RESUMO
Chromoblastomycosis is a chronic and progressive subcutaneous mycosis caused mainly by the fungus Fonsecaea pedrosoi. The infection is characterized by erythematous papules and histological sections demonstrating an external layer of fibrous tissue and an internal layer of thick granulomatous inflammatory tissue containing mainly macrophages and neutrophils. Several groups are studying the roles of the innate and adaptive immune systems in F. pedrosoi infection; however, few studies have focused on the role of neutrophils in this infection. In the current study, we verify the importance of murine neutrophils in the killing of F. pedrosoi conidia and hyphae. We demonstrate that phagocytosis and reactive oxygen species during infection with conidia are TLR-2- and TLR-4-dependent and are essential for conidial killing. Meanwhile, hyphal killing occurs by NET formation in a TLR-2-, TLR-4-, and ROS-independent manner. In vivo experiments show that TLR-2 and TLR-4 are also important in chromoblastomycosis infection. TLR-2KO and TLR-4KO animals had lower levels of CCL3 and CXCL1 chemokines and impaired neutrophil migration to the infected site. These animals also had higher fungal loads during infection with F. pedrosoi conidia, confirming that TLR-2 and TLR-4 are essential receptors for F. pedrosoi recognition and immune system activation. Therefore, this study demonstrates for the first time that neutrophil activation during F. pedrosoi is conidial or hyphal-specific with TLR-2 and TLR-4 being essential during conidial infection but unnecessary for hyphal killing by neutrophils.
Assuntos
Cromoblastomicose/imunologia , Fonsecaea/imunologia , Hifas/imunologia , Neutrófilos/imunologia , Esporos Fúngicos/imunologia , Receptor 2 Toll-Like/imunologia , Receptor 4 Toll-Like/imunologia , Animais , Quimiocina CCL3/genética , Quimiocina CCL3/imunologia , Quimiocina CXCL1/genética , Quimiocina CXCL1/imunologia , Cromoblastomicose/genética , Cromoblastomicose/patologia , Camundongos , Camundongos Knockout , Neutrófilos/patologia , Receptor 2 Toll-Like/genética , Receptor 4 Toll-Like/genéticaRESUMO
OBJECTIVE: This study aims to investigate the role of protease-activated receptor (PAR) 2 and mast cell (MC) tryptase in LPS-induced lung inflammation and neutrophil recruitment in the lungs of C57BL/6 mice. METHODS: C57BL/6 mice were pretreated with the PAR2 antagonist ENMD-1068, compound 48/80 or aprotinin prior to intranasal instillation of MC tryptase or LPS. Blood leukocytes, C-X-C motif chemokine ligand (CXCL) 1 production leukocytes recovered from bronchoalveolar lavage fluid (BALF), and histopathological analysis of the lung were evaluated 4 h later. Furthermore, we performed experiments to determine intracellular calcium signaling in RAW 264.7 cells stimulated with LPS in the presence or absence of a protease inhibitor cocktail or ENMD-1068 and evaluated PAR2 expression in the lungs of LPS-treated mice. RESULTS: Pharmacological blockade of PAR2 or inhibition of proteases reduced neutrophils recovered in BALF and LPS-induced calcium signaling. PAR2 blockade impaired LPS-induced lung inflammation, PAR2 expression in the lung and CXCL1 release in BALF, and increased circulating blood neutrophils. Intranasal instillation of MC tryptase increased the number of neutrophils recovered in BALF, and MC depletion with compound 48/80 impaired LPS-induced neutrophil migration. CONCLUSION: Our study provides, for the first time, evidence of a pivotal role for MCs and MC tryptase in neutrophil migration, lung inflammation and macrophage activation triggered by LPS, by a mechanism dependent on PAR2 activation.
Assuntos
Mastócitos/imunologia , Infiltração de Neutrófilos , Pneumonia/imunologia , Receptor PAR-2/imunologia , Triptases/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Sinalização do Cálcio , Quimiocina CXCL1/imunologia , Feminino , Lipopolissacarídeos , Pulmão/imunologia , Pulmão/patologia , Ativação de Macrófagos , Camundongos , Camundongos Endogâmicos C57BL , Piperazinas/farmacologia , Pneumonia/induzido quimicamente , Pneumonia/patologia , Células RAW 264.7 , Receptor PAR-2/antagonistas & inibidoresRESUMO
BACKGROUND: ß-Blockers reduce temporomandibular joint (TMJ) pain. We asked whether they also reduce TMJ inflammation and, if so, whether this anti-inflammatory effect contributes to its analgesic action. METHODS: We measured many parameters of the inflammatory response after co-administration of the ß-blocker propranolol with the inflammatory agent carrageenan in the TMJ of female rats. We also hypothesized that the activation of ß-adrenoceptors in the TMJ induces nociception mediated, at least in part, by the inflammatory response. To test this hypothesis, we examined the nociceptive response induced by the activation of the ß-adrenoceptors in the TMJ in female rats pretreated with thalidomide and fucoidan. RESULTS: We found that the co-administration of propranolol with carrageenan in the TMJ of female rats significantly reduced several parameters of the inflammatory response induced by carrageenan such as plasma extravasation, neutrophil migration and the release of the pro-inflammatory cytokines TNF-α, IL-1ß and CINC-1. Furthermore, the injection of the ß-adrenergic receptor agonist isoproterenol in the TMJ induced nociception that was significantly reduced by thalidomide, fucoidan and by the co-administration of propranolol but not of the α-adrenergic receptor antagonist phentolamine. CONCLUSIONS: Propranolol has anti-inflammatory effects that contribute to its antinociceptive action in the TMJ of females. SIGNIFICANCE: ß-Blockers have an anti-inflammatory effect on temporomandibular joint (TMJ) that contributes to its analgesic effect. The results of this work suggest that ß-blockers can be used to treat the painful conditions of TMJ, especially when they are associated with an inflammatory process.
Assuntos
Antagonistas Adrenérgicos beta/farmacologia , Nociceptividade/efeitos dos fármacos , Propranolol/farmacologia , Articulação Temporomandibular/efeitos dos fármacos , Antagonistas Adrenérgicos alfa/farmacologia , Analgésicos/uso terapêutico , Animais , Anti-Inflamatórios/uso terapêutico , Anticoagulantes/farmacologia , Carragenina/farmacologia , Quimiocina CXCL1/efeitos dos fármacos , Quimiocina CXCL1/imunologia , Feminino , Imunossupressores/farmacologia , Interleucina-1beta/efeitos dos fármacos , Interleucina-1beta/imunologia , Dor/tratamento farmacológico , Medição da Dor/efeitos dos fármacos , Fentolamina/farmacologia , Polissacarídeos/farmacologia , Ratos , Ratos Wistar , Articulação Temporomandibular/imunologia , Transtornos da Articulação Temporomandibular/tratamento farmacológico , Talidomida/farmacologiaRESUMO
AIMS: Inflammation induced by hyperglycemia triggers the toll-like receptor (TLR) pathway into cells. Our hypothesis was that metformin treatment attenuates the TLR signaling pathways triggered by inflammation in skeletal muscle of hypoinsulinemic/hyperglycemic STZ-induced rats. Thus, we examined TLR signaling under hypoinsulinemia and hyperglycemia conditions and its correlation with insulin resistance in muscle of diabetic rats treated with metformin. METHODS: Ten-day diabetic rats were submitted to 7 days of saline (D group) or metformin (500 mg/kg once per day) (D + M group). The skeletal muscle was collected before the insulin tolerance test. Then, Western blotting analysis of skeletal muscle supernatant was probed with TLR4, TLR2, NF-κB, IκB, p-AMPK and p-JNK. TNF-α and CXCL1/KC content was analyzed by ELISA. RESULTS: Metformin treatment increased whole-body insulin sensitivity. This regulation was accompanied by a parallel change of p-AMPK and by an inverse regulation of TLR4 and NF-κB contents in the soleus muscle (r = 0.7229, r = -0.8344 and r = -0.7289, respectively, Pearson correlation; p < 0.05). Metformin treatment increased IκB content when compared to D rats. In addition, metformin treatment decreased p-JNK independently of TLR2 signal in diabetic rats. CONCLUSION: In summary, the results indicate a relationship between muscular TLR4, p-AMPK and NF-κB content and insulin sensitivity. The study also highlights that in situations of insulin resistance, such as in diabetic subjects, metformin treatment may prevent attenuation of activation of the inflammatory pathway.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Hipoglicemiantes/administração & dosagem , Metformina/administração & dosagem , Músculo Esquelético/efeitos dos fármacos , Receptor 4 Toll-Like/imunologia , Animais , Quimiocina CXCL1/genética , Quimiocina CXCL1/imunologia , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/imunologia , Humanos , Resistência à Insulina , Masculino , Músculo Esquelético/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Ratos , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Receptor 4 Toll-Like/genética , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
UNLABELLED: BACKGROUND: Uncontrolled hapatic inflammatory response is regarded as the primary pathological mechanism of acute liver failure and impairs the regeneration of hepatocytes and stem cell grafts. Interleukin-1 plays a key role for activating immune and inflammatory response. Recently, siRNA has made quite a few progresses in treating inflammatory response. AIM: To assess the effect of IL-1? siRNA adenovirus on MSC and the therapeutic effect of MSC combined with IL-1? siRNA adenovirus in ALF. MATERIAL AND METHODS: We implanted MSC or/and IL-1? siRNA adenovirus via the tail vein, using CCl4-induced ALF in a mice model. Mice were sacrificed at different time points. Blood samples and liver tissues were collected. Hepatic injury, liver regeneration, cytokines (CXCL1, IL-1?, IL-10, IL-6, VEGF and HGF), animal survival and vital MSC were assessed after cell transplantation. RESULTS: MSC combined with IL-1? siRNA reduced the inflammatory levels and prevented liver failure. These animals administrated with MSC and IL-1? siRNA also exhibited improved liver regeneration and increased survival rates. Immunohistochemistry and fluorescence microscopy revealed the number of vital MSC in ALF + MSC + IL-1? siRNA group were significantly more than that in ALF + MSC group. CONCLUSION: IL-1? siRNA adenovirus could enhance MSC ability of tissue regeneration through increasing its survival rate. Accordingly, combination of IL-1? siRNA adenovirus and MSC had a synergistic effect on acute liver failure.
Assuntos
Interleucina-1beta/efeitos dos fármacos , Falência Hepática Aguda , Regeneração Hepática/efeitos dos fármacos , Fígado/efeitos dos fármacos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/efeitos dos fármacos , RNA Interferente Pequeno/farmacologia , Adenoviridae , Animais , Quimiocina CXCL1/efeitos dos fármacos , Quimiocina CXCL1/imunologia , Fator de Crescimento de Hepatócito/metabolismo , Inflamação , Interleucina-10/imunologia , Interleucina-6/imunologia , Fígado/imunologia , Fígado/metabolismo , Regeneração Hepática/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Terapêutica com RNAi , Fator A de Crescimento do Endotélio Vascular/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
OBJECTIVE AND DESIGN: The activation of proteinase-activated receptors (PARs) has been implicated in the development of important hallmarks of inflammation, including in vivo leukocyte recruitment; however, its role in the regulation of leukocyte migration in response to inflammatory stimuli has not been elucidated until now. Here, we examined the effects of the PAR4 antagonist YPGKF-NH 2 (tcY-NH2) on neutrophil recruitment in experimentally induced inflammation. METHODS: BALB/c mice were intrapleurally injected with tcY-NH2 (40 ng/kg) prior to intrapleural injection of carrageenan (Cg) or neutrophil chemoattractant CXCL8; the number of infiltrating neutrophils was evaluated after 4 h, and KC production was assessed at different times after Cg injection. Neutrophil adhesion and rolling cells were studied using a brain circulation preparation 4 h after the Cg or CXCL8 challenge in tcY-NH2-treated mice. RESULTS: PAR4 blockade inhibited CXCL8- and Cg-induced neutrophil migration into the pleural cavity of BALB/c mice and reduced neutrophil rolling and adherence. Surprisingly, PAR4 blockade increased the level of KC in response to carrageenan. CONCLUSION: These results demonstrated that PAR4 blockade impairs neutrophil migration in vivo, suggesting that PAR4 plays an important role in the regulation of inflammation, at least in part because of its ability to inhibit the actions of the neutrophil chemoattractant CXCL8.
Assuntos
Infiltração de Neutrófilos/efeitos dos fármacos , Oligopeptídeos/farmacologia , Receptores de Trombina/antagonistas & inibidores , Animais , Carragenina , Quimiocina CXCL1/imunologia , Modelos Animais de Doenças , Feminino , Inflamação/induzido quimicamente , Inflamação/imunologia , Interleucina-8 , Camundongos Endogâmicos BALB CRESUMO
OBJECTIVE: Interleukin-4 (IL-4) is a multifunctional cytokine involved in many diseases such as autoimmune hepatitis and idiosyncratic drug reactions. However, its role in acetaminophen (APAP)-induced liver injury remains unclear. Our objective was to evaluate the contribution of IL-4 to the pathogenesis of APAP-induced liver injury. METHODS: Balb/C (WT) and IL-4 knockout (IL-4(-/-)) mice were orally overdosed with APAP. After 24 h, survival percentage, biochemical and morphological markers of liver injury, and tissue inflammation were assessed. RESULTS: IL-4(-/-) mice were protected from APAP toxicity. Intravital confocal microscopy, tissue histology and serum ALT levels showed significantly less liver injury and inflammation than in the WT group, which may explain the increased survival rate of IL-4(-/-) mice. In addition, IL-4(-/-) mice had decreased production of tumor necrosis factor α, CXCL1 and interleukin-1ß in the liver, but not in a remote site such as the lungs. Hepatic macrophage activation was markedly reduced in IL-4-deficient mice. In addition, glutathione depletion-a primary cause of APAP-mediated injury-was significantly attenuated in IL-4(-/-) mice. CONCLUSIONS: Taken together, our data demonstrate that IL-4(-/-) mice are protected from APAP-induced liver injury due to reduced depletion of glutathione, which prevented liver damage and tissue inflammation.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/imunologia , Glutationa/imunologia , Interleucina-4/imunologia , Acetaminofen , Animais , Doença Hepática Induzida por Substâncias e Drogas/patologia , Quimiocina CXCL1/imunologia , Inflamação/imunologia , Interleucina-4/genética , Fígado/imunologia , Fígado/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Fator de Necrose Tumoral alfa/imunologiaRESUMO
BACKGROUND AND PURPOSE: Chemokines orchestrate neutrophil recruitment to inflammatory foci. In the present study, we evaluated the participation of three chemokines, KC/CXCL1, MIP-2/CXCL2 and LIX/CXCL5, which are ligands for chemokine receptor 2 (CXCR2), in mediating neutrophil recruitment in immune inflammation induced by antigen in immunized mice. EXPERIMENTAL APPROACH: Neutrophil recruitment was assessed in immunized mice challenged with methylated bovine serum albumin, KC/CXCL1, LIX/CXCL5 or tumour necrosis factor (TNF)-alpha. Cytokine and chemokine levels were determined in peritoneal exudates and in supernatants of macrophages and mast cells by elisa. CXCR2 and intercellular adhesion molecule 1 (ICAM-1) expression was determined using immunohistochemistry and confocal microscopy. KEY RESULTS: Antigen challenge induced dose- and time-dependent neutrophil recruitment and production of KC/CXCL1, LIX/CXCL5 and TNF-alpha, but not MIP-2/CXCL2, in peritoneal exudates. Neutrophil recruitment was inhibited by treatment with reparixin (CXCR1/2 antagonist), anti-KC/CXCL1, anti-LIX/CXCL5 or anti-TNF-alpha antibodies and in tumour necrosis factor receptor 1-deficient mice. Intraperitoneal injection of KC/CXCL1 and LIX/CXCL5 induced dose- and time-dependent neutrophil recruitment and TNF-alpha production, which were inhibited by reparixin or anti-TNF-alpha treatment. Macrophages and mast cells expressed CXCR2 receptors. Increased macrophage numbers enhanced, while cromolyn sodium (mast cell stabilizer) diminished, LIX/CXCL5-induced neutrophil recruitment. Macrophages and mast cells from immunized mice produced TNF-alpha upon LIX/CXCL5 stimulation. Methylated bovine serum albumin induced expression of ICAM-1 on mesenteric vascular endothelium, which was inhibited by anti-TNF-alpha or anti-LIX/CXCL5. CONCLUSION AND IMPLICATIONS: Following antigen challenge, CXCR2 ligands are produced and act on macrophages and mast cells triggering the production of TNF-alpha, which synergistically contribute to neutrophil recruitment through induction of the expression of ICAM-1.
Assuntos
Quimiocina CXCL1/imunologia , Quimiocina CXCL5/imunologia , Neutrófilos/imunologia , Peritonite/imunologia , Fator de Necrose Tumoral alfa/fisiologia , Animais , Anticorpos/farmacologia , Bovinos , Quimiocina CXCL1/farmacologia , Quimiocina CXCL5/farmacologia , Molécula 1 de Adesão Intercelular/biossíntese , Macrófagos Peritoneais/imunologia , Macrófagos Peritoneais/metabolismo , Mastócitos/imunologia , Mastócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Peritonite/metabolismo , Receptores de Interleucina-8A/antagonistas & inibidores , Receptores de Interleucina-8B/antagonistas & inibidores , Receptores de Interleucina-8B/biossíntese , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Albumina Sérica/imunologia , Sulfonamidas/farmacologia , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/imunologiaRESUMO
OBJECTIVE: To investigate the mechanism underlying neutrophil migration into the articular cavity in experimental arthritis and, by extension, human inflammatory synovitis. METHODS: Antigen-induced arthritis (AIA) was generated in mice with methylated bovine serum albumin (mBSA). Migration assays and histologic analysis were used to evaluate neutrophil recruitment to knee joints. Levels of inflammatory mediators were measured by enzyme-linked immunosorbent assay. Antibodies and pharmacologic inhibitors were used in vivo to determine the role of specific disease mediators. Samples of synovial tissue and synovial fluid from rheumatoid arthritis (RA) or osteoarthritis patients were evaluated for CXCL1 and CXCL5 expression. RESULTS: High levels of CXCL1, CXCL5, and leukotriene B4 (LTB4) were expressed in the joints of arthritic mice. Confirming their respective functional roles, repertaxin (a CXCR1/CXCR2 receptor antagonist), anti-CXCL1 antibody, anti-CXCL5 antibody, and MK886 (a leukotriene synthesis inhibitor) reduced mBSA-induced neutrophil migration to knee joints. Repertaxin reduced LTB4 production in joint tissue, and neutrophil recruitment induced by CXCL1 or CXCL5 was inhibited by MK886, suggesting a sequential mechanism. Levels of both CXCL1 and CXCL5 were elevated in synovial fluid and were released in vitro by RA synovial tissues. Moreover, RA synovial fluid neutrophils stimulated with CXCL1 or CXCL5 released significant amounts of LTB4. CONCLUSION: Our data implicate CXCL1, CXCL5, and LTB4, acting sequentially, in neutrophil migration in AIA. Elevated levels of CXCL1 and CXCL5 in the synovial compartment of RA patients provide robust comparative data indicating that this mechanism plays a role in inflammatory joint disease. Together, these results suggest that inhibition of CXCL1, CXCL5, or LTB4 may represent a potential therapeutic strategy in RA.
Assuntos
Artrite/imunologia , Quimiotaxia de Leucócito/imunologia , Leucotrieno B4/imunologia , Neutrófilos/imunologia , Receptores de Interleucina-8B/imunologia , Animais , Quimiocina CXCL1/imunologia , Quimiocina CXCL5/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Articulação do Joelho/imunologia , Articulação do Joelho/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
In the present study, we used the electronic version of the von Frey test to investigate the role of cytokines (TNF-alpha and IL-1beta) and chemokines (KC/CXCL-1) in the genesis of mechanical hypernociception during antigen-induced inflammation in mice. The nociceptive test consisted of evoking a hindpaw flexion reflex with a hand-held force transducer (electronic anesthesiometer) adapted with a 0.5 mm(2) polypropylene tip. The intraplantar administration of methylated bovine serum albumin (mBSA) in previously immunized (IM), but not in sham-immunized (SI) mice, induced mechanical hypernociception in a dose-dependent manner. Hypernociception induced by antigen was reduced in animals pretreated with IL-1ra and reparixin (a non-competitive allosteric inhibitor of CXCR2), and in TNF receptor type 1 deficient (TNFR1-/-) mice. Consistently, antigen challenge induced a time-dependent release of TNF-alpha, IL-1beta and KC/CXCL-1 in IM, but not in SI, mice. The increase in TNF-alpha levels preceded the increase in IL-1beta and KC/CXCL1. Antigen-induced release of IL-1beta and KC/CXCL1 was reduced in TNFR1-/- mice, and TNF-alpha-induced hypernociception was inhibited by IL-1ra and reparixin. Hypernociception induced by IL-1beta in immunized mice was inhibited by indomethacin, whereas KC/CXCL1-induced hypernociception was inhibited by indomethacin and guanethidine. Antigen-induced hypernociception was reduced by indomethacin and guanethidine and abolished by the two drugs combined. Together, these results suggest that inflammation associated with an adaptive immune response induces hypernociception that is mediated by an initial release of TNF-alpha, which triggers the subsequent release of IL-1beta and KC/CXCL1. The latter cytokines in turn stimulate the release of the direct-acting final mediators, prostanoids and sympathetic amines.