RESUMO
Lipid bodies (LBs) are intracellular accumulations of neutral lipids surrounded by a single membrane. These organelles are involved in the production of eicosanoids, which modulate immunity by either promoting or dampening inflammatory responses. Leishmania infantum, the etiological agent of visceral leishmaniasis in Brazil, is an intracellular parasite that causes disease by suppressing macrophage microbicidal responses. C57BL/6 mouse bone marrow-derived macrophages infected with L. infantum strain LcJ had higher numbers of LB+ cells (P<.0001) and total LBs than noninfected cultures. Large (>3 µm) LBs were present inside parasitophorous vacuoles (PVs). These results contrast with those of L. infantum-infected BALB/c macrophages, in which the only LBs are derived from parasite, not macrophage origin. Increased LBs in C57BL/6 macrophages in close association with parasites would position host LBs where they could modulate L. infantum infection. These results imply a potential influence of the host genetics on the role of LBs in host-pathogen interactions. Overall, our data support a model in which the expression, and the role of LBs upon infection, ultimately depends on the specific combination of host-pathogen interactions.
Assuntos
Leishmania infantum/imunologia , Leishmaniose Visceral/imunologia , Gotículas Lipídicas/metabolismo , Macrófagos/microbiologia , Animais , Brasil , Feminino , Leishmaniose Visceral/patologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BLRESUMO
Sepsis is a severe disorder characterized by systemic inflammatory responses in the presence of an infection and may progress to multiple organ dysfunction and death. Alterations in cerebral microcirculation fulfill a crucial role in the pathogenesis of severe sepsis, and include a decrease in capillary density and disturbances in leukocyte movement along capillaries. Nevertheless, the mechanisms involved in sepsis-associated cerebral microcirculatory alterations have so far not been defined. We investigated the effect of the peroxisome proliferator-activated receptor gamma (PPARγ) selective agonist rosiglitazone on leukocyte/endothelial cell interaction and functional capillary density in the brain in the cecal ligation and puncture (CLP) model of sepsis. Anti-inflammatory effects of rosiglitazone on the cerebral microcirculation were marked. Functional capillary density increased and leukocyte rolling and adhesion were decreased in animals submitted to CLP and treated with rosiglitazone. Our data provide evidence for involvement of PPARγ activation in leukocyte-endothelium interactions and alterations in capillary density. Improved cerebral perfusion in animals treated with rosiglitazone, suggests that PPARγ activation is protective against cerebral microvascular dysfunction in sepsis.
Assuntos
Anti-Inflamatórios/farmacologia , Encéfalo/irrigação sanguínea , Circulação Cerebrovascular/efeitos dos fármacos , Transtornos Cerebrovasculares/prevenção & controle , Microcirculação/efeitos dos fármacos , Microvasos/efeitos dos fármacos , PPAR gama/agonistas , Sepse/tratamento farmacológico , Tiazolidinedionas/farmacologia , Animais , Ceco/cirurgia , Transtornos Cerebrovasculares/imunologia , Transtornos Cerebrovasculares/metabolismo , Transtornos Cerebrovasculares/fisiopatologia , Citoproteção , Modelos Animais de Doenças , Migração e Rolagem de Leucócitos/efeitos dos fármacos , Ligadura , Masculino , Camundongos , Microvasos/imunologia , Microvasos/metabolismo , Microvasos/fisiopatologia , Infiltração de Neutrófilos/efeitos dos fármacos , PPAR gama/metabolismo , Punções , Rosiglitazona , Sepse/imunologia , Sepse/metabolismo , Sepse/fisiopatologiaRESUMO
Esophageal cancer (EC) is an aggressive disease, presenting two main histological subtypes: adenocarcinoma (EAC) and squamous cell carcinoma (ESCC). The two EC subtypes widely differ concerning virtually all factors. ESCC development is mainly associated with tobacco and alcohol abuse, whereas obesity and chronic gastroesophageal reflux disease (GERD) are important risk factors not only for EAC, but also for for Barrett's esophagus (BE), an intestinal metaplasia that precedes EAC. Obesity triggers ectopic lipid droplets (LD) accumulation in non-adipose tissues. LD are organelles involved in cell metabolism, signaling, proliferation and production of inflammatory mediators. Therefore, the aim of this work was to investigate LD occurrence and role in EC. This study shows progressive LD levels increase along EAC development, in esophageal samples from non-obese through obese individuals, as well as BE, and EAC patients, whereas no significant changes were observed in ESCC samples, when compared to non-tumor samples. Additionally, in order to mimic BE and EAC risk factors exposure, a non-tumor esophageal cell line was incubated with oleic acid (OA) and acidified medium and/or deoxycholic acid (DCA), revealing a significant increment in LD amount as well as in COX-2 and CXCL-8 expression, and in IL-8 secretion. Further, COX-2 expression and LD amount presented a significant positive correlation and were detected co-localized in EAC, but not in ESCC, suggesting that LD may be the site for eicosanoid production in EAC. In conclusion, this study shows that obesity, and BE- and EAC-associated inflammatory stimuli result in a gradual increase of LD, that may be responsible for orchestrating inflammatory mediators' production and/or action, thus contributing to BE and EAC genesis and progression.
Assuntos
Adenocarcinoma/metabolismo , Esôfago de Barrett/metabolismo , Ciclo-Oxigenase 2/metabolismo , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas do Esôfago/metabolismo , Gotículas Lipídicas/metabolismo , Transdução de Sinais/fisiologia , Adenocarcinoma/patologia , Esôfago de Barrett/patologia , Linhagem Celular , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/patologia , Esôfago/metabolismo , Esôfago/patologia , Refluxo Gastroesofágico/metabolismo , Refluxo Gastroesofágico/patologia , Humanos , Inflamação/metabolismo , Inflamação/patologia , Fatores de RiscoRESUMO
Lipid bodies, lipid rich cytoplasmic inclusions, are characteristically abundant in vivo in leukocytes associated with inflammation. Because lipid bodies are potential reservoirs of esterified arachidonate and sites at which eicosanoid-forming enzymes may localize, we evaluated mechanisms of lipid body formation in neutrophils (PMN). Among receptor-mediated agonists, platelet activating factor (PAF), but not C5a, formyl-methyl-phenylalanine, interleukin 8, or leukotriene (LT) B4, induced the rapid formation of lipid bodies in PMN. This action of PAF was receptor mediated, as it was dose dependently inhibited by the PAF receptor antagonist WEB 2086 and blocked by pertussis toxin. Lipid body induction by PAF required 5-lipoxygenase (LO) activity and was inhibited by the 5-lipoxygenase-activating protein antagonist MK 886 and the 5-LO inhibitor zileuton, but not by cyclooxygenase inhibitors. Corroborating the dependency of PAF-induced lipid body formation on 5-LO, PMN and macrophages from wild-type mice, but not from 5-LO genetically deficient mice, formed lipid bodies on exposure to PAF both in vitro and in vivo within the pleural cavity. The 5-LO product inducing lipid body formation was not LTB4 but was 5(S)-hydroxyeicosatetraenoic acid [5(S)-HETE], which was active at 10-fold lower concentrations than PAF and was also inhibited by pertussis toxin but not by zileuton or WEB 2086. Furthermore, 5-HETE was equally effective in inducing lipid body formation in both wild-type and 5-LO genetically deficient mice. Both PAF- and 5(S)-HETE-induced lipid body formation were inhibited by protein kinase C (PKC) inhibitors staurosporine and chelerythrine, the phospholipase C (PLC) inhibitors D609 and U-73122, and by actinomycin D and cycloheximide. Prior stimulation of human PMN with PAF to form lipid bodies enhanced eicosanoid production in response to submaximal stimulation with the calcium ionophore A23187; and the levels of both prostaglandin (PG) E2 and LTB4 correlated with the number of lipid bodies. Furthermore, pretreatment of cells with actinomycin D or cycloheximide inhibited not only the induction of lipid body formation by PAF, but also the PAF-induced "priming" for enhanced PGE2 and LTB4 in PMN. Thus, the compartmentalization of lipids to form lipid bodies in PMN is dependent on specific cellular responses that can be PAF receptor mediated, involves signaling through 5-LO to form 5-HETE and then through PKC and PLC, and requires new protein synthesis. Since increases in lipid body numbers correlated with priming for enhanced PGE2 and LTB4 production in PMN, the induction of lipid bodies may have a role in the formation of eicosanoid mediators by leukocytes involved in inflammation.
Assuntos
Araquidonato 5-Lipoxigenase/metabolismo , Compartimento Celular , Corpos de Inclusão/efeitos dos fármacos , Metabolismo dos Lipídeos , Neutrófilos/efeitos dos fármacos , Fator de Ativação de Plaquetas/farmacologia , Receptores de Superfície Celular , Receptores Acoplados a Proteínas G , Animais , Araquidonato 5-Lipoxigenase/genética , Relação Dose-Resposta a Droga , Proteínas de Ligação ao GTP/metabolismo , Humanos , Inibidores de Lipoxigenase/farmacologia , Camundongos , Camundongos Knockout , Neutrófilos/enzimologia , Glicoproteínas da Membrana de Plaquetas/metabolismo , Proteína Quinase C/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Transdução de Sinais , Fosfolipases Tipo C/metabolismoRESUMO
The specific intracellular sites at which enzymes act to generate arachidonate-derived eicosanoid mediators of inflammation are uncertain. We evaluated the formation and function of cytoplasmic lipid bodies. Lipid body formation in eosinophils was a rapidly (<1 h) inducible response which was platelet-activating factor (PAF) receptor-mediated, involved signaling through protein kinase C, and required new protein synthesis. In intact and enucleated eosinophils, the PAF-induced increases in lipid body numbers correlated with enhanced production of both lipoxygenase- and cyclooxygenase-derived eicosanoids. All principal eosinophil eicosanoid-forming enzymes, 5-lipoxygenase, leukotriene C4 synthase, and cyclooxygenase, were immunolocalized to native as well as newly induced lipid bodies in intact and enucleated eosinophils. Thus, lipid bodies are structurally distinct, inducible, nonnuclear sites for enhanced synthesis of paracrine eicosanoid mediators of inflammation.
Assuntos
Eicosanoides/biossíntese , Eosinófilos/metabolismo , Eosinófilos/ultraestrutura , Corpos de Inclusão/metabolismo , Corpos de Inclusão/ultraestrutura , Metabolismo dos Lipídeos , Adulto , Araquidonato 5-Lipoxigenase/metabolismo , Sítios de Ligação , Núcleo Celular/metabolismo , Inibidores Enzimáticos/farmacologia , Eosinófilos/efeitos dos fármacos , Glutationa Transferase/metabolismo , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Corpos de Inclusão/efeitos dos fármacos , Mediadores da Inflamação/metabolismo , Fator de Ativação de Plaquetas/farmacologia , Prostaglandina-Endoperóxido Sintases/metabolismo , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Transdução de SinaisRESUMO
The present study compared the effects of early short-term with prolonged low-dose corticosteroid therapy in acute lung injury (ALI). In total, 120 BALB/c mice were randomly divided into five groups. In the control group, saline was intratracheally (i.t.) instilled. In the ALI group, mice received Escherichia coli lipopolysaccharide (10 microg i.t.). ALI animals were further randomised into four subgroups to receive saline (0.1 mL i.v.) or methylprednisolone (2 mg x kg(-1) i.v.) at 6 h, 24 h or daily (for 7 days, beginning at day 1). At 1, 3 and 8 weeks, in vivo and in vitro lung mechanics and histology (light and electron microscopy), collagen and elastic fibre content, cytokines in bronchoalveolar lavage fluid and the expression of matrix metalloproteinase (MMP)-9 and -2 were measured. In vivo (static elastance and viscoelastic pressure) and in vitro (tissue elastance and resistance) lung mechanics, alveolar collapse, cell infiltration, collagen and elastic fibre content and the expression of MMP-9 and MMP-2 were increased in ALI at 1 week. Methylprednisolone led to a complete resolution of lung mechanics, avoided fibroelastogenesis and the increase in the expression of MMP-9 and MMP-2 independent of steroid treatment design. Thus, early short-term, low-dose methylprednisolone is as effective as prolonged therapy in acute lung injury.
Assuntos
Lesão Pulmonar/tratamento farmacológico , Lesão Pulmonar/patologia , Metilprednisolona/administração & dosagem , Doença Aguda , Animais , Anti-Inflamatórios/administração & dosagem , Colágeno/química , Citocinas/metabolismo , Escherichia coli/metabolismo , Inflamação , Lipopolissacarídeos/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Fatores de TempoRESUMO
Transcription factors of the NFAT family are thought to play a major role in regulating the expression of cytokine genes and other inducible genes during the immune response. The role of NFAT1 was investigated by targeted disruption of the NFAT1 gene. Unexpectedly, cells from NFAT1 -/- mice showed increased primary responses to Leishmania major and mounted increased secondary responses to ovalbumin in vitro. In an in vivo model of allergic inflammation, the accumulation of eosinophils and levels of serum immunoglobulin E were increased in NFAT1 -/- mice. These results suggest that NFAT1 exerts a negative regulatory influence on the immune response.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Hipersensibilidade/imunologia , Imunidade , Ativação Linfocitária , Proteínas Nucleares , Fatores de Transcrição/fisiologia , Sequência de Aminoácidos , Animais , Antígenos de Protozoários/imunologia , Linhagem Celular , Citocinas/biossíntese , Proteínas de Ligação a DNA/genética , Eosinófilos/imunologia , Marcação de Genes , Imunoglobulina E/biossíntese , Memória Imunológica , Leishmania major/imunologia , Camundongos , Dados de Sequência Molecular , Fatores de Transcrição NFATC , Ovalbumina/imunologia , Linfócitos T/imunologia , Fatores de Transcrição/genéticaRESUMO
Programmed cell death by apoptosis of unnecessary or potentially harmful cells is clearly beneficial to multicellular organisms. Proper functioning of such a program demands that the removal of dying cells proceed without an inflammatory reaction. Phosphatidylserine (PS) is one of the ligands displayed by apoptotic cells that participates in their noninflammatory removal when recognized by neighboring phagocytes. PS ligation induces the release of transforming growth factor-beta (TGF-beta), an antiinflammatory cytokine that mediates the suppression of macrophage-mediated inflammation. In Hydra vulgaris, an organism that stands at the base of metazoan evolution, the selective advantage provided by apoptosis lies in the fact that Hydra can survive recycling apoptotic cells by phagocytosis. In unicellular organisms, it has been proposed that altruistic death benefits clonal populations of yeasts and trypanosomatids. Now we show that advantageous features of the apoptotic process can operate without death as the necessary outcome. Leishmania spp are able to evade the killing activity of phagocytes and establish themselves as obligate intracellular parasites. Amastigotes, responsible for disease propagation, similar to apoptotic cells, inhibit macrophage activity by exposing PS. Exposed PS participates in amastigote internalization. Recognition of this moiety by macrophages induces TGF-beta secretion and IL-10 synthesis, inhibits NO production, and increases susceptibility to intracellular leishmanial growth.
Assuntos
Apoptose , Regulação para Baixo/fisiologia , Hydra/fisiologia , Leishmania/fisiologia , Macrófagos/imunologia , Macrófagos/microbiologia , AnimaisRESUMO
BACKGROUND: The immunologic repercussions due to cavity insufflation are the focus of great discussion. The aim of this study was to compare the inflammatory response and bacterial dissemination after laparotomy and abdominal CO2 insufflation in a murine model of peritonitis. METHODS: Swiss mice were inoculated intraperitoneally with 0.5 ml of a solution containing 1 x 10(8) colony-forming units (CFU)/ml of Escherichia coli and were divided into three groups as follow: control (anesthesia for 30 min), laparotomy (2.5-cm midline incision for 30 min), and CO2 pneumoperitoneum (CO2 cavity insufflation for 30 min). The number of leukocytes, CFU/ml counting, and the levels of interleukin (IL)-6, tumor necrosis factor-alpha (TNF-alpha), and IL-10 were evaluated in blood, peritoneal, and pleural fluid samples obtained at 90 min and 18 h after the procedures. RESULTS: The laparotomy group showed a greater bacterial dissemination to the blood, peritoneum, and pleural cavity and also greater neutrophil migration to the peritoneal cavity compared to the CO2 insufflated and control groups. The 24-h mortality was also significantly higher in the laparotomy group. The IL-6 levels showed a precocious rise in all groups submitted to bacterial inoculation at the 90-min time point. At the 18-h time point, IL-6 levels in the peritoneum were significantly higher in the laparotomy group than in the control or CO2 insufflated groups. At the same time, TNF-alpha levels were higher in the laparotomy and CO2 insufflated groups than in controls; IL-10 levels showed no differences among the groups. CONCLUSIONS: Our results suggest that cavity insufflation with CO2 is a more effective method of access, inducing less bacterial dissemination and also a less intense inflammatory response. Cavity insufflation with CO2 may present a good option for the surgical treatment of patients with bacterial peritonitis.
Assuntos
Translocação Bacteriana , Dióxido de Carbono , Inflamação/etiologia , Insuflação/efeitos adversos , Laparotomia/efeitos adversos , Peritonite/cirurgia , Animais , Sangue/microbiologia , Contagem de Células Sanguíneas , Citocinas/sangue , Escherichia coli/fisiologia , Insuflação/normas , Laparotomia/mortalidade , Masculino , CamundongosRESUMO
Toll like receptors (TLRs) are involved in the modulation of diverse host genes expression through a complex network of signalling events that allow for an appropriate response to a microbial pathogen. In the present work we used TLR6KO mice in order to study the role of TLR6 in the immune discrimination of lipids from two Babesia bovis strains, attenuated R1A (LA) and virulent S2P (LV), and the consequent macrophage activation. We demonstrated that TLR6 is required for lipid body induction in murine peritoneal macrophages by both LA and LV. Interestingly, as regards IL-10 and COX-2/PGE2 pathway induction by LA and LV, we observed differences in the biological effects produced by these lipid extracts. Our results indicate a role of TLR6 in the down-modulation of these immunoregulators only in the case of LA, whereas this receptor was not implicated in pro-inflammatory TNFα, IL-6 and KC release induced by LA. Remarkably, LV did not exert the down-modulatory effect observed for LA, supporting the notion that LA and LV possess different lipid composition that could correlate with the polar pathogenic effect of both B. bovis strains.
Assuntos
Babesia bovis/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/metabolismo , Interleucina-10/metabolismo , Macrófagos Peritoneais/efeitos dos fármacos , Receptor 6 Toll-Like/metabolismo , Animais , Babesia bovis/patogenicidade , Ciclo-Oxigenase 2/genética , Dinoprostona/genética , Interleucina-10/genética , Gotículas Lipídicas/fisiologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Camundongos , Camundongos Knockout , Receptor 6 Toll-Like/genética , VirulênciaRESUMO
Although lipid bodies, inducible cytoplasmic inclusions active in arachidonic acid metabolism, are abundant in activated leukocytes, including eosinophils, mechanisms for eosinophil lipid body formation are not certain. Eosinophils from hypereosinophilic syndrome (HES) donors contained about twice (approximately 18/cell) as many lipid bodies as eosinophils froin normal donors (approximately 10/cell). By immunocytochemistry both 5- and 15-lipoxygenases were localized at lipid bodies in HES eosinophils. Platelet-activating factor (PAF) induced rapid, receptor-mediated increases in lipid bodies in normal and HES eosinophils. Protein kinase C (PKC) inhibitors, chelerythrine and calphostin C, inhibited PAF-induced lipid body formation partially in normal and HES eosinophils. In HES, but not normal, eosinophils, PAF-induced lipid body formation was completely blocked by two tyrosine kinase inhibitors, herbimycin A and genistein, which were not acting on 5-lipoxygenase because they also blocked 5-HETE-induced lipid body formation in HES, and not normal, eosinophils. After 24 h culture with eosinophil growth factor cytokines [interleukin (IL)-3, IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) or GM-CSF alone but not IL-5 or IL-3 alone], normal eosinophils were induced to exhibit an HES-like phenotype, including increased lipid body numbers and tyrosine kinase-dependent signaling for PAF-induced lipid body formation. Thus, signal transduction mechanisms involved in PAF-induced lipid body formation in eosinophils can be differentially recruited. Tyrosine kinase-dependent signaling is not involved in normal eosinophils, but is active in HES eosinophils and in normal eosinophils cultured with GM-CSF. PKC- and tyrosine kinase-dependent pathways are involved in the formation of eosinophil lipid bodies, which may facilitate enhanced synthesis of lipoxygenase-derived eicosanoids.
Assuntos
Eosinofilia/sangue , Eosinófilos/fisiologia , Corpos de Inclusão/ultraestrutura , Transdução de Sinais/fisiologia , Adulto , Araquidonato 15-Lipoxigenase/sangue , Araquidonato 5-Lipoxigenase/sangue , Benzoquinonas , Inibidores Enzimáticos/farmacologia , Eosinófilos/enzimologia , Eosinófilos/ultraestrutura , Genisteína/farmacologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Humanos , Técnicas In Vitro , Corpos de Inclusão/efeitos dos fármacos , Corpos de Inclusão/enzimologia , Interleucina-3/farmacologia , Interleucina-5/farmacologia , Lactamas Macrocíclicas , Lipídeos/sangue , Fosfoproteínas/sangue , Fosfotirosina , Fator de Ativação de Plaquetas/farmacologia , Proteínas Tirosina Quinases/antagonistas & inibidores , Proteínas Tirosina Quinases/sangue , Quinonas/farmacologia , Valores de Referência , Rifabutina/análogos & derivadosRESUMO
Selective platelet-activating factor (PAF) antagonists and autodesensitization to this lipid were used to investigate the role of PAF in antigen-induced pleurisy in the rat. Pleural inflammation was triggered by the intrathoracic (i.t.) injection of ovalbumin (12 micrograms/cavity) into animals actively sensitized 14 days before. Successive daily i.t. injections of PAF (1 microgram/cavity) led to selective autodesensitization, which was apparent after the third injection and maximal after the fifth. The PAF antagonists BN 52021 and WEB 2086 inhibited the late pleural eosinophil accumulation caused by antigen but, as also noted with WEB 2170, failed to modify the early antigen-induced plasma exudation and leukocyte infiltration. In contrast to the antagonists, desensitization to PAF was clearly effective against these early alterations. To further investigate this discrepancy, the antigenic challenge was performed 24 h after a single prestimulation with PAF, when sensitivity to the lipid was still intact. Under this condition, plasma exudation and cellular influx triggered by the antigen were also abrogated, indicating that this protective effect was accounted for by a mechanism other than refractoriness to PAF. Because 24 h after PAF injection only eosinophil counts remained elevated, an alternative eosinophilotactic substance was used to further study the mechanism of PAF versus antigen-induced pleural inflammation. Prior treatment with the peptide Ala-Gly-Ser-Glu (ECF-A, 20 micrograms/cavity) also inhibited the allergic pleurisy, whereas the noneosinophilotactic substances histamine (200 micrograms/cavity) and serotonin (100 micrograms/cavity) were inactive. Furthermore, drugs that share the ability to impair PAF-induced eosinophilia, including azelastine and cetirizine, prevented the inhibitory effect of PAF on the antigen-induced pleurisy. These findings suggest that PAF may account for the late eosinophilia, but not for the acute phase of the rat allergic pleurisy, which is clearly attenuated by PAF or ECF-A pretreatment.
Assuntos
Diterpenos , Hipersensibilidade a Drogas , Eosinófilos/efeitos dos fármacos , Ovalbumina/imunologia , Fator de Ativação de Plaquetas/fisiologia , Glicoproteínas da Membrana de Plaquetas , Pleurisia/imunologia , Receptores de Superfície Celular/fisiologia , Receptores Acoplados a Proteínas G , Análise de Variância , Animais , Azepinas/farmacologia , Fatores Quimiotáticos de Eosinófilos/farmacologia , Feminino , Adjuvante de Freund , Ginkgolídeos , Inflamação , Lactonas/farmacologia , Contagem de Leucócitos/efeitos dos fármacos , Masculino , Oligopeptídeos/farmacologia , Fator de Ativação de Plaquetas/antagonistas & inibidores , Fator de Ativação de Plaquetas/farmacologia , Pleurisia/sangue , Ratos , Ratos Wistar , Receptores de Superfície Celular/antagonistas & inibidores , Triazóis/farmacologiaRESUMO
Blood leukocyte count alterations induced by PAF-acether in anesthetized and nonanesthetized rats were investigated. Intravenous injection of increasing amounts of PAF-acether (1.5-8 micrograms/kg) in nonanesthetized animals induced dose-dependent hemoconcentration and leukocytosis. The former was apparent within 10 min, peaked from 30 min to 1 h, and diminished thereafter. The leukocytosis was noted within 30 min, was maximal at 1 h, and was over 4 h after injection of PAF-acether (4 micrograms/kg). It was characterized by a marked increase in the blood neutrophil counts under conditions in which the number of lymphocytes, monocytes, and eosinophils remained unchanged. PAF-acether-induced leukocytosis occurred in parallel with a marked decrease in the number of bone marrow nucleated cells, suggesting that the latter phenomenon may determine the former one. Leukocytosis by PAF-acether was inhibited dose-dependently by specific PAF-acether antagonists including BN 52021 (median effective dose ED50 = 4.99 mg/kg), WEB 2086 (ED50 = 4.59 mg/kg), and 48740 RP (ED50 = 9.02 mg/kg). General anesthesia by either pentobarbital, urethane, or ether inhalation, but not by ketamine, also impaired the PAF-acether-induced blood leukocytosis under conditions in which the hemoconcentration was not modified. In addition, pentobarbital-anesthetized rats did not have reduced bone marrow nucleated cell counts after PAF-acether stimulation. These findings are consistent with the assessment that PAF-acether-induced rat leukocytosis is accounted for by a bone marrow neutrophil mobilization process that is clearly suppressed in animals anesthetized by pentobarbital.
Assuntos
Anestésicos/farmacologia , Contagem de Leucócitos/efeitos dos fármacos , Fator de Ativação de Plaquetas/farmacologia , Anestesia Geral , Animais , Células da Medula Óssea , Hematócrito , Leucocitose/induzido quimicamente , Masculino , Ratos , Ratos WistarRESUMO
Tumor necrosis factor-stimulated gene 14 (TSG-14)/PTX3 was identified originally as a TNF-alpha and IL-1beta-stimulated gene from normal, human foreskin fibroblasts and vascular endothelial cells, respectively. TSG-14 gene encodes a 42-kDa-secreted glycoprotein with a carboxy-terminal half that shares homology with the entire sequence of C-reactive protein (CRP) and serum amyloid P component (SAP), acute-phase proteins of the pentraxin family. Some experimental evidence suggests that TSG-14 plays a role in inflammation, yet its function and mechanism of action remain unclear. We have generated transgenic mice that overexpress the murine TSG-14 gene under the control of its own promoter. From eight transgenic founders, two lineages were derived and better characterized: Tg2 and Tg4, carrying two and four copies of the transgene, respectively. TSG-14 transgenic mice were found to be more resistant to the endotoxic shock induced by LPS and to the polymicrobial sepsis caused by cecal ligation and puncture (CLP). Moreover, macrophages derived from the transgenic mice produced higher amounts of nitric oxide in response to IFN-gamma, TNF-alpha, and LPS as compared with macrophages from wild-type animals, and the augmented response appears to be the consequence of a higher responsiveness of transgenic macrophages to IFN-gamma. The data shown here are the first in vivo evidence of the involvement of TSG-14 in the inflammatory process and suggest a role for TSG-14 in the defense against bacterial infections.
Assuntos
Proteína C-Reativa/fisiologia , Endotoxemia/genética , Sepse/genética , Componente Amiloide P Sérico/fisiologia , Animais , Animais não Endogâmicos , Proteína C-Reativa/genética , Ceco/lesões , Ceco/microbiologia , Modelos Animais de Doenças , Endotoxemia/imunologia , Humanos , Imunidade Inata , Inflamação , Interferon gama/farmacologia , Perfuração Intestinal/complicações , Ligadura , Lipopolissacarídeos/toxicidade , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/fisiologia , Camundongos , Camundongos Transgênicos , Óxido Nítrico/biossíntese , Proteínas Recombinantes de Fusão/fisiologia , Sepse/etiologia , Sepse/imunologia , Componente Amiloide P Sérico/genética , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
In this study we investigated the involvement of inflammatory cells in the pleural accumulation of eosinophils induced by lipopolysaccharide (LPS). Intrathoracic (i.t.) injection of LPS (250 ng/cavity) into rats induced a significant eosinophil accumulation that developed within 24 h, was maximal at 48 h, and returned to control values within 120 h. This eosinophil influx was preceded by a huge neutrophil influx within 4 h and accompanied by a mononuclear cell accumulation between 24 and 48 h. Pretreatment with an antineutrophil monoclonal antibody (RP-3, 2 ml per animal) selectively reduced the number of circulating neutrophils within 8 h but failed to alter the LPS-induced eosinophilia. Similarly, platelet depletion with an anti-rat platelet antiserum did not alter the LPS-induced eosinophil accumulation. Cyclosporine (50 mg/kg, 12 and 2 h before) partially inhibited (51%) the LPS-induced pleural eosinophilia, whereas the eosinophilia was not changed by prior degranulation of pleural mast cells with polymyxin B (10 micrograms/cavity, 24 h before). Moreover, selective depletion of T lymphocytes using an anti-Thy 1.0 monoclonal antibody significantly inhibited the eosinophilia triggered by LPS. The i.t. injection of liposomes containing dichloromethylene diphosphonate significantly reduced (65%) the number of resident macrophages after 5 days. Under this condition, the eosinophil infiltration induced by LPS was completely inhibited. Accordingly, the i.t. injection of supernatant from macrophage monolayers, obtained from the pleural cavities of LPS-injected rats, into naive recipient animals led to a twofold increase in the number of pleural eosinophils. In conclusion, our data suggest an important role for resident macrophages and T lymphocytes in the eosinophil accumulation induced by LPS.
Assuntos
Eosinófilos/citologia , Eosinófilos/efeitos dos fármacos , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/fisiologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/fisiologia , Animais , Degranulação Celular/fisiologia , Fatores Quimiotáticos de Eosinófilos/biossíntese , Quimiotaxia de Leucócito/efeitos dos fármacos , Quimiotaxia de Leucócito/fisiologia , Ciclosporina/farmacologia , Eosinófilos/fisiologia , Injeções Espinhais , Macrófagos/metabolismo , Masculino , Mastócitos/fisiologia , Camundongos , Eosinofilia Pulmonar/induzido quimicamente , Ratos , Ratos Wistar , Estimulação QuímicaRESUMO
The role of catecholamines in regulating pleural neutrophilia evoked by intrathoracic (i.t.) injection of lipopolysaccharide (LPS) was investigated in Wistar rats by means of surgical adrenalectomy, depletion of catecholamine stores or adrenoceptor blockade. Treatment of animals with a single dose of LPS evoked a dramatic increase in the number of pleural neutrophils concomitant with an increase in the number of these cells in blood at 4 h. Although blood neutrophilia was drastically reduced when catecholamine stores were depleted with intraperitoneal (i.p.) injection of reserpine, pleural neutrophilia was not modified. However, the i.t. injection of reserpine reduced the increase in pleural neutrophils after LPS stimulation. Adrenalectomy failed to inhibit the increase in neutrophil counts in the blood or pleural cavity after LPS challenge. Pretreatment with intravenous (i.v.) injection of prazosin, an alpha(1)/alpha(2B) antagonist, reduced LPS-induced blood but not pleural neutrophilia. On the other hand, although pleural neutrophilia was not affected by systemic pretreatment with the alpha(2)-adrenoceptor antagonist, yohimbine, the local treatment (i. t. injection) with this antagonist markedly reduced the increase in pleural neutrophil counts observed after stimulation by LPS. In contrast, pleural neutrophilia induced by i.t injection of formyl-methionyl-leucyl-phenylalanine (fMLP) was not modified by local treatment with yohimbine. Taken together, our results suggest that catecholamines, through activation of alpha(1) and alpha(2)-adrenoceptors, play a role in the regulation of blood and pleural neutrophilia observed during the inflammatory response evoked by LPS in the pleural cavity.
Assuntos
Lipopolissacarídeos/farmacologia , Neutrófilos/imunologia , Pleura/imunologia , Receptores Adrenérgicos/imunologia , Adrenalectomia , Inibidores da Captação Adrenérgica/farmacologia , Antagonistas Adrenérgicos alfa/farmacologia , Animais , Catecolaminas/metabolismo , Quimiotaxia de Leucócito/efeitos dos fármacos , Quimiotaxia de Leucócito/imunologia , Masculino , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/citologia , Pleura/química , Pleura/citologia , Pleura/efeitos dos fármacos , Pleurisia/induzido quimicamente , Pleurisia/imunologia , Ratos , Ratos Wistar , Receptores Adrenérgicos/metabolismo , Reserpina/farmacologia , Ioimbina/farmacologiaRESUMO
The injection of PAF (6 micrograms/kg, i.v.) induced, in rats, haemoconcentration accompanied by an increase in the platelet number, as attested by the counts of platelets in blood samples diluted in formalin-free EDTA solution. This increase was significant at 15 min, peaked from 1 to 4 h and returned to basal levels 24 h after the lipid administration. The release of platelets induced by PAF was inhibited dose-dependently by specific PAF receptor antagonist such as WEB 2086 (0.5-2 mg/kg), BN 52021 and 48740 RP (5-25 mg/kg). Furthermore, platelet mobilization was clearly impaired in splenectomized animals stimulated by PAF, whereas thrombocytopenia and haemoconcentration by the same stimulus were intact. It was also noted that a second injection of PAF, 24 h after the initial stimulation with the lipid, failed to induce an increase in platelet counts, indicating autodesensitization. Desensitization to PAF or pretreatment with PAF antagonists clearly prevented the increase in the platelet counts after stimulation by adrenaline (15 micrograms/kg). These findings suggest that, in rats, PAF can induce release of platelets by a spleen-dependent mechanism and that this lipid may be relevant to the thrombocytosis triggered by adrenaline.
Assuntos
Epinefrina/farmacologia , Fator de Ativação de Plaquetas/farmacologia , Agregação Plaquetária/fisiologia , Trombocitose/sangue , Animais , Hematócrito , Masculino , Fator de Ativação de Plaquetas/antagonistas & inibidores , Contagem de Plaquetas/efeitos dos fármacos , Ratos , Ratos Endogâmicos , Baço/fisiologia , Esplenectomia , Trombocitose/induzido quimicamenteRESUMO
1. The intrapleural injection of Paf-acether into rats caused, at 30 min, a marked exudation accompanied by a reduction in the pleural leucocyte count. At 6 h, the exudate volume had decreased and a significant increase in the total leucocyte count, particularly eosinophils was noted. 2. Two Paf-acether antagonists, WEB 2086 and 48740 RP abrogated the pleural leucopenia observed 30 min after Paf-acether administration, whereas the exudation was inhibited only by the former. Pleurisy was also reduced by about 60% with dexamethasone, by about 45% with BW 755C or LY 171883, a mixed cyclo-oxygenase/lipoxygenase inhibitor and a peptido-leukotriene antagonist respectively, and by about 30% with indomethacin, flurbiprofen or piroxicam. 3. Repeated daily intrapleural injections of Paf-acether led to a state of progressive desensitization to Paf-acether itself, whereas responsiveness to 5-hydroxytryptamine was maintained. In addition, the Paf-induced auto-desensitization was largely inhibited by WEB 2086. 4. Pleurisy induced by zymosan, but not by carrageenin, was significantly reduced in Paf-acether-desensitized animals. These results were consistent with those obtained with WEB 2086 which suppressed zymosan-induced but not carrageenin-induced pleurisy. 5. This study suggests that Paf-acether-induced pleurisy in the rat may be mediated by lipoxygenase arachidonic acid metabolites and that pleurisy induced by zymosan, but not by carrageenin, is largely dependent upon Paf-acether.
Assuntos
Azepinas/farmacologia , Fator de Ativação de Plaquetas/antagonistas & inibidores , Pleurisia/prevenção & controle , Piridinas/farmacologia , Tiazóis/farmacologia , Triazinas/farmacologia , Triazóis , Zimosan/antagonistas & inibidores , Animais , Carragenina/antagonistas & inibidores , Carragenina/toxicidade , Contagem de Leucócitos , Masculino , Pleurisia/induzido quimicamente , Ratos , Ratos Endogâmicos , Zimosan/toxicidadeRESUMO
1 The intrathoracic injection of platelet activating factor (PAF) into rats induced a decrease in the pleural leucocyte numbers within 15 min, accompanied by a marked exudation, maximal 1 h later. After 6 h, concomitantly with the reduction of exudation, a marked increase in the number of mononuclear cells, neutrophils and eosinophils was observed. Within 24 h, the pleural eosinophil accumulation peaked and persisted up to 96 h. 2 Topical treatment with nedocromil sodium affected pleural exudation by PAF under conditions where systemic meclizine was ineffective. Nedocromil sodium blocked, dose-dependently, the increase in the pleural content of mononuclear cells, neutrophils and eosinophils, observed 6 h after PAF administration, as well as the eosinophilia 24 h later. Moreover, the co-incubation of peritoneal eosinophils with nedocromil sodium did not interfere with the migration triggered by PAF. 3 The transfer of the 6 h-PAF pleural washings from donor to recipient rats caused a selective pleural eosinophilia, which was clearly inhibited when nedocromil sodium was administered to donor, but not to recipient animals, showing that this drug interferes with the generation rather than with the expression of the eosinophilotactic activity(ies). 4 These findings indicate that the nedocromil sodium interferes with PAF-induced exudation and leucocyte accumulation, by a mechanism other than its ability to reduce the local effects of histamine and which may relate to suppression of the eosinophilotactic principle generation.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Linfocinas/biossíntese , Fator de Ativação de Plaquetas/farmacologia , Quinolonas/farmacologia , Animais , Quimiotaxia de Leucócito/efeitos dos fármacos , Eosinófilos/efeitos dos fármacos , Feminino , Histamina/farmacologia , Contagem de Leucócitos , Masculino , Nedocromil , Fator de Ativação de Plaquetas/antagonistas & inibidores , Pleurisia/induzido quimicamente , Pleurisia/metabolismo , Proteínas/metabolismo , Ratos , Ratos EndogâmicosRESUMO
1. The role of both exogenously administered and endogenously generated bradykinin (BK) on LPS-induced eosinophil accumulation in the mice pleural cavity was investigated by means of treatment with BK selective receptor agonists/antagonists and captopril. 2. Intrathoracic (i.t.) injection of LPS (250 ng cavity(-1)) induced eosinophil influx at 24 h as previously described (Bozza et al., 1993). Pretreatment with the B1 receptor antagonist des-Arg9-[leu-8]BK (0.025 and 0.25 nmol cavity(-1)) showed no effect on this phenomenon, whereas pretreatment with the B2 receptor antagonists, NPC 17731 (0.025 and 0.25 nmol cavity(-1)) or HOE 140 (2.5 nmol cavity(-1)), increased LPS-induced eosinophil influx. Accordingly, pretreatment with captopril at 10 mg kg(-1) i.p., inhibited eosinophil infiltration induced by LPS in the pleural cavity, suggesting that endogenous BK is down-regulating LPS-induced eosinophil accumulation. 3. BK administered at 15 and 25 nmol cavity(-1), i.t. or i.p. also inhibited LPS-induced eosinophil accumulation. BK alone had no effect on the basal number of leucocytes in the pleural or peritoneal cavity in doses up to 25 nmol cavity(-1). Nevertheless, when injected at doses of 50 and 100 nmol cavity(-1) BK induced leucocyte influx characterized by neutrophil and eosinophil accumulation at 24 h. 4. Similarly to what was observed with BK, a specific B2 receptor agonist, Tyr8BK, administered at 0.25 nmol cavity(-1) i.p., significantly inhibited the eosinophil influx induced by LPS. 5. The mechanism by which B2 receptor agonists inhibit LPS-induced eosinophil accumulation was investigated by pretreating the animals with indomethacin or a selective cyclooxygenase-2 inhibitor, NS-398. Pretreatment with either indomethacin or NS-398 had no effect on eosinophil influx induced by LPS alone, but those drugs were able to restore the LPS-induced eosinophil influx in Tyr8BK (0.25 nmol cavity(-1)) injected mice. 6. In conclusion, endogenously generated bradykinin seems to modulate, through activation of B2 receptors, eosinphil accumulation induced by LPS via a mechanism dependent on prostanoid synthesis.