RESUMO
Chronic use of statins may have anti-inflammatory action, promoting immunomodulation and survival in patients with sepsis. This study aimed to analyze the effects of pretreatment with simvastatin in lethal sepsis induced by cecal ligation and puncture (CLP). Male Swiss mice received prophylactic treatment with simvastatin or pyrogen-free water orally in a single daily dose for 30 days. After this period, the CLP was performed. Naïve and Sham groups were performed as non-infected controls. Animal survival was monitored for 60 h after the CLP. Half of mice were euthanized after 12 h to analyze colony-forming units (CFUs); hematological parameters; production of IL-10, IL-12, IL-6, TNF-α, IFN-γ, and MCP-1; cell counts on peritoneum, bronchoalveolar lavage (BAL), bone marrow, spleen, and mesenteric lymph node; immunephenotyping of T cells and antigen presenting cells and production of hydrogen peroxide (H2O2). Simvastatin induced an increase in survival and a decrease in the CFU count on peritoneum and on BAL cells number, especially lymphocytes. There was an increase in the platelets and lymphocytes number in the Simvastatin group when compared to the CLP group. Simvastatin induced a greater activation and proliferation of CD4+ T cells, as well as an increase in IL-6 and MCP-1 production, in chemotaxis to the peritoneum and in H2O2 secretion at this site. These data suggest that simvastatin has an impact on the survival of animals, as well as immunomodulatory effects in sepsis induced by CLP in mice.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Sepse , Sinvastatina/farmacologia , Animais , Linfócitos T CD4-Positivos/patologia , Modelos Animais de Doenças , Peróxido de Hidrogênio/imunologia , Masculino , Camundongos , Sepse/imunologia , Sepse/patologia , Sepse/prevenção & controleRESUMO
Alveolar macrophages (AMs) are multitasking cells that maintain lung homeostasis by clearing apoptotic cells (efferocytosis) and performing antimicrobial effector functions. Different PRRs have been described to be involved in the binding and capture of non-opsonized Streptococcus pneumoniae, such as TLR-2, mannose receptor (MR) and scavenger receptors (SRs). However, the mechanism by which the ingestion of apoptotic cells negatively influences the clearance of non-opsonized S. pneumoniae remains to be determined. In this study, we evaluated whether the prostaglandin E2 (PGE2) produced during efferocytosis by AMs inhibits the ingestion and killing of non-opsonized S. pneumoniae. Resident AMs were pre-treated with an E prostanoid (EP) receptor antagonist, inhibitors of cyclooxygenase and protein kinase A (PKA), incubated with apoptotic Jurkat T cells, and then challenged with S. pneumoniae. Efferocytosis slightly decreased the phagocytosis of S. pneumoniae but greatly inhibited bacterial killing by AMs in a manner dependent on PGE2 production, activation of the EP2-EP4/cAMP/PKA pathway and inhibition of H2O2 production. Our data suggest that the PGE2 produced by AMs during efferocytosis inhibits H2O2 production and impairs the efficient clearance non-opsonized S. pneumoniae by EP2-EP4/cAMP/PKA pathway.
Assuntos
Dinoprostona/metabolismo , Macrófagos Alveolares/imunologia , Fagocitose , Infecções Pneumocócicas/imunologia , Streptococcus pneumoniae/imunologia , Animais , Apoptose , Bacteriólise , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Feminino , Homeostase , Humanos , Peróxido de Hidrogênio/metabolismo , Células Jurkat , Macrófagos Alveolares/microbiologia , Ratos , Ratos Wistar , Receptores de Prostaglandina E Subtipo EP2/metabolismo , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Transdução de SinaisRESUMO
Platelet-activating factor receptor (PAFR) is a G protein-coupled receptor (GPCR) implicated in many diseases. Toll-like receptors (TLRs) play a critical role in shaping innate and adaptive immune responses. In this study, we investigated whether PAFR signaling changes the macrophages responsiveness to agonists of TLR2 (Pam3Cys), TLR4 (LPS), and TLR3 agonist Poly(I:C). Exogenous PAF inhibited the production of pro-inflammatory cytokines (IL-12p40, IL-6, and TNF-α) and increased anti-inflammatory IL-10 in macrophages challenged with Pam3Cys and LPS, but not with Poly (I:C). PAF did not affect mRNA expression of MyD88, suggesting that PAF acts downstream the adaptor. PAF inhibited LPS-induced phosphorylation of NF-κB p65 and increased NF-κB p105 phosphorylation, which is processed in the proteasome to generate p50 subunit. The PAF potentiation of IL-10 production was dependent on proteasome processing but independent of NF-κB transactivation domain. Inhibition of p50 abolished the PAF-induced IL-10 production. These findings indicate that the impaired transcriptional activity of the p65 subunit and the enhanced p105 phosphorylation induced by PAF are responsible for down regulation of pro-inflammatory cytokines and up regulation of IL-10, respectively, in LPS-challenged macrophages. Together, our data unveil a heretofore unrecognized role for PAFR in modulating activation of NF-κB in macrophages.
Assuntos
Inflamação/metabolismo , Subunidade p50 de NF-kappa B/metabolismo , Glicoproteínas da Membrana de Plaquetas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Receptores Toll-Like/metabolismo , Fator de Transcrição RelA/metabolismo , Animais , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Interleucina-10/metabolismo , Lipopolissacarídeos/farmacologia , Lipoproteínas/farmacologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Macrófagos Peritoneais/patologia , Masculino , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/genética , Fator 88 de Diferenciação Mieloide/metabolismo , Fator de Ativação de Plaquetas/metabolismo , Fator de Ativação de Plaquetas/farmacologia , Receptor 2 Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismoRESUMO
Tityus serrulatus sting causes thousands of deaths annually worldwide. T. serrulatus-envenomed victims exhibit local or systemic reaction that culminates in pulmonary oedema, potentially leading to death. However, the molecular mechanisms underlying T. serrulatus venom (TsV) activity remain unknown. Here we show that TsV triggers NLRP3 inflammasome activation via K(+) efflux. Mechanistically, TsV triggers lung-resident cells to release PGE2, which induces IL-1ß production via E prostanoid receptor 2/4-cAMP-PKA-NFκB-dependent mechanisms. IL-1ß/IL-1R actions account for oedema and neutrophil recruitment to the lungs, leading to TsV-induced mortality. Inflammasome activation triggers LTB4 production and further PGE2 via IL-1ß/IL-1R signalling. Activation of LTB4-BLT1/2 pathway decreases cAMP generation, controlling TsV-induced inflammation. Exogenous administration confirms LTB4 anti-inflammatory activity and abrogates TsV-induced mortality. These results suggest that the balance between LTB4 and PGE2 determines the amount of IL-1ß inflammasome-dependent release and the outcome of envenomation. We suggest COX1/2 inhibition as an effective therapeutic intervention for scorpion envenomation.
Assuntos
Proteínas de Transporte/genética , Dinoprostona/farmacologia , Interleucina-1beta/efeitos dos fármacos , Leucotrieno B4/farmacologia , Macrófagos Peritoneais/efeitos dos fármacos , Picadas de Escorpião/imunologia , Venenos de Escorpião/farmacologia , Animais , Araquidonato 5-Lipoxigenase/genética , Western Blotting , Proteínas de Transporte/imunologia , Celecoxib/farmacologia , AMP Cíclico/imunologia , Proteínas Quinases Dependentes de AMP Cíclico/efeitos dos fármacos , Proteínas Quinases Dependentes de AMP Cíclico/imunologia , Inibidores de Ciclo-Oxigenase/farmacologia , Dinoprostona/imunologia , Técnicas In Vitro , Indóis/farmacologia , Indometacina/farmacologia , Inflamassomos/imunologia , Interleucina-1beta/imunologia , Leucotrieno B4/imunologia , Inibidores de Lipoxigenase/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos Peritoneais/imunologia , Camundongos , Camundongos Knockout , NF-kappa B/efeitos dos fármacos , NF-kappa B/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR , Fosfoproteínas , Antagonistas de Prostaglandina/farmacologia , Receptores de Prostaglandina E Subtipo EP2/efeitos dos fármacos , Receptores de Prostaglandina E Subtipo EP2/imunologia , Receptores de Prostaglandina E Subtipo EP4/efeitos dos fármacos , Receptores de Prostaglandina E Subtipo EP4/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Picadas de Escorpião/mortalidade , Escorpiões , Xantonas/farmacologiaRESUMO
Although Leishmania (Viannia) braziliensis is the most prevalent species that cause American tegumentary leishmaniasis (ATL), the immune response against this parasite has been poorly investigated. Upon activation, macrophages produce a series of pro-inflammatory molecules, including the lipid mediator leukotriene B4 (LTB4). LTB4 has been shown to enhance several macrophage functions, but its role in human macrophages is less known. Here, we investigated the role of LTB4 on human monocyte-derived macrophages infected with human isolate of L. (V.) braziliensis (IMG3). It was found that human macrophages produce LTB4 upon infection with Leishmania, which by autocrine or paracrine activation of its high affinity receptor BLT1, potentiates macrophage leishmanicidal activity. This LTB4 effect is mediated by increased secretion of reactive oxygen species (ROS). Moreover, Leishmania infection decreased the expression of BLT1, leading to the speculation that this could represent a parasite escape mechanism to establish a chronic inflammatory infection. Therefore, our data suggest that LTB4 could be used in therapeutic strategies to control Leishmania infection.
Assuntos
Leishmania braziliensis/imunologia , Leishmania braziliensis/fisiologia , Leucotrieno B4/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Receptores do Leucotrieno B4/metabolismo , Sobrevivência Celular , Células Cultivadas , Interações Hospedeiro-Patógeno , Humanos , Macrófagos/parasitologia , Espécies Reativas de Oxigênio/metabolismoRESUMO
Clinical and experimental observations have supported the notion that free heme released during hemorrhagic and hemolytic episodes may have a major role in lung inflammation. With alveolar macrophages (AM) being the main line of defense in lung environments, the influence of free heme on AM activity and function was investigated. We observed that heme in a concentration range found during hemolytic episodes (3-30 µM) elicits AM to present a proinflammatory profile, stimulating reactive oxygen species (ROS) and nitric oxide (NO) generation and inducing IL-1ß, IL-6, and IL-10 secretion. ROS production is NADPH oxidase-dependent, being inhibited by DPI and apocynin, and involves p47 subunit phosphorylation. Furthermore, heme induces NF- κB nuclear translocation, iNOS, and also HO-1 expression. Moreover, AM stimulated with free heme show enhanced phagocytic and bactericidal activities. Taken together, the data support a dual role for heme in the inflammatory response associated with lung hemorrhage, acting as a proinflammatory molecule that can either act as both an adjuvant of the innate immunity and as an amplifier of the inflammatory response, leading tissue injury. The understanding of heme effects on pulmonary inflammatory processes can lead to the development of new strategies to ameliorate tissue damage associated with hemorrhagic episodes.
Assuntos
Heme/metabolismo , Inflamação/metabolismo , Macrófagos Alveolares/metabolismo , Síndrome Metabólica/imunologia , Pneumonia/metabolismo , Animais , Humanos , Camundongos , RatosRESUMO
Acute lung injury (ALI) develops in response to a direct insult to the lung or secondarily to a systemic inflammatory response, such as sepsis. There is clinical evidence that the incidence and severity of ALI induced by direct insult are lower in diabetics. In the present study we investigated whether the same occurs in ALI secondarily to sepsis and the molecular mechanisms involved. Diabetes was induced in male Wistar rats by alloxan and sepsis by caecal ligation and puncture surgery (CLP). Six hours later, the lungs were examined for oedema and cell infiltration in bronchoalveolar lavage. Alveolar macrophages (AMs) were cultured in vitro for analysis of IκB and p65 subunit of NFκB phosphorylation and MyD88 and SOCS-1 mRNA. Diabetic rats were more susceptible to sepsis than non-diabetics. In non-diabetic rats, the lung presented oedema, leukocyte infiltration and increased COX2 expression. In diabetic rats these inflammatory events were significantly less intense. To understand why diabetic rats despite being more susceptible to sepsis develop milder ALI, we examined the NFκB activation in AMs of animals with sepsis. Whereas in non-diabetic rats the phosphorylation of IκB and p65 subunit occurred after 6 h of sepsis induction, this did not occur in diabetics. Moreover, in AMs from diabetic rats the expression of MyD88 mRNA was lower and that of SOCS-1 mRNA was increased compared with AMs from non-diabetic rats. These results show that ALI secondary to sepsis is milder in diabetic rats and this correlates with impaired activation of NFκB, increased SOCS-1 and decreased MyD88 mRNA.
Assuntos
Lesão Pulmonar Aguda/etiologia , Lesão Pulmonar Aguda/metabolismo , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 1/metabolismo , NF-kappa B/metabolismo , Sepse/complicações , Lesão Pulmonar Aguda/patologia , Animais , Ciclo-Oxigenase 2/metabolismo , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/mortalidade , Diabetes Mellitus Tipo 1/mortalidade , Suscetibilidade a Doenças , Ativação Enzimática , Macrófagos Alveolares/metabolismo , Masculino , Ratos , Ratos Wistar , Fatores de TempoRESUMO
Candida albicans is the most common opportunistic fungal pathogen and causes local and systemic disease in immunocompromised patients. Alveolar macrophages (AMs) are pivotal for the clearance of C. albicans from the lung. Activated AMs secrete 5-lipoxygenase-derived leukotrienes (LTs), which in turn enhance phagocytosis and microbicidal activity against a diverse array of pathogens. Our aim was to investigate the role of LTB(4) and LTD(4) in AM antimicrobial functions against C. albicans and the signaling pathways involved. Pharmacologic and genetic inhibition of LT biosynthesis as well as receptor antagonism reduced phagocytosis of C. albicans when compared with untreated or WT controls. Conversely, exogenous LTs of both classes augmented base-line C. albicans phagocytosis by AMs. Although LTB(4) enhanced mainly mannose receptor-dependent fungal ingestion, LTD(4) enhanced mainly dectin-1 receptor-mediated phagocytosis. LT enhancement of yeast ingestion was dependent on protein kinase C-δ (PKCδ) and PI3K but not PKCα and MAPK activation. Both LTs reduced activation of cofilin-1, whereas they enhanced total cellular F-actin; however, LTB(4) accomplished this through the activation of LIM kinases (LIMKs) 1 and 2, whereas LTD(4) did so exclusively via LIMK-2. Finally, both exogenous LTB(4) and LTD(4) enhanced AM fungicidal activity in an NADPH oxidase-dependent manner. Our data identify LTB(4) and LTD(4) as key mediators of innate immunity against C. albicans, which act by both distinct and conserved signaling mechanisms to enhance multiple antimicrobial functions of AMs.
Assuntos
Actinas/metabolismo , Candida albicans/metabolismo , Candidíase/metabolismo , Cofilina 1/metabolismo , Imunidade Inata/fisiologia , Leucotrieno B4/metabolismo , Macrófagos Alveolares/metabolismo , Actinas/genética , Actinas/imunologia , Animais , Candida albicans/imunologia , Candidíase/genética , Candidíase/imunologia , Cofilina 1/genética , Cofilina 1/imunologia , Ativação Enzimática/genética , Ativação Enzimática/imunologia , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/imunologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Lectinas Tipo C , Leucotrieno B4/genética , Leucotrieno B4/imunologia , Quinases Lim/genética , Quinases Lim/imunologia , Quinases Lim/metabolismo , Macrófagos Alveolares/imunologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/imunologia , Proteínas do Tecido Nervoso/metabolismo , Fagocitose/genética , Fagocitose/imunologia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/imunologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteína Quinase C-delta , Ratos , Ratos WistarRESUMO
Leukotrienes (LTs) are potent lipid mediators involved in the control of host defense. LTB(4) induces leukocyte accumulation, enhances phagocytosis and bacterial clearance, and increases NO synthesis. LTB(4) is also important in early effector T cell recruitment that is mediated by LTB(4) receptor 1, the high-affinity receptor for LTB(4). The aims of this study were to evaluate whether LTs are involved in the secondary immune response to vaccination in a murine model of Histoplasma capsulatum infection. Our results demonstrate that protection of wild-type mice immunized with cell-free Ags from H. capsulatum against histoplasmosis was associated with increased LTB(4) and IFN-gamma production as well as recruitment of memory T cells into the lungs. In contrast, cell-free Ag-immunized mice lacking 5-lipoxygenase(-/-), a critical enzyme involved in LT synthesis, displayed a marked decrease on recruitment of memory T cells to the lungs associated with increased synthesis of TGF-beta as well as IL-10. Strikingly, these effects were associated with increased mortality to 5-lipoxygenase(-/-)-infected mice. These data establish an important immunomodulatory role of LTs, in both the primary and secondary immune responses to histoplasmosis.
Assuntos
Adjuvantes Imunológicos/fisiologia , Histoplasmose/imunologia , Imunização Secundária , Memória Imunológica , Leucotrieno B4/fisiologia , Subpopulações de Linfócitos T/imunologia , Adjuvantes Imunológicos/antagonistas & inibidores , Adjuvantes Imunológicos/biossíntese , Animais , Antígenos de Fungos/administração & dosagem , Antígenos de Fungos/imunologia , Movimento Celular/imunologia , Citocinas/biossíntese , Vacinas Fúngicas/administração & dosagem , Vacinas Fúngicas/imunologia , Histoplasma/imunologia , Histoplasmose/microbiologia , Histoplasmose/prevenção & controle , Humanos , Imunidade Inata , Leucotrieno B4/antagonistas & inibidores , Leucotrieno B4/biossíntese , Pneumopatias Fúngicas/imunologia , Pneumopatias Fúngicas/patologia , Pneumopatias Fúngicas/prevenção & controle , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/microbiologiaRESUMO
Leukotrienes (LTs) are known to be produced by macrophages when challenged with Leishmania, but it is not known whether these lipid mediators play a role in host defense against this important protozoan parasite. In this study, we investigated the involvement of LTs in the in vitro and in vivo response to Leishmania amazonensis infection in susceptible (BALB/c) and resistant (C3H/HePAS) mice. Pharmacologic or genetic deficiency of LTs resulted in impaired leishmanicidal activity of peritoneal macrophages in vitro. In contrast, addition of LTB4 increased leishmanicidal activity and this effect was dependent on the BLT1 receptor. LTB4 augmented NO production in response to L. amazonensis challenge, and studies with a NO synthesis inhibitor revealed that NO was critical for the enhancement of macrophage leishmanicidal activity. Interestingly, macrophages from resistant mice produced higher levels of LTB4 upon L. amazonensis challenge than did those from susceptible mice. In vivo infection severity, as assessed by footpad swelling following s.c. promastigote inoculation, was increased when endogenous LT synthesis was abrogated either pharmacologically or genetically. Taken together, these results for the first time reveal an important role for LTB4 in the protective response to L. amazonensis, identify relevant leishmanicidal mechanisms, and suggest that genetic variation in LTB4 synthesis might influence resistance and susceptibility patterns to infection.