppression. We found that ovarian tumor antigen-specific CD4(+) T cells induced by DC(IL-15/p38inhib) migrated in response to CXCL12 and CCL22 (both highly expressed in ovarian cancer) and to ascites CD14(+) myeloid cells. Cocultures showed that ascites CD14(+) cells markedly suppressed antigen-specific CD4(+) T responses, but suppression could be alleviated by treatment with anti-IL-10 or inhibition of indoleamine 2,3-dioxygenase. These results suggest that the efficacy of DC vaccination against ovarian cancer may be boosted by agents that inhibit tumor-associated CD14(+) myeloid cell suppression or indoleamine 2,3-dioxygenase activity.
Assuntos
Ascite/imunologia , Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Indolamina-Pirrol 2,3,-Dioxigenase/imunologia , Receptores de Lipopolissacarídeos/imunologia , Neoplasias Ovarianas/imunologia , Antígenos de Neoplasias/imunologia , Vacinas Anticâncer , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/imunologia , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Leishmania (L.) chagasi is the etiologic agent of visceral leishmaniasis (VL) that can be transmitted to humans and dogs. VL in Brazil represents a serious public health problem; therefore, it is important to study new alternatives to treat infected dogs. In dogs, the therapeutic arsenal against canine VL is limited. The immunomodulator protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride (P-MAPA) improves immunocompetence when the immune system is impaired, but its dependence on Toll-like receptors (TLRs) and the mechanisms involved in immune response remain unclear. The in vitro action of P-MAPA on the expression of TLR2 and TLR4, reactive oxygen species (ROS), nitric oxide (NO) and p38 mitogen-activated protein kinase (p38 MAPK) and IKK phosphorylation was studied in mononuclear cells from peripheral blood and macrophages from healthy and Leishmania-infected dogs. The PBMC or macrophages were isolated and cultured with different concentrations of P-MAPA (20,100 and 200 µg/ml) in a humid environment at 37°C with 5% CO(2). Observation revealed that Leishmania-infected dogs showed a decrease in TLR2 in macrophages compared with healthy dogs and in induction with P-MAPA. ROS were increased in PBMCs from Leishmania spp.-infected dogs compared with healthy dogs and P-MAPA improved ROS production. NO production was increased in culture supernatant from macrophages stimulated by P-MAPA in both healthy and Leishmania spp. infected dogs. Treatment of macrophages from healthy dogs with immunomodulatory P-MAPA induced p38 MAPK and IKK phosphorylation, suggesting signal transduction by this pathway. These findings suggest that P-MAPA has potential as a therapeutic drug in the treatment of canine visceral leishmaniasis.
Assuntos
Doenças do Cão/imunologia , Fatores Imunológicos/farmacologia , Leishmaniose Visceral/imunologia , Leucócitos Mononucleares/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Animais , Cães , Feminino , Fatores de Transcrição Kruppel-Like/imunologia , Leishmaniose Visceral/veterinária , Leucócitos Mononucleares/imunologia , Macrófagos/imunologia , Masculino , Óxido Nítrico/imunologia , Espécies Reativas de Oxigênio/imunologia , Receptor 2 Toll-Like/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Curcumin is the active compound in the extract of Curcuma longa rhizomes with anti-inflammatory properties mediated by inhibition of intracellular signalling. SOCS and MAPKinases are involved in the signalling events controlling the expression of IL-6, TNF-α and PGE2, which have important roles on chronic inflammatory diseases. The aim was to assess if these pathways are involved in curcumin-mediated effects on LPS-induced expression of these cytokines in macrophages. RAW 264.7 murine macrophages were stimulated with Escherichia coli LPS in the presence and absence of non-cytotoxic concentrations of curcumin. Curcumin potently inhibited LPS-induced expression of IL-6, TNF-α and COX-2 mRNA and prevented LPS-induced inhibition of SOCS-1 and -3 expression and the inhibition of the activation of p38 MAPKinase by modulation of its nuclear translocation. In conclusion, curcumin potently inhibits expression of LPS-induced inflammatory cytokines in macrophages via mechanisms that involve modulation of expression and activity of SOCS-1 and SOCS-3 and of p38 MAPK.
Assuntos
Curcumina/farmacologia , Imunidade Inata , Macrófagos/imunologia , Proteínas Supressoras da Sinalização de Citocina/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Animais , Ciclo-Oxigenase 2/imunologia , Citocinas/imunologia , Interleucina-6/imunologia , Lipopolissacarídeos/farmacologia , Camundongos , RNA Mensageiro/imunologia , Transdução de Sinais/imunologia , Proteína 1 Supressora da Sinalização de Citocina , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Tuberculosis (TB) is one of the world's most pernicious diseases mainly due to immune evasion strategies displayed by its causative agent Mycobacterium tuberculosis (Mtb). Blood monocytes (Mos) represent an important source of DCs during chronic infections; consequently, the alteration of their differentiation constitutes an escape mechanism leading to mycobacterial persistence. We evaluated whether the CD16(+)/CD16(-) Mo ratio could be associated with the impaired Mo differentiation into DCs found in TB patients. The phenotype and ability to stimulate Mtb-specific memory clones DCs from isolated Mo subsets were assessed. We found that CD16(-) Mos differentiated into CD1a(+) DC-SIGN(high) cells achieving an efficient recall response, while CD16(+) Mos differentiated into a CD1a(-) DC-SIGN(low) population characterized by a poor mycobacterial Ag-presenting capacity. The high and sustained phosphorylated p38 expression observed in CD16(+) Mos was involved in the altered DC profile given that its blockage restored DC phenotype and its activation impaired CD16(-) Mo differentiation. Furthermore, depletion of CD16(+) Mos indeed improved the differentiation of Mos from TB patients toward CD1a(+) DC-SIGN(high) DCs. Therefore, Mos from TB patients are less prone to differentiate into DCs due to their increased proportion of CD16(+) Mos, suggesting that during Mtb infection Mo subsets may have different fates after entering the lungs.