RESUMEN
The gut microbiome orchestrates epithelial homeostasis and both local and remote immunological responses. Critical to these regulatory interactions are innate immune receptors termed Toll-like receptors (TLRs). Studies to date have implicated innate immunity and Toll-like receptors in shaping key features of the gut microbiome. However, a variety of biological and environmental variables are also implicated in determining gut microbiota composition. In this report, we hypothesized that cohousing and environment dominated the regulation of the gut microbiota in animal models independent of innate immunity. To determine the importance of these variables, innate immunity, or environment in shaping gut microbiota, we used a randomized cohousing strategy and transgenic TLR-deficient mice. We have found that mice cohoused together by genotype exhibited limited changes over time in the composition of the gut microbiota. However, for mice randomized to cage, we report extensive changes in the gut microbiota, independent of TLR function, whereby the fecal microbiota of TLR-deficient mice converges with that of wild-type mice. TLR5-deficient mice in these experiments exhibit greater susceptibility to comparative changes in the microbiota than other TLR-deficient mice and wild-type mice. Our work has broad implications for the study of innate immunity and host-microbiota interactions. Given the profound impact that gut dysbiosis may have on immunity, this report highlights the potential impact of cohousing on the gut microbiota and indices of inflammation as outcomes in biological models of infectious or inflammatory disease.
Asunto(s)
Microbioma Gastrointestinal , Homeostasis , Interacciones Microbiota-Huesped , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiología , Receptores Toll-Like/metabolismo , Animales , Susceptibilidad a Enfermedades , Disbiosis , Inmunidad Innata , Inmunidad Mucosa , Ratones , Modelos AnimalesRESUMEN
Idiopathic pneumonia syndrome (IPS) is a common, often fatal, complication following hematopoietic stem cell transplantation (HSCT) characterized by severe pneumonitis and interstitial fibrosis. Fully reconstituted syngeneic bone marrow transplant (BMT) mice infected with murine γ-herpesvirus-68 develop interleukin-17 (IL-17)-driven pneumonitis and fibrosis, which mimics clinical manifestations of IPS. We found CD103+ and CD11b+ dendritic cells (DCs) are selectively deficient for the Notch ligand, DLL4, following BMT and CD4+ T cells isolated from lungs and spleens of infected BMT mice display Notch signaling defects. Mice transplanted with CD4-Cre-driven dominant-negative Notch transcriptional regulator Mastermind-Like (CD4-Cre-DNMAML (CCD) mice) bone marrow displayed elevated IL-17 and transforming growth factor-ß (TGF ß) in the lungs, a further expansion of T-helper type 17 (Th17) cells, and developed more fibrosis than wild-type (WT)-BMT mice. Culture of BMT lung leukocytes with recombinant Notch ligand, DLL4, restored Notch signaling and decreased production of IL-17. Adoptive transfer of CD11c+ DCs could restore Th1 and limit Th17 in WT-BMT but not CCD-BMT mice, indicating that a specific DC/CD4+ T-cell Notch interaction modulates IL-17 production following reconstitution in syngeneic BMT mice. Given recent clinical observations showing that patients with pulmonary complications post-transplant harbor occult herpesvirus infections, these data provide mechanistic insight and suggest potential therapies for these devastating conditions.
Asunto(s)
Células Dendríticas/inmunología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Infecciones por Herpesviridae/inmunología , Interleucina-17/metabolismo , Pulmón/patología , Neumonía/inmunología , Complicaciones Posoperatorias/inmunología , Rhadinovirus/inmunología , Células Th17/inmunología , Proteínas Adaptadoras Transductoras de Señales , Animales , Proteínas de Unión al Calcio , Células Cultivadas , Fibrosis , Infecciones por Herpesviridae/etiología , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Pulmón/virología , Activación de Linfocitos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neumonía/etiología , Neumonía/virología , Complicaciones Posoperatorias/virología , Receptores Notch/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
The utility of transbronchial biopsy in the management of pulmonary complications following hematopoietic stem cell transplantation (HSCT) has shown variable results. Herein, we examine the largest case series of patients undergoing transbronchial biopsy following HSCT. We performed a retrospective analysis of 130 transbronchial biopsy cases performed in patients with pulmonary complications post HSCT. Logistic regression models were applied to examine diagnostic yield, odds of therapy change and complications. The most common histologic finding on transbronchial biopsy was a nonspecific interstitial pneumonitis (n=24 cases, 18%). Pathogens identified by transbronchial biopsy were rare, occurring in <5% of cases. A positive transbronchial biopsy significantly increased the odds of a subsequent change in corticosteroid therapy (odds ratio (OR)=3.12; 95% confidence interval (CI) 1.18-8.23; P=0.02) but was not associated with a change in antibiotic therapy (OR=1.01; 95% CI 0.40-2.54; P=0.98) or changes in overall therapy (OR=1.92; 95% CI 0.79-4.70; P=0.15). Patients who underwent a transbronchial biopsy had increased odds of complications related to the bronchoscopy (OR=3.33, 95% CI 1.63-6.79; P=0.001). In conclusion, transbronchial biopsy may contribute to the diagnostic management of noninfectious lung injury post HSCT, whereas its utility in the management of infectious pulmonary complications of HSCT remains low.
Asunto(s)
Broncoscopía/métodos , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Pulmón/patología , Acondicionamiento Pretrasplante/efectos adversos , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
Hematopoietic stem cell transplant (HSCT) treats or cures a variety of hematological and inherited disorders. Unfortunately, patients who undergo HSCT are susceptible to infections by a wide array of opportunistic pathogens. Pseudomonas aeruginosa bacteria can have life-threatening effects in HSCT patients by causing lung pathology that has been linked to high levels of the potent pro-inflammatory cytokine, interleukin-1ß (IL-1ß). Using a murine bone marrow transplant (BMT) model, we show that overexpression of prostaglandin E2 (PGE2) post-BMT signals via EP2 or EP4 to induce cyclic adenosine monophosphate (cAMP), which activates protein kinase A or the exchange protein activated by cAMP (Epac) to induce cAMP response element binding-dependent transcription of IL-1ß leading to exacerbated lung injury in BMT mice. Induction of IL-1ß by PGE2 is time and dose dependent. Interestingly, IL-1ß processing post-P. aeruginosa infection occurs via the enzymatic activity of either caspase-1 or caspase-8. Furthermore, PGE2 can limit autophagy-mediated killing of P. aeruginosa in alveolar macrophages, yet autophagy does not have a role in PGE2-mediated upregulation of IL-1ß. Reducing PGE2 levels with indomethacin improved bacterial clearance and reduced IL-1ß-mediated acute lung injury in P. aeruginosa-infected BMT mice.
Asunto(s)
Dinoprostona/metabolismo , Trasplante de Células Madre Hematopoyéticas , Lesión Pulmonar/inmunología , Macrófagos Alveolares/inmunología , Complicaciones Posoperatorias/inmunología , Infecciones por Pseudomonas/inmunología , Pseudomonas aeruginosa/fisiología , Animales , Células Cultivadas , AMP Cíclico/metabolismo , Modelos Animales de Enfermedad , Humanos , Indometacina/uso terapéutico , Interleucina-1beta/metabolismo , Lesión Pulmonar/tratamiento farmacológico , Lesión Pulmonar/etiología , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Complicaciones Posoperatorias/tratamiento farmacológico , Infecciones por Pseudomonas/tratamiento farmacológico , Infecciones por Pseudomonas/etiología , Subtipo EP2 de Receptores de Prostaglandina E/metabolismo , Transducción de Señal , Regulación hacia ArribaRESUMEN
Interleukin-36γ (IL-36γ) is a member of novel IL-1-like proinflammatory cytokine family that are highly expressed in epithelial tissues and several myeloid-derived cell types. Little is known about the role of the IL-36 family in mucosal immunity, including lung anti-bacterial responses. We used murine models of IL-36γ deficiency to assess the contribution of IL-36γ in the lung during experimental pneumonia. Induction of IL-36γ was observed in the lung in response to Streptococcus pneumoniae (Sp) infection, and mature IL-36γ protein was secreted primarily in microparticles. IL-36γ-deficient mice challenged with Sp demonstrated increased mortality, decreased lung bacterial clearance and increased bacterial dissemination, in association with reduced local expression of type-1 cytokines, and impaired lung macrophage M1 polarization. IL-36γ directly stimulated type-1 cytokine induction from dendritic cells in vitro in a MyD88-dependent manner. Similar protective effects of IL-36γ were observed in a Gram-negative pneumonia model (Klebsiella pneumoniae). Intrapulmonary delivery of IL-36γ-containing microparticles reconstituted immunity in IL-36γ-/- mice. Enhanced expression of IL-36γ was also observed in plasma and bronchoalveolar lavage fluid of patients with acute respiratory distress syndrome because of pneumonia. These studies indicate that IL-36γ assumes a vital proximal role in the lung innate mucosal immunity during bacterial pneumonia by driving protective type-1 responses and classical macrophage activation.
Asunto(s)
Interleucina-1/sangre , Interleucina-1/metabolismo , Infecciones por Klebsiella/inmunología , Klebsiella pneumoniae/fisiología , Pulmón/inmunología , Macrófagos/inmunología , Infecciones Neumocócicas/inmunología , Neumonía/inmunología , Síndrome de Dificultad Respiratoria/inmunología , Streptococcus pneumoniae/fisiología , Adulto , Animales , Células Cultivadas , Femenino , Humanos , Inmunidad Innata , Inmunidad Mucosa , Interleucina-1/genética , Pulmón/microbiología , Macrófagos/microbiología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Regulación hacia ArribaRESUMEN
Fibrocytes are circulating mesenchymal precursors (CD45+, col 1+) recruited to fibrotic areas. Fibrocytes secrete profibrotic mediators including periostin; a matricellular protein that regulates cellular interactions with extracellular matrix (ECM) components. In bleomycin-induced fibrosis, periostin deficiency in structural or hematopoietic cells limits development of pulmonary fibrosis. To determine if hematopoietic-derived fibrocytes might secrete soluble factors to activate structural myofibroblast differentiation, wild-type (WT) fibroblasts were treated with conditioned medium from fibrocytes isolated from bleomycin-treated WT or periostin-/- mice. After 24 h we saw less α-smooth muscle actin expression in cells treated with conditioned medium from periostin-/- fibrocytes. Adoptive transfer of WT fibrocytes augmented lung fibrosis to a greater extent than transfer of fibrocytes from periostin-/- mice. In vitro analysis of fibrocytes and fibroblasts isolated from WT and periostin-/- mice treated with TGFß1 or periostin demonstrated co-regulation of mesenchymal activation and beta 1 integrin as a potential receptor for periostin on fibrocytes. Additionally, connective tissue growth factor (CTGF) mRNA expression was increased in fibrocytes treated with periostin whereas CTGF and lysl oxidase (LOX) mRNA expression was low in bleomycin-treated periostin-/- fibrocytes. These data suggest fibrocytes may augment bleomycin-induced fibrosis via secretion of periostin and other soluble factors that promote myofibroblast differentiation.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Pulmón/metabolismo , Células Madre Mesenquimatosas/fisiología , Miofibroblastos/fisiología , Fibrosis Pulmonar/inmunología , Traslado Adoptivo , Animales , Bleomicina , Moléculas de Adhesión Celular/genética , Diferenciación Celular , Células Cultivadas , Colágeno/metabolismo , Factor de Crecimiento del Tejido Conjuntivo/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Modelos Animales de Enfermedad , Proteínas de la Matriz Extracelular/genética , Proteínas de la Matriz Extracelular/metabolismo , Integrina beta1/metabolismo , Pulmón/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína-Lisina 6-Oxidasa/genética , Proteína-Lisina 6-Oxidasa/metabolismo , Fibrosis Pulmonar/inducido químicamenteRESUMEN
Hematopoietic stem cell transplantation (HSCT) efficacy is limited by numerous pulmonary complications. We developed a model of syngeneic bone marrow transplantion (BMT) followed by infection with murine gamma herpesvirus-68 that results in pneumonitis and fibrosis and mimics human "noninfectious" HSCT complications. BMT mice experience increased early lytic replication, but establish viral latency by 21 days post infection. CD4 T cells in BMT mice are skewed toward interleukin (IL)-17A rather than interferon (IFN)-γ production. Transplantation of bone marrow from Il-17a(-/-) donors or treatment with anti-IL-17A neutralization antibodies at late stages attenuates pneumonitis and fibrosis in infected BMT mice, suggesting that hematopoietic-derived IL-17A is essential for development of pathology. IL-17A directly influences activation and extracellular matrix production by lung mesenchymal cells. Lung CD11c+ cells of BMT mice secrete more transforming growth factor beta-ß1, and pro-TH17 mRNAs for IL-23 and IL-6, and less TH1-promoting cytokine mRNA for IFN-γ but slightly more IL-12 mRNA in response to viral infection. Adoptive transfer of non-BMT lung CD11c-enriched cells restores robust TH1 response and suppresses aberrant TH17 response in BMT mice to improve lung pathology. Our data suggest that "noninfectious" HSCT lung complications may reflect preceding viral infections and demonstrate that IL-17A neutralization may offer therapeutic advantage even after disease onset.
Asunto(s)
Células Presentadoras de Antígenos/inmunología , Trasplante de Médula Ósea , Infecciones por Herpesviridae/inmunología , Pulmón/patología , Neumonía/inmunología , Complicaciones Posoperatorias/inmunología , Rhadinovirus/fisiología , Células Th17/inmunología , Animales , Anticuerpos Neutralizantes/administración & dosificación , Células Cultivadas , Modelos Animales de Enfermedad , Fibrosis , Humanos , Interleucina-17/genética , Interleucina-17/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neumonía/etiología , Neumonía/prevención & control , Complicaciones Posoperatorias/prevención & control , Células Th17/virología , Latencia del Virus , Replicación ViralRESUMEN
BACKGROUND/AIMS: Idiopathic interstitial pneumonias (IIPs) are a diverse grouping of chronic pulmonary diseases characterised by varying degrees of pulmonary fibrosis. The triggers of the fibroproliferative process in IIP remain enigmatic but recent attention has been directed towards chemokine involvement in this process. METHODS: The expression of two chemokine receptors, CCR7 and CXCR4, and their respective ligands, CCL19, CCL21, and CXCL12, were examined in surgical lung biopsies (SLBs) from patients with IIP. Transcript and protein expression of these receptors and their ligands was compared with that detected in histologically normal margin SLBs. RESULTS: CCR7 and CXCR4 were detected by gene array and real time polymerase chain reaction analysis and CCR7, but not CXCR4, expression was significantly raised in usual interstitial pneumonia (UIP) relative to biopsies from patients diagnosed with non-specific interstitial pneumonia (NSIP) or respiratory bronchiolitis/interstitial lung disease (RBILD). CCR7 protein was expressed in interstitial areas of all upper and lower lobe UIP SLBs analysed. CCR7 expression was present in 50% of NSIP SLBs, and CCR7 was restricted to blood vessels and mononuclear cells in 75% of RBILD SLBs. Immune cell specific CXCR4 expression was seen in IIP and normal margin biopsies. CCR7 positive areas in UIP biopsies were concomitantly positive for CD45 (the leucocyte common antigen) but CCR7 positive areas in all IIP SLBs lacked the haemopoietic stem cell antigen CD34, collagen 1, and alpha smooth muscle actin. CONCLUSION: This molecular and immunohistochemical analysis showed that IIPs are associated with abnormal CCR7 transcript and protein expression.
Asunto(s)
Enfermedades Pulmonares Intersticiales/metabolismo , Receptores de Quimiocina/metabolismo , Actinas/metabolismo , Quimiocina CCL19 , Quimiocina CCL21 , Quimiocina CXCL12 , Quimiocinas CC/genética , Quimiocinas CC/metabolismo , Quimiocinas CXC/genética , Quimiocinas CXC/metabolismo , Ensayo de Inmunoadsorción Enzimática/métodos , Expresión Génica , Humanos , Antígenos Comunes de Leucocito/metabolismo , Ligandos , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Reacción en Cadena de la Polimerasa/métodos , Receptores CCR7 , Receptores CXCR4/metabolismo , Receptores de Quimiocina/genéticaRESUMEN
Pulmonary fibrosis can be modeled in animals by intratracheal instillation of FITC, which results in acute lung injury, inflammation, and extracellular matrix deposition. We have previously shown that despite chronic inflammation, this model of pulmonary fibrosis is lymphocyte independent. The CC chemokine monocyte-chemoattractant protein-1 is induced following FITC deposition. Therefore, we have investigated the contribution of the main monocyte-chemoattractant protein-1 chemokine receptor, CCR2, to the fibrotic disease process. We demonstrate that CCR2(-/-) mice are protected from fibrosis in both the FITC and bleomycin pulmonary fibrosis models. The protection is specific for the absence of CCR2, as CCR5(-/-) mice are not protected. The protection is not explained by differences in acute lung injury, or the magnitude or composition of inflammatory cells. FITC-treated CCR2(-/-) mice display differential patterns of cellular activation as evidenced by the altered production of cytokines and growth factors following FITC inoculation compared with wild-type controls. CCR2(-/-) mice have increased levels of GM-CSF and reduced levels of TNF-alpha compared with FITC-treated CCR2(+/+) mice. Thus, CCR2 signaling promotes a profibrotic cytokine cascade following FITC administration.
Asunto(s)
Fibrosis Pulmonar/etiología , Receptores de Quimiocina/deficiencia , Animales , Bleomicina/farmacología , Quimiocina CCL2/biosíntesis , Citocinas/metabolismo , Fluoresceína-5-Isotiocianato/farmacología , Ratones , Ratones Mutantes , Fibrosis Pulmonar/inducido químicamente , Receptores CCR2 , Receptores CCR5/deficiencia , Receptores CCR5/genética , Receptores de Quimiocina/genética , Transducción de SeñalRESUMEN
We hypothesized that pulmonary granulocyte-macrophage colony-stimulating factor (GM-CSF) is critically involved in determining the functional capabilities of alveolar macrophages (AM) for host defense. To test this hypothesis, cells were collected by lung lavage from GM-CSF mutant mice [GM(-/-)] and C57BL/6 wild-type mice. GM(-/-) mice yielded almost 4-fold more AM than wild-type mice. The percentage of cells positive for the beta(2)-integrins CD11a and CD11c was reduced significantly in GM(-/-) AM compared with wild-type cells, whereas expression of CD11b was similar in the two groups. The phagocytic activity of GM(-/-) AM for FITC-labeled microspheres was impaired significantly compared with that of wild-type AM both in vitro and in vivo (after intratracheal inoculation with FITC-labeled beads). Stimulated secretion of tumor necrosis factor-alpha (TNF-alpha) and leukotrienes by AM from the GM(-/-) mice was greatly reduced compared with wild-type AM, whereas secretion of monocyte chemoattractant protein-1 was increased. Transgenic expression of GM-CSF exclusively in the lungs of GM(-/-) mice resulted in AM with normal or supranormal expression of CD11a and CD11c, phagocytic activity, and TNF-alpha secretion. Thus, in the absence of GM-CSF, AM functional capabilities for host defense were significantly impaired but were restored by lung-specific expression of GM-CSF.
Asunto(s)
Factor Estimulante de Colonias de Granulocitos y Macrófagos/fisiología , Macrófagos Alveolares/fisiología , Animales , Lavado Broncoalveolar , Antígenos CD18/metabolismo , Adhesión Celular , Quimiocina CCL2/metabolismo , Citometría de Flujo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/deficiencia , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Leucotrienos/metabolismo , Macrófagos Alveolares/citología , Macrófagos Alveolares/inmunología , Ratones , Ratones Transgénicos , Microesferas , Fagocitosis , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
BACKGROUND: The purpose of this study was to describe the community-based impact of near-fatal asthma within the District of Columbia (Washington, DC). METHODS: The design was a prospective cohort study. Subjects included all persons in 1993 who presented to Washington, DC hospitals alive, requiring intubation for respiratory failure (including subjects who subsequently died in the hospital). Washington, DC hospitals were contacted on a biweekly basis to identify subjects. Patients were contacted by mail, followed by an interview with the subject or proxy. RESULTS: Of the 35 case subjects identified, 31 (88.6%) were interviewed. Sixty-one percent of the subjects were female; 84% were African-American; and 45.2% were less than 18 years old. Forty-five percent had asthma for 10 or more years. Twenty-three percent reported the emergency department as their usual source of health care, and 32% saw a provider on a weekly basis. Fifty-two percent were taking four or more prescription medications, and 29% were taking no anti-inflammatory medications. In the 24 hours before the event, 77% reported difficulty breathing, but only 64% reported contacting a health care provider. CONCLUSIONS: Community-based investigation of near-fatal asthma may lead to a better characterization of risk factors associated with this event. Findings from this study suggest that some of the factors associated with near-fatal events may be different from those associated with fatal asthma and that up to one third of the events may have been preventable.
Asunto(s)
Asma/mortalidad , Adolescente , Adulto , Asma/epidemiología , Causas de Muerte , Estudios Transversales , District of Columbia , Femenino , Humanos , Seguro de Salud , Masculino , Persona de Mediana Edad , Factores de Riesgo , Tasa de SupervivenciaRESUMEN
Pulmonary infectious diseases cause significant morbidity and mortality in both industrialized and developing countries. Adaptive immune responses are required to defend the lung against pathogens that survive in normal macrophages and extracellular organisms that evade phagocytosis. Microbes initiate both innate immune responses and specific adaptive immune responses. Innate immune response molecules regulate T-lymphocyte differentiation. Activated T-lymphocytes provide cytokines, which activate macrophages and lytic signals that lyse infected antigen-presenting cells. Antibodies produced by plasma cells facilitate microbial clearance through diverse effector mechanisms including opsonization, complement fixation and antibody-dependent cytotoxicity. Lymphocytes determine the specificity of the immune response and orchestrate effector limbs of the immune response.
Asunto(s)
Formación de Anticuerpos/inmunología , Linfocitos B/inmunología , Inmunidad Celular/inmunología , Infecciones/inmunología , Pulmón/inmunología , Linfocitos T/inmunología , Animales , Citocinas/inmunología , Citocinas/metabolismo , Citotoxicidad Inmunológica , Humanos , Inmunoglobulinas/inmunología , Activación de Linfocitos , Fagocitosis , Subgrupos de Linfocitos T/inmunologíaRESUMEN
Mice deleted for the plasminogen activator inhibitor-1 (PAI-1) gene are relatively protected from developing pulmonary fibrosis induced by bleomycin. We hypothesized that PAI-1 deficiency reduces fibrosis by promoting plasminogen activation and accelerating the clearance of fibrin matrices that accumulate within the damaged lung. In support of this hypothesis, we found that the lungs of PAI-1(-/-) mice accumulated less fibrin after injury than wild-type mice, due in part to enhanced fibrinolytic activity. To further substantiate the importance of fibrin removal as the mechanism by which PAI-1 deficiency limited bleomycin-induced fibrosis, bleomycin was administered to mice deficient in the gene for the Aalpha-chain of fibrinogen (fib). Contrary to our expectation, fib(-/-) mice developed pulmonary fibrosis to a degree similar to fib(+/-) littermate controls, which have a plasma fibrinogen level that is 70% of that of wild-type mice. Although elimination of fibrin from the lung was not in itself protective, the beneficial effect of PAI-1 deficiency was still associated with proteolytic activity of the plasminogen activation system. In particular, inhibition of plasmin activation and/or activity by tranexamic acid reversed both the accelerated fibrin clearance and the protective effect of PAI-1 deficiency. We conclude that protection from fibrosis by PAI-1 deficiency is dependent upon increased proteolytic activity of the plasminogen activation system; however, complete removal of fibrin is not sufficient to protect the lung.
Asunto(s)
Bleomicina/farmacología , Fibrinógeno/metabolismo , Fibrosis Pulmonar/metabolismo , Animales , Antifibrinolíticos/farmacología , Líquido del Lavado Bronquioalveolar/citología , Permeabilidad Capilar/efectos de los fármacos , Colágeno/efectos de los fármacos , Colágeno/metabolismo , Femenino , Fibrina/efectos de los fármacos , Fibrina/metabolismo , Fibrina/farmacocinética , Fibrinógeno/genética , Fibrinolisina/efectos de los fármacos , Fibrinolisina/metabolismo , Genotipo , Cinética , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Pulmón/patología , Masculino , Tasa de Depuración Metabólica , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Inhibidor 1 de Activador Plasminogénico/deficiencia , Inhibidor 1 de Activador Plasminogénico/genética , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/genética , Análisis de Supervivencia , Ácido Tranexámico/farmacologíaRESUMEN
Macrophage inflammatory protein-1alpha (MIP-1alpha/CCL3) is a CC chemokine required for optimal recruitment of leukocytes in response to cryptococcal Ags. MIP-1alpha is expressed in the lungs by day 6 post Cryptococcus neoformans infection and could play a role in the development of cell-mediated immunity. To address this possibility, wild-type (MIP-1alpha(+/+)) mice and MIP-1alpha knockout (MIP-1alpha(-/-)) mice were infected intratracheally with a highly virulent strain of C. neoformans (145A). MIP-1alpha message was detected in the lungs on days 3, 7, and 14 in MIP-1alpha(+/+) mice, but it was undetectable in MIP-1alpha(-/-) mice. On day 16, MIP-1alpha(-/-) mice had a 7-fold increase in C. neoformans burden in the lungs, but no decrease in pulmonary leukocyte recruitment. MIP-1alpha(+/+) and MIP-1alpha(-/-) mice had similar numbers of recruited lymphocytes and monocytes/macrophages. Notably, MIP-1alpha(-/-) mice had a significantly greater number of eosinophils. MIP-1alpha(-/-) mice had extremely high levels of serum IgE. This switch of immune response to a T(2) phenotype was associated with enhanced IL-4 and IL-13 expression in the lungs of MIP-1alpha(-/-) mice compared with MIP-1alpha (+/+) mice. Progression of pulmonary cryptococcosis in the presence of nonprotective T(2) immunity resulted in profound lung damage in MIP-1alpha(-/-) mice (eosinophilic crystal deposition, destruction of lung parenchyma, and pulmonary hemorrhage). Twelve-week survival was dramatically decreased in MIP-1alpha(-/-) mice. These studies, together with our previous studies, demonstrate that MIP-1alpha plays a role in both the afferent (T(1)/T(2) development) and efferent (T(1)-mediated leukocyte recruitment) phases of cell-mediated immunity to C. neoformans.
Asunto(s)
Quimiocinas CC/fisiología , Criptococosis/inmunología , Proteínas Inflamatorias de Macrófagos/fisiología , Linfocitos T/inmunología , Animales , Movimiento Celular/inmunología , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocinas CC/biosíntesis , Quimiocinas CC/genética , Quimiocinas CC/inmunología , Criptococosis/genética , Criptococosis/microbiología , Criptococosis/patología , Cryptococcus neoformans/crecimiento & desarrollo , Cryptococcus neoformans/inmunología , Eliminación de Gen , Sueros Inmunes/administración & dosificación , Sueros Inmunes/farmacología , Inmunidad Celular , Inmunoglobulina E/sangre , Inyecciones Intraperitoneales , Interferón gamma/biosíntesis , Interleucina-12/biosíntesis , Interleucina-13/biosíntesis , Interleucina-4/biosíntesis , Leucocitos/inmunología , Pulmón/inmunología , Pulmón/microbiología , Pulmón/patología , Enfermedades Pulmonares Fúngicas/genética , Enfermedades Pulmonares Fúngicas/inmunología , Enfermedades Pulmonares Fúngicas/microbiología , Enfermedades Pulmonares Fúngicas/patología , Proteínas Inflamatorias de Macrófagos/biosíntesis , Proteínas Inflamatorias de Macrófagos/genética , Proteínas Inflamatorias de Macrófagos/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fenotipo , Eosinofilia Pulmonar/genética , Eosinofilia Pulmonar/inmunología , Análisis de Supervivencia , Linfocitos T/metabolismoRESUMEN
To characterize the role of GM-CSF in pulmonary fibrosis, we have studied bleomycin-induced fibrosis in wild-type mice vs mice with a targeted deletion of the GM-CSF gene (GM-CSF-/- mice). Without GM-CSF, pulmonary fibrosis was worse both histologically and quantitatively. These changes were not related to enhanced recruitment of inflammatory cells because wild-type and GM-CSF-/- mice recruited equivalent numbers of cells to the lung following bleomycin. Interestingly, recruitment of eosinophils was absent in GM-CSF-/- mice. We investigated whether the enhanced fibrotic response in GM-CSF-/- animals was due to a deficiency in an endogenous down-regulator of fibrogenesis. Analysis of whole lung homogenates from saline- or bleomycin-treated mice revealed that GM-CSF-/- animals had reduced levels of PGE2. Additionally, alveolar macrophages were harvested from wild-type and GM-CSF-/- mice that had been exposed to bleomycin. Although bleomycin treatment impaired the ability of alveolar macrophages from wild-type mice to synthesize PGE2, alveolar macrophages from GM-CSF-/- mice exhibited a significantly greater defect in PGE2 synthesis than did wild-type cells. Exogenous addition of GM-CSF to alveolar macrophages reversed the PGE2 synthesis defect in vitro. Administration of the PG synthesis inhibitor, indomethacin, to wild-type mice during the fibrogenic phase postbleomycin worsened the severity of fibrosis, implying a causal role for PGE2 deficiency in the evolution of the fibrotic lesion. These data demonstrate that GM-CSF deficiency results in enhanced fibrogenesis in bleomycin-induced pulmonary fibrosis and indicate that one mechanism for this effect is impaired production of the potent antifibrotic eicosanoid, PGE2.
Asunto(s)
Bleomicina/farmacología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/fisiología , Prostaglandinas/fisiología , Fibrosis Pulmonar/etiología , Fibrosis Pulmonar/metabolismo , Animales , Bleomicina/administración & dosificación , División Celular/genética , Dinoprostona/antagonistas & inhibidores , Dinoprostona/biosíntesis , Dinoprostona/deficiencia , Esquema de Medicación , Factor Estimulante de Colonias de Granulocitos y Macrófagos/administración & dosificación , Factor Estimulante de Colonias de Granulocitos y Macrófagos/deficiencia , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Indometacina/administración & dosificación , Inyecciones Intramusculares , Inyecciones Intraperitoneales , Intubación Intratraqueal , Cinética , Recuento de Leucocitos , Leucotrieno C4/biosíntesis , Leucotrieno C4/deficiencia , Metabolismo de los Lípidos , Lípidos/biosíntesis , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/patología , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/genética , Cloruro de Sodio/administración & dosificaciónRESUMEN
The mechanism by which IFN-gamma up-regulates invariant chain mRNA in antigen-presenting cells has been under intensive investigation. This study shows that in murine monocytic cells the transcriptional up-regulation mediated by the invariant chain (Ii) upstream enhancer only accounts for part of the induction of Ii mRNA by IFN-gamma. We have identified an intronic region in the murine Ii gene that contributes to the transcriptional up-regulation by IFN-gamma. The region has S (H), X/X2 and Y boxes similar to those in MHC class II promoters and the Ii upstream enhancer. Mutations at the putative S, X and Y boxes have abolished the ability of the region to mediate Ii up-regulation by IFN-gamma. Consistent with the functions of these boxes, our findings reveal that the up-regulation of Ii transcription by IFN-gamma mediated by the intronic region is dependent on the induction of class II transactivator by IFN-gamma.
Asunto(s)
Antígenos de Diferenciación de Linfocitos B/genética , Antígenos de Histocompatibilidad Clase II/genética , Interferón gamma/farmacología , Intrones , Elementos de Respuesta , Secuencia de Bases , Células Cultivadas , Humanos , Datos de Secuencia Molecular , ARN Mensajero/análisis , Transcripción Genética/efectos de los fármacosRESUMEN
Although cells of the innate inflammatory response, such as macrophages and neutrophils, have been extensively studied in the arena of Gram-negative bacterial pneumonia, a role for T cells remains unknown. To study the role of specific T cell populations in bacterial pneumonia, mice deleted of their TCR beta- and/or delta-chain were intratracheally inoculated with Klebsiella pneumoniae. Gamma delta T cell knockout mice displayed increased mortality at both early and late time points. In contrast, mice specifically lacking only alpha beta-T cells were no more susceptible than wild-type mice. Pulmonary bacterial clearance in gamma delta-T cell knockout mice was unimpaired. Interestingly, these mice displayed increased peripheral blood dissemination. Rapid up-regulation of IFN-gamma and TNF-alpha gene expression, critical during bacterial infections, was markedly impaired in lung and liver tissue from gamma delta-T cell-deficient mice 24 h postinfection. The increased peripheral blood bacterial dissemination correlated with impaired hepatic bacterial clearance following pulmonary infection and increased hepatic injury as measured by plasma aspartate aminotransferase activity. Combined, these data suggest that mice lacking gamma delta-T cells have an impaired ability to resolve disseminated bacterial infections subsequent to the initial pulmonary infection. These data indicate that gamma delta-T cells comprise a critical component of the acute inflammatory response toward extracellular Gram-negative bacterial infections and are vital for the early production of the proinflammatory cytokines IFN-gamma and TNF-alpha.
Asunto(s)
Citocinas/genética , Regulación de la Expresión Génica/inmunología , Infecciones por Klebsiella/inmunología , Infecciones por Klebsiella/mortalidad , Klebsiella pneumoniae/inmunología , Neumonía Bacteriana/inmunología , Neumonía Bacteriana/mortalidad , Receptores de Antígenos de Linfocitos T gamma-delta/fisiología , Animales , Citocinas/biosíntesis , Predisposición Genética a la Enfermedad , Interferón gamma/biosíntesis , Interferón gamma/genética , Intubación Intratraqueal , Infecciones por Klebsiella/genética , Infecciones por Klebsiella/patología , Hígado/inmunología , Hígado/metabolismo , Hígado/microbiología , Hígado/patología , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/microbiología , Pulmón/patología , Recuento de Linfocitos , Linfopenia/genética , Linfopenia/inmunología , Linfopenia/mortalidad , Linfopenia/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neumonía Bacteriana/genética , Neumonía Bacteriana/patología , ARN Mensajero/biosíntesis , Receptores de Antígenos de Linfocitos T gamma-delta/deficiencia , Receptores de Antígenos de Linfocitos T gamma-delta/genética , Especificidad de la Especie , Análisis de Supervivencia , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Subgrupos de Linfocitos T/patología , Factor de Necrosis Tumoral alfa/biosíntesis , Factor de Necrosis Tumoral alfa/genéticaRESUMEN
The authors surveyed all 125 allopathic medical schools to determine the number of schools that had implemented a formal curriculum in managed care and how many had a substantial interest in a Web-based clearinghouse for managed care curricular resources. They describe the results of their survey and the Web site they developed, the Managed Care Education Clearinghouse.
Asunto(s)
Curriculum , Educación Médica Continua , Educación de Pregrado en Medicina , Internet , Programas Controlados de Atención en Salud , Recolección de Datos , Facultades de Medicina , Estados UnidosRESUMEN
Few studies have addressed the importance of vascular remodeling in the lung during the development of bleomycin-induced pulmonary fibrosis. For fibroplasia and deposition of extracellular matrix to occur, there must be a geometric increase in neovascularization. We hypothesized that net angiogenesis during the pathogenesis of fibroplasia and deposition of extracellular matrix during bleomycin-induced pulmonary fibrosis are dependent in part upon an overexpression of the angiogenic CXC chemokine, macrophage inflammatory protein-2 (MIP-2). To test this hypothesis, we measured MIP-2 by specific ELISA in whole lung homogenates in either bleomycin-treated or control CBA/J mice and correlated these levels with lung hydroxyproline. We found that lung tissue from mice treated with bleomycin, compared with that from saline-treated controls, demonstrated a significant increase in the presence of MIP-2 that was correlated to a greater angiogenic response and total lung hydroxyproline content. Neutralizing anti-MIP-2 Abs inhibited the angiogenic activity of day 16 bleomycin-treated lung specimens using an in vivo angiogenesis bioassay. Furthermore, when MIP-2 was depleted in vivo by passive immunization, bleomycin-induced pulmonary fibrosis was significantly reduced without a change in the presence of pulmonary neutrophils, fibroblast proliferation, or collagen gene expression. This was also paralleled by a reduction in angiogenesis. These results demonstrate that the angiogenic CXC chemokine, MIP-2, is an important factor that regulates angiogenesis/fibrosis in pulmonary fibrosis.
Asunto(s)
Bleomicina/toxicidad , Quimiocinas CXC/inmunología , Monocinas/inmunología , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/inmunología , Animales , Diferenciación Celular/inmunología , Movimiento Celular/inmunología , Quimiocina CXCL2 , Quimiocinas CXC/biosíntesis , Quimiocinas CXC/fisiología , Colágeno/biosíntesis , Colágeno/genética , Femenino , Fibroblastos/patología , Regulación de la Expresión Génica/inmunología , Sueros Inmunes/administración & dosificación , Inmunización Pasiva , Inmunohistoquímica , Pulmón/irrigación sanguínea , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Ratones , Ratones Endogámicos CBA , Monocinas/biosíntesis , Monocinas/fisiología , Neovascularización Patológica/inmunología , Neovascularización Patológica/fisiopatología , Neovascularización Patológica/prevención & control , Neutrófilos/patología , Fibrosis Pulmonar/metabolismo , Fibrosis Pulmonar/fisiopatología , Ratas , Ratas Long-EvansRESUMEN
Prostate cancer is the second leading cause of malignancy-related mortality in males in the United States. As a solid tumor, clinically significant tumor growth and metastasis are dependent on nutrients and oxygen supplied by tumor-associated neovasculature. As such, there is a selective tumorigenic advantage for those neoplasms that can produce angiogenic mediators. We show here that human prostate cancer cell lines can constitutively produce angiogenic CXC chemokines. Tumorigenesis of PC-3 prostate cancer cells was shown to be attributable, in part, to the production of the angiogenic CXC chemokine, interleukin (IL)-8. Neutralizing antisera to IL-8 inhibits PC-3 tumor growth in a human prostate cancer/SCID mouse model. Furthermore, angiogenic activity in PC-3 tumor homogenates was attributable to IL-8. In contrast, the Du145 prostate cancer cell line uses a different angiogenic CXC chemokine, GRO-alpha, to mediate tumorigenicity. Neutralizing antisera to GRO-alpha but not IL-8 reduced tumor growth in vivo and reduced the angiogenic activity in tumor homogenates. Thus, prostate cancer cell lines can use distinct CXC chemokines to mediate their tumorigenicity.