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1.
Allergy ; 69(12): 1666-72, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25118837

RESUMO

BACKGROUND: Glutathione S-transferase M1 (GSTM1) is a phase II enzyme and regulator of inflammatory signaling in airway epithelial cells. We have found upregulation of neutrophilic airway inflammation in atopic asthmatics expressing GSTM1 gene (GSTM1+) compared to GSTM1null asthmatics. We hypothesized that GSTM1 modulates NF-κB activation in bronchial epithelium in atopic asthmatics. We determined regulation of allergen-induced NF-κB activation in bronchial epithelium by GSTM1 in human atopic asthmatics in vivo. METHODS: Endobronchial biopsies and bronchoalveolar lavage fluid samples were collected from 13 GSTM1+ and 12 GSTM1null human atopic asthmatics at baseline and 24 h after segmental allergen challenge. A quantitative analysis of NF-κB activation in airway epithelium was accomplished using a polyclonal antibody against the phosphorylated p65 component of NF-κB. Elastase-positive neutrophils in the bronchial wall were quantified. RESULTS: Postallergen neutrophilia in airway subepithelium and epithelial lining fluid was greater in GSTM1+ compared to GSTM1null asthmatics. Airway eosinophilia was similar in GSTM1+ and GSTM1null asthmatics. Allergen-provoked NF-κB induction in bronchial epithelium was significantly greater in GSTM1+ compared to GSTM1null asthmatics. Activation of NF-κB activation in airway epithelial cells correlated with interleukin-8 concentrations and absolute neutrophil numbers in bronchoalveolar lavage fluid in GSTM1+ but not GSTM1null asthmatics. CONCLUSIONS: Allergen-induced neutrophilic airway inflammation in GSTM1+ asthmatics is associated with NF-κB activation in airway epithelial cells in vivo. These novel data provide a potential mechanism of the genomic link between GSTM1 polymorphism and airway neutrophilia in atopic asthma.


Assuntos
Alérgenos/imunologia , Asma/imunologia , Asma/metabolismo , Glutationa Transferase/metabolismo , NF-kappa B/metabolismo , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Adulto , Animais , Asma/diagnóstico , Asma/tratamento farmacológico , Líquido da Lavagem Broncoalveolar/imunologia , Citocinas/metabolismo , Ativação Enzimática , Eosinófilos/imunologia , Eosinófilos/patologia , Feminino , Genótipo , Glutationa Transferase/genética , Humanos , Masculino , Pessoa de Meia-Idade , Neutrófilos/imunologia , Neutrófilos/patologia , Mucosa Respiratória/patologia , Adulto Jovem
2.
Oncogene ; 31(26): 3164-76, 2012 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-22002309

RESUMO

The mechanisms by which chronic inflammatory lung diseases, particularly chronic obstructive pulmonary disease, confer enhanced risk for lung cancer are not well-defined. To investigate whether nuclear factor (NF)-κB, a key mediator of immune and inflammatory responses, provides an interface between persistent lung inflammation and carcinogenesis, we utilized tetracycline-inducible transgenic mice expressing constitutively active IκB kinase ß in airway epithelium (IKTA (IKKß trans-activated) mice). Intraperitoneal injection of ethyl carbamate (urethane), or 3-methylcholanthrene (MCA) and butylated hydroxytoluene (BHT) was used to induce lung tumorigenesis. Doxycycline-treated IKTA mice developed chronic airway inflammation and markedly increased numbers of lung tumors in response to urethane, even when transgene expression (and therefore epithelial NF-κB activation) was begun after exposure to carcinogen. Studies using a separate tumor initiator/promoter model (MCA+BHT) indicated that NF-κB functions as an independent tumor promoter. Enhanced tumor formation in IKTA mice was preceded by increased proliferation and reduced apoptosis of alveolar epithelium, resulting in increased formation of premalignant lesions. Investigation of inflammatory cells in lungs of IKTA mice revealed a substantial increase in macrophages and lymphocytes, including functional CD4+/CD25+/FoxP3+ regulatory T lymphocytes (Tregs). Importantly, Treg depletion using repetitive injections of anti-CD25 antibodies limited excessive tumor formation in IKTA mice. At 6 weeks following urethane injection, antibody-mediated Treg depletion in IKTA mice reduced the number of premalignant lesions in the lungs in association with an increase in CD8 lymphocytes. Thus, persistent NF-κB signaling in airway epithelium facilitates carcinogenesis by sculpting the immune/inflammatory environment in the lungs.


Assuntos
Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/patologia , NF-kappa B/metabolismo , Transdução de Sinais , Linfócitos T Reguladores/imunologia , Animais , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Doença Crônica , Epitélio/efeitos dos fármacos , Epitélio/imunologia , Epitélio/metabolismo , Epitélio/patologia , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/induzido quimicamente , Neoplasias Pulmonares/metabolismo , Camundongos , Comunicação Parácrina/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Fatores de Tempo , Uretana/efeitos adversos
3.
Oncogene ; 30(12): 1402-12, 2011 Mar 24.
Artigo em Inglês | MEDLINE | ID: mdl-21076466

RESUMO

The transcription factor nuclear factor kappa B (NF-κB) is activated in human breast cancer tissues and cell lines. However, it is unclear whether NF-κB activation is a consequence of tumor formation or a contributor to tumor development. We developed a doxycycline (dox)-inducible mouse model, termed DNMP, to inhibit NF-κB activity specifically within the mammary epithelium during tumor development in the polyoma middle T oncogene (PyVT) mouse mammary tumor model. DNMP females and PyVT littermate controls were treated with dox from 4 to 12 weeks of age. We observed an increase in tumor latency and a decrease in final tumor burden in DNMP mice compared with PyVT controls. A similar effect with treatment from 8 to 12 weeks indicates that outcome is independent of effects on postnatal virgin ductal development. In both cases, DNMP mice were less likely to develop lung metastases than controls. Treatment from 8 to 9 weeks was sufficient to impact primary tumor formation. Inhibition of NF-κB increases apoptosis in hyperplastic stages of tumor development and decreases proliferation at least in part by reducing Cyclin D1 expression. To test the therapeutic potential of NF-κB inhibition, we generated palpable tumors by orthotopic injection of PyVT cells and then treated systemically with the NF-κB inhibitor thymoquinone (TQ). TQ treatment resulted in a reduction in tumor volume and weight as compared with vehicle-treated control. These data indicate that epithelial NF-κB is an active contributor to tumor progression and demonstrate that inhibition of NF-κB could have a significant therapeutic impact even at later stages of mammary tumor progression.


Assuntos
Epitélio/metabolismo , Epitélio/patologia , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , NF-kappa B/metabolismo , Carga Tumoral , Animais , Apoptose , Linhagem Celular Tumoral , Modelos Animais de Doenças , Doxiciclina/toxicidade , Epitélio/efeitos dos fármacos , Feminino , Glândulas Mamárias Animais/efeitos dos fármacos , Glândulas Mamárias Animais/metabolismo , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Animais/induzido quimicamente , Camundongos , Camundongos Transgênicos , NF-kappa B/antagonistas & inibidores
4.
Gut ; 58(9): 1267-74, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19625278

RESUMO

BACKGROUND AND AIMS: Autoimmune pancreatitis (AIP) is a poorly understood human disease affecting the exocrine pancreas. The goal of the present study was to elucidate the pathogenic mechanisms underlying pancreatic autoimmunity in a murine disease model. METHODS: A transgenic mouse with an S100A4/fibroblast-specific protein 1 (FSP1) Cre-mediated conditional knockout of the transforming growth factor beta (TGFbeta) type II receptor, termed Tgfbr2(fspKO), was used to determine the direct role of TGFbeta in S100A4(+) cells. Immunohistochemical studies suggested that Tgfbr2(fspKO) mice develop mouse AIP (mAIP) characterised by interlobular ductal inflammatory infiltrates and pancreatic autoantibody production. Fluorescence-activated cell sorting (FACS)-isolated dendritic cells (DCs) from diseased pancreata were verified to have S100A4-Cre-mediated DNA recombination. RESULTS: The Tgfbr2(fspKO) mice spontaneously developed mAIP by 6 weeks of age. DCs were confirmed to express S100A4, a previously reported protein expressed by fibroblasts. Adoptive transfer of bone marrow-derived DCs from Tgfbr2(fspKO) mice into 2-week-old syngenic wild-type C57BL/6 mice resulted in reproduction of pancreatitis within 6 weeks. Similar adoptive transfer of wild-type DCs had no effect on pancreas pathology of the host mice. The inability to induce pancreatitis by adoptive transfer of Tgfbr2(fspKO) DCs in adult mice suggested a developmental event in mAIP pathogenesis. Tgfbr2(fspKO) DCs undergo elevated maturation in response to antigen and increased activation of naïve CD4-positive T cells. CONCLUSION: The development of mAIP in the Tgfbr2(fspKO) mouse model illustrates the role of TGFbeta in maintaining myeloid DC immune tolerance. The loss of immune tolerance in myeloid S100A4(+) DCs can mediate mAIP and may explain some aspects of AIP disease pathogenesis.


Assuntos
Doenças Autoimunes/imunologia , Células Dendríticas/imunologia , Pancreatite/imunologia , Proteínas S100/metabolismo , Transdução de Sinais/fisiologia , Fator de Crescimento Transformador beta/metabolismo , Transferência Adotiva , Animais , Autoanticorpos/análise , Biomarcadores/análise , Proliferação de Células , Quimera , Células Dendríticas/metabolismo , Citometria de Fluxo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Animais , Proteínas Serina-Treonina Quinases/genética , Receptor do Fator de Crescimento Transformador beta Tipo II , Receptores de Fatores de Crescimento Transformadores beta/genética , Proteína A4 de Ligação a Cálcio da Família S100 , Linfócitos T/imunologia , beta-Galactosidase/análise
5.
Clin Exp Immunol ; 153(3): 420-8, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18647324

RESUMO

Pseudomonas aeruginosa pneumonia usually results from a deficit of the innate immune system. To investigate whether inflammatory signalling by airway epithelial cells provides a pivotal line of defence against P. aeruginosa infection, we utilized two separate lines of inducible transgenic mice that express a constitutive activator of the nuclear factor kappa-B (NF-kappaB) pathway (IKTA) or a dominant inhibitor of NF-kappaB (DNTA) in airway epithelial cells. Compared with control mice, IKTA mice showed an enhanced host response to P. aeruginosa infection with greater neutrophil influx into the lungs, increased expression of Glu-Leu-Arg-positive (ELR(+)) CXC chemokines macrophage inflammatory protein-2 and keratinocyte chemoattractant (KC), superior bacterial clearance and improved survival at 24 h after infection. Neutrophil depletion abrogated the improvement in host defence identified in IKTA mice. In contrast, DNTA mice showed impaired responses to P. aeruginosa infection with higher bacterial colony counts in the lungs, decreased neutrophilic lung inflammation and lower levels of KC in lung lavage fluid. DNTA mice given recombinant KC at the time of P. aeruginosa infection demonstrated improved neutrophil recruitment to the lungs and enhanced bacterial clearance. Our data indicate that the NF-kappaB pathway in airway epithelial cells plays an essential role in defence against P. aeruginosa through generation of CXC chemokines and recruitment of neutrophils.


Assuntos
Quimiocinas CXC/metabolismo , NF-kappa B/imunologia , Infiltração de Neutrófilos/imunologia , Infecções por Pseudomonas/imunologia , Pseudomonas aeruginosa/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Quimiocina CXCL2/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Epitélio/imunologia , Queratinócitos/metabolismo , Pulmão/imunologia , Camundongos , Camundongos Transgênicos , NF-kappa B/metabolismo
6.
Clin Exp Immunol ; 150(2): 245-54, 2007 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17672868

RESUMO

Cyclooxygenase-2 (COX-2) gene expression in the lung is induced in pathological conditions such as asthma and pneumonia; however, the exact impact of COX-2 gene expression in the airway in regulating inflammatory and immunological response in the lung is not understood. To define a physiological role of inducible COX-2 in airway epithelial cells, we developed a novel line of transgenic mice, referred to as CycloOxygenase-2 TransActivated (COTA) mice, that overexpress a COX-2 transgene in the distribution of the CC-10 promoter in response to doxycycline. In response to doxycycline treatment, COX-2 expression was increased in airway epithelium of COTA mice and whole lung tissue contained a three- to sevenfold increase in prostaglandin E(2) (PGE(2)), prostaglandin D(2) (PGD(2)) thromboxane B(2) (TXB(2)) and 6-Keto prostaglandin F(2alpha) (PGF(2alpha)) compared to wild-type and untreated COTA mice. Interestingly, primary mouse tracheal epithelial cells from COTA mice produced only PGE(2) by doxycycline-induced COX-2 activation, providing an indication of cellular specificity in terms of mediator production. In the ovalbumin model, in which doxycycline was given at the sensitization stage, there was an increase in interleukin (IL)-4 level in lung tissue from COTA mice compared to untreated COTA and wild-type mice. In addition, COTA mice that were treated with doxycycline had impaired clearance of Pseudomonas aeruginosa pneumonia compared to wild-type mice. COX-2 gene expression in airway epithelial cells has an important role in determining immunological response to infectious and allergic agents.


Assuntos
Brônquios/enzimologia , Ciclo-Oxigenase 2/imunologia , Regulação Enzimológica da Expressão Gênica/imunologia , Mucosa Respiratória/imunologia , Traqueia/enzimologia , Animais , Brônquios/imunologia , Células Cultivadas , Ciclo-Oxigenase 2/genética , Dinoprostona/biossíntese , Células Epiteliais/enzimologia , Células Epiteliais/imunologia , Genótipo , Imunidade nas Mucosas , Interleucina-4/biossíntese , Camundongos , Camundongos Transgênicos , Prostaglandinas/biossíntese , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/microbiologia , Pseudomonas aeruginosa/crescimento & desenvolvimento , Traqueia/imunologia
7.
Thorax ; 59(11): 977-80, 2004 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-15516475

RESUMO

BACKGROUND: While idiopathic pulmonary fibrosis (IPF) is one of the most common forms of interstitial lung disease, the aetiology of IPF is poorly understood. Familial cases of pulmonary fibrosis suggest a genetic basis for some forms of the disease. Recent reports have linked genetic mutations in surfactant protein C (SFTPC) with familial forms of pulmonary fibrosis, including one large family in which a number of family members were diagnosed with usual interstitial pneumonitis (UIP), the pathological correlate to IPF. Because of this finding in familial cases of pulmonary fibrosis, we searched for SFTPC mutations in a cohort of sporadic cases of UIP and non-specific interstitial pneumonitis (NSIP). METHODS: The gene for SFTPC was sequenced in 89 patients diagnosed with UIP, 46 patients with NSIP, and 104 normal controls. RESULTS: Ten single nucleotide polymorphisms in the SFTPC sequence were found in IPF patients and not in controls. Only one of these created an exonic change resulting in a change in amino acid sequence. In this case, a T to C substitution resulted in a change in amino acid 73 of the precursor protein from isoleucine to threonine. Of the remaining polymorphisms, one was in the 5' UTR, two were exonic without predicted amino acid sequence changes, and six were intronic. One intronic mutation suggested a potential enhancement of a splicing site. CONCLUSIONS: Mutations in SFTPC are identified infrequently in this patient population. These findings indicate that SFTPC mutations do not contribute to the pathogenesis of IPF in the majority of sporadic cases.


Assuntos
Doenças Pulmonares Intersticiais/genética , Mutação/genética , Proteína C Associada a Surfactante Pulmonar/genética , Feminino , Amplificação de Genes , Heterozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Polimorfismo Genético
8.
Curr Drug Targets ; 5(6): 581-8, 2004 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-15270205

RESUMO

The inflammatory response of the lung and airways is one of the main targets for tile development of new therapies for variety of disorders including the acute respiratory distress syndrome, cystic fibrosis, idiopathic pulmonary fibrosis, and chronic obstructive pulmonary disease. Over the last decade our understanding of the molecular biology of the inflammatory response has advanced considerably and has opened up new avenues for therapeutic intervention. Furthermore, the mechanism of action of many of the existing anti-inflammatory agents has been revealed by this burgeoning information. Here, we discuss the functions and therapeutic potential of molecules that might prove promising as targets for treatment of inflammatory lung diseases. These possible molecular targets include cell surface proteins/receptors [toll like receptors (TLRs), triggering receptors expressed on myeloid cells (TREMs), and syndecans)], transcription factors [NF-kappaB, AP-1, PU.1, and high mobility group box 1 (HMGB1)], and regulatory proteins [macrophage migration inhibitory factor (MIF), granulocyte macrophage colony stimulating factor (GM-CSF), cyclooxygenase 2 (COX-2), heme oxygenase 1 (HO-1)].


Assuntos
Lesão Pulmonar , Pneumonia/tratamento farmacológico , Receptores de Superfície Celular/efeitos dos fármacos , Humanos , Pulmão/efeitos dos fármacos , Modelos Biológicos , Neutrófilos/química , Pneumonia/etiologia , Pneumonia/patologia , Receptores de Superfície Celular/fisiologia , Transdução de Sinais/efeitos dos fármacos
9.
Eur Respir J ; 22(2): 197-202, 2003 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12952247

RESUMO

Conventional pleurodesing agents often provoke acute pleural inflammation followed by fibrosis. The inflammation frequently causes pain and fever. Transforming growth factor (TGF)-beta is a pro-fibrotic but anti-inflammatory cytokine. Intrapleural TGF-beta2 administration produces effective pleurodesis in animals, but its effects on mesothelial cells are unknown. The authors hypothesised that, unlike conventional pleurodesing agents, TGF-beta2 can induce collagen synthesis without stimulating pleural inflammation. In the in vitro studies, TGF-beta2, talc and doxycycline were administered to rabbit mesothelial cells for 24 h. These agents were also injected intrapleurally in rabbits and the induced pleural fluids collected at 24 h. TGF-beta2 was as potent as talc and doxycycline in upregulating mesothelial cell collagen expression. Talc and doxycycline both induced significant increases in interleukin (IL)-8 production from mesothelial cells in vitro and in rabbit pleural fluids in vivo. TGF-beta2, however, did not stimulate mesothelial cell IL-8 release in vitro and induced a dose-dependent suppression of pleural fluid IL-8. Pleural fluid IL-8 levels correlated significantly with leukocyte and lactate dehydrogenase concentrations in the fluids. In summary, transforming growth factor-beta was a potent inducer of mesothelial cell collagen synthesis. Unlike talc and tetracycline, which provoked pleural inflammation, transforming growth factor-beta2 suppressed pleural inflammation in vivo. Transforming growth factor-beta2 can produce effective pleural fibrosis without necessitating acute pleural inflammation.


Assuntos
Colágeno/biossíntese , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Interleucina-8/biossíntese , Pleura/efeitos dos fármacos , Fator de Crescimento Transformador beta/administração & dosagem , Animais , Antibacterianos/administração & dosagem , Células Cultivadas , Relação Dose-Resposta a Droga , Doxiciclina/administração & dosagem , Injeções , Pleura/metabolismo , Coelhos , Talco/administração & dosagem
10.
Am J Respir Crit Care Med ; 164(5): 873-8, 2001 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-11549548

RESUMO

We examined the effects of dexamethasone treatment on nuclear factor (NF)-kappa B activation and lung inflammation in transgenic reporter mice expressing photinus luciferase under the control of an NF-kappa B-dependent promoter (HLL mice). In vitro studies with bone marrow and peritoneal macrophages derived from these mice showed that treatment with dexamethasone blocked luciferase induction after treatment with Escherichia coli lipopolysaccharide (LPS); however, treatment of mice with intraperitoneal injection of dexamethasone at doses of 0.3 microg/g and 1 microg/g failed to inhibit NF-kappa B-dependent luciferase activity in the lungs. Furthermore, intraperitoneal treatment with 10 microg/g of dexamethasone prior to LPS paradoxically resulted in augmented luciferase activity as compared with that of mice treated with LPS alone. NF-kappa B-dependent luciferase expression in the lungs was detected by bioluminescence imaging and by measurement of luciferase activity in homogenized lung tissue. In these studies, there was an excellent correlation between indirect measurement of luciferase activity by bioluminescence in living mice and direct measurement of luciferase activity in lung tissue. Dexamethasone treatment did not affect LPS-induced neutrophilic influx or the concentration of macrophage inflammatory protein-2 in lung lavage fluid. These findings emphasize the potential error of extrapolating in vitro findings to complex in vivo events such as regulation of inflammation.


Assuntos
Dexametasona/administração & dosagem , Glucocorticoides/administração & dosagem , NF-kappa B/efeitos dos fármacos , NF-kappa B/fisiologia , Animais , Lipopolissacarídeos/administração & dosagem , Macrófagos/metabolismo , Camundongos , Camundongos Transgênicos
11.
Infect Immun ; 69(10): 5991-6, 2001 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-11553535

RESUMO

Reactive oxygen species (ROS) are thought to be involved in intracellular signaling, including activation of the transcription factor NF-kappaB. We investigated the role of NADPH oxidase in the NF-kappaB activation pathway by utilizing knockout mice (p47phox-/-) lacking the p47phox component of NADPH oxidase. Wild-type (WT) controls and p47phox-/- mice were treated with intraperitoneal (i.p.) Escherichia coli lipopolysaccharide (LPS) (5 or 20 microg/g of body weight). LPS-induced NF-kappaB binding activity and accumulation of RelA in nuclear protein extracts of lung tissue were markedly increased in WT compared to p47phox-/- mice 90 min after treatment with 20 but not 5 microg of i.p. LPS per g. In another model of lung inflammation, RelA nuclear translocation was reduced in p47phox-/- mice compared to WT mice following treatment with aerosolized LPS. In contrast to NF-kappaB activation in p47phox-/- mice, LPS-induced production of macrophage inflammatory protein 2 in the lungs and neutrophilic lung inflammation were not diminished in these mice compared to WT mice. We conclude that LPS-induced NF-kappaB activation is deficient in the lungs of p47phox-/- mice compared to WT mice, but this abnormality does not result in overt alteration in the acute inflammatory response.


Assuntos
Pulmão/imunologia , NADPH Oxidases/imunologia , NF-kappa B/imunologia , Fosfoproteínas/imunologia , Animais , Transporte Biológico , Extratos Celulares , Núcleo Celular , Quimiocina CXCL2 , Quimiocinas/biossíntese , Quimiocinas CXC/biossíntese , Escherichia coli , Feminino , Hepatócitos , Injeções Intraperitoneais , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/imunologia , Fígado/citologia , Fígado/imunologia , Pulmão/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NADPH Oxidases/genética , Neutrófilos/imunologia , Fosfoproteínas/genética , Fator de Transcrição RelA
14.
Inflammation ; 25(3): 145-55, 2001 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-11403205

RESUMO

Although pulmonary inflammation is an important pathologic event in cystic fibrosis (CF), the relationship between expression of the CF gene and the inflammatory response is unclear. We studied tumor necrosis factor (TNF) alpha and IL-1beta stimulated production of IL-6 and IL-8 by CF, corrected CF, and normal human bronchial epithelial cells in culture. During the first 24 hours of TNFalpha stimulation, CF cells produced significantly more IL-8 than normal or corrected CF cells. In the second 24 hours of TNFalpha stimulation, IL-6 and IL-8 generation ceased in normal and corrected CF cells but accelerated in CF cells, resulting in marked IL-6 and IL-8 accumulation in CF cells. Similar results were found when cells were stimulated with IL-1beta. Finally, when CF cells were grown at 27 degrees C (a culture condition which results in transport of CF transmembrane conductance regulator, CFTR, to the cell membrane and normalization of chloride conductance) TNFalpha-stimulated production of IL-6 and IL-8 reverted to normal. We conclude that dysregulation of cytokine generation by CF bronchial epithelial cells is directly related to expression of mutant CFTR and these observations provide a potential mechanism for persistence of airway inflammation in CF.


Assuntos
Brônquios/imunologia , Fibrose Cística/imunologia , Citocinas/biossíntese , Células Cultivadas , Fibrose Cística/etiologia , Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Células Epiteliais/imunologia , Humanos , Inflamação/etiologia , Mediadores da Inflamação/metabolismo , Interleucina-1/farmacologia , Interleucina-6/biossíntese , Interleucina-8/biossíntese , Fator de Necrose Tumoral alfa/farmacologia
15.
Inflammation ; 25(1): 25-31, 2001 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-11293663

RESUMO

Activation of NF-kappaB and production of NF-kappaB-dependent chemokines are thought to be involved in the pathogenesis of neutrophilic lung inflammation. Calpain-1 inhibitor (CI-1) blocks activation of NF-kappaB by preventing proteolysis of the inhibitory protein IkappaB-alpha by the ubiquitin/proteasome pathway. We hypothesized that inhibition of proteasome function with CI-1 would block NF-kappaB activation in vivo after intraperitoneal (i.p.) treatment with bacterial lipopolysaccharide (LPS), and that NF-kappaB inhibition would be associated with suppression of chemokine gene expression and attenuation of neutrophilic alveolitis. We treated rats with a single i.p. injection of CI-1 (10 mg/kg) two hours prior to i.p. LPS (7 mg/kg). Treatment with Cl-1 prevented degradation of IkappaB-alpha and activation of NF-kappaB in the liver in response to LPS; however, Cl-1 treatment had no detected effect on NF-kappaB activation in lung tissue. CI-1 treatment prior to LPS resulted in 40% lower MIP-2 concentration in lung lavage fluid compared to rats treated with vehicle prior to LPS (502 +/- 112 pg/ml vs. 859 +/-144 pg/ml, P < 0.05). In addition, CI-1 treatment substantially inhibited LPS-induced neutrophilic alveolitis (2.7+ /- 1.2 x 10(5) vs. 43.7 +/- 12.2 x 10(5) lung lavage neutrophils, P < 0.01). These data indicate that NF-kappaB inhibition in the liver can alter lung inflammation induced by systemic LPS treatment and suggest that a liver-lung interaction contributes to the inflammatory response of the lung.


Assuntos
Glicoproteínas/uso terapêutico , Proteínas I-kappa B , Fígado/efeitos dos fármacos , Pulmão/metabolismo , Complexos Multienzimáticos/antagonistas & inibidores , NF-kappa B/antagonistas & inibidores , Neutrófilos/fisiologia , Pneumonia/prevenção & controle , Inibidores de Proteases/uso terapêutico , Animais , Líquido da Lavagem Broncoalveolar/química , Quimiocina CXCL2 , Quimiocinas/análise , Quimiocinas/biossíntese , Cisteína Endopeptidases , Proteínas de Ligação a DNA/metabolismo , Endotoxemia/complicações , Endotoxemia/genética , Endotoxemia/imunologia , Regulação da Expressão Gênica/efeitos dos fármacos , Glicoproteínas/farmacologia , Lipopolissacarídeos/toxicidade , Fígado/metabolismo , Pulmão/patologia , Macrófagos Alveolares/efeitos dos fármacos , Masculino , Inibidor de NF-kappaB alfa , Fosforilação/efeitos dos fármacos , Pneumonia/etiologia , Pneumonia/genética , Pneumonia/imunologia , Inibidores de Proteases/farmacologia , Complexo de Endopeptidases do Proteassoma , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley
16.
Expert Opin Ther Targets ; 5(2): 197-204, 2001 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-15992176

RESUMO

The transcription factor NF-kappaB has been implicated in the pathogenesis of a variety of both acute and chronic inflammatory diseases. Many agents have shown promise and potential to abrogate NF-kappaB activity in both in vitro and in vivo systems. These include antioxidants, corticosteroids, proteasome inhibitors, arachadonic acid pathway metabolites, salicylates, molecular interventions and cell-permeable peptides. Unfortunately, therapies aimed at blocking its activation have not proven clinically feasible at this time. As the complex signal transduction pathways leading to NF-kappaB activation are further elucidated, more specific inhibitors of NF-kappaB are likely to be identified.

17.
Am J Respir Crit Care Med ; 162(3 Pt 1): 1095-101, 2000 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-10988136

RESUMO

We utilized a line of transgenic mice expressing Photinus luciferase complementary DNA (cDNA) under the control of a nuclear factor kappa B (NF-kappaB)-dependent promoter (from the 5' human immunodeficiency virus-1 [HIV-1] long terminal repeat) to examine the role of NF-kappaB activation in the pathogenesis of systemic inflammation induced by bacterial endotoxin (lipopolysaccharide [LPS]). After intraperitoneal injection of E. coli LPS, these mice displayed a time- and dose-dependent, organ-specific pattern of luciferase expression, showing that NF-kappaB-dependent gene transcription is transiently activated in multiple organs by systemic LPS administration. Luciferase expression in liver could be specifically blocked by intravenous administration of replication-deficient adenoviral vectors expressing a dominant inhibitor of NF-kappaB (IkappaB-alphaDN), confirming that luciferase gene expression is a surrogate marker for NF-kappaB activation in this line of mice. After treatment with intraperitoneal LPS, the mice were found to have increased lung tissue messenger RNA (mRNA) expression of a variety of cytokines that are thought to be NF-kappaB-dependent, as well as elevated serum concentrations of presumed NF-kappaB-dependent cytokines. In lung tissue homogenates, a close correlation was identified between luciferase activity and KC levels. These studies show that systemic treatment with LPS orchestrates a multiorgan NF-kappaB-dependent response that likely regulates the pathobiology of systemic inflammation.


Assuntos
Ampliador HIV/genética , Síndrome de Resposta Inflamatória Sistêmica/genética , Animais , Citocinas/genética , Citocinas/metabolismo , DNA Complementar/genética , Humanos , Lipopolissacarídeos/imunologia , Luciferases/genética , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Camundongos Transgênicos , RNA Mensageiro/genética , Síndrome de Resposta Inflamatória Sistêmica/imunologia
18.
Am J Respir Cell Mol Biol ; 23(3): 396-403, 2000 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-10970832

RESUMO

In cystic fibrosis (CF), inflammatory mediator production by airway epithelial cells is a critical determinant of chronic airway inflammation. To determine whether altered signal transduction through the nuclear factor (NF)-kappaB pathway occurs in CF epithelial cells and results in excessive generation of inflammatory cytokines, we evaluated tumor necrosis factor (TNF)-alpha-induced production of the NF-kappaB-dependent cytokine interleukin (IL)-8 and activation of NF-kappaB in three different human bronchial epithelial cell lines: (1) BEAS cells that express wild-type CF transmembrane conductance regulator (CFTR), (2) IB3 cells with mutant CFTR, and (3) C38 cells, which are "corrected" IB3 cells complemented with wild-type CFTR. Treatment of cells with TNF-alpha (30 ng/ml) resulted in markedly elevated NF-kappaB activation and production of IL-8 by IB3 cells compared with BEAS and C38 cells. Despite the differences in NF- kappaB activation, no differences in basal levels of IkappaB-alpha or TNF-alpha- induced IkappaB-alpha processing and degradation were detected among the cell lines. In contrast, the basal level of IkappaB-beta was increased in the IB3 cells. Treatment with TNF-alpha resulted in increased formation of hypophosphorylated IkappaB-beta and increased nuclear localization of IkappaB-beta in IB3 cells compared with the other cell types. These findings provide additional evidence of a dysregulated inflammatory response in CF.


Assuntos
Fibrose Cística/imunologia , Fibrose Cística/metabolismo , Proteínas I-kappa B/metabolismo , NF-kappa B/metabolismo , Mucosa Respiratória/metabolismo , Brônquios/citologia , Brônquios/imunologia , Brônquios/metabolismo , Células Cultivadas , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Expressão Gênica/imunologia , Humanos , Proteínas I-kappa B/imunologia , Immunoblotting , Interleucina-8/metabolismo , Ligases/metabolismo , Mutagênese/fisiologia , NF-kappa B/imunologia , Fosforilação , Mucosa Respiratória/citologia , Mucosa Respiratória/imunologia , Fator de Necrose Tumoral alfa/farmacologia
19.
Arch Surg ; 135(6): 667-72; discussion 672-3, 2000 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10843362

RESUMO

HYPOTHESIS: Hepatic cryoablation of 30% to 35% or more of liver parenchyma in a sheep model results in eicosanoid and nuclear factor-kappaB (NF-kappaB)-mediated changes in pulmonary hemodynamics and lung permeability. SETTING: Laboratory. INTERVENTIONS: At initial thoracotomy, catheters were placed in the main pulmonary artery, left atrium, right carotid artery, and efferent duct of the caudal mediastinal lymph node for subsequent monitoring in adult sheep. After a 1- to 2-week period of recovery, animals underwent laparotomy and left-lobe cryoablation (approximately 35% by volume) with subsequent awake monitoring and on postoperative days 1 to 3. MAIN OUTCOME MEASURES: Cryoablation-induced lung permeability and hemodynamic changes were compared with baseline values in sheep that underwent instrumentation. Similarly handled sheep underwent resection of a similar volume of hepatic parenchyma or had pulmonary artery pressure increases induced by mechanical left atrial obstruction. Activation of NF-kappaB was assessed with electrophoretic mobility shift assay, and serum thromboxane levels were measured with mass spectroscopy. RESULTS: Cryoablation resulted in acutely increased mean pulmonary (20 to 35 cm water) and systemic pressures, which returned to baseline at 24 hours with no change in cardiac output. Serum thromboxane levels increased 30 minutes after cryoablation (9-fold) and returned to baseline at 24 hours. Activation of NF-kappaB was present in liver and lung tissue by 30 minutes after cryoablation. Lung lymph-plasma protein clearance markedly exceeded the expected increase from pulmonary pressures alone, and increased lymph-plasma protein ratio persisted after pulmonary artery pressures normalized. Similar changes were not associated with 35% hepatic resection. CONCLUSIONS: This study demonstrates that 35% hepatic cryoablation results in an acute but transient increase in pulmonary artery pressure that may be mediated by increased thromboxane levels. Increases in pulmonary capillary permeability are not accounted for by pressure changes alone, and may be a result of NF-kappaB-mediated inflammatory mechanisms. These data show that cryosurgery causes pathophysiological changes similar to those observed with endotoxin and other systemic inflammatory stimuli.


Assuntos
Criocirurgia/efeitos adversos , Fígado/cirurgia , Síndrome do Desconforto Respiratório/etiologia , Animais , Proteínas Sanguíneas/metabolismo , Permeabilidade Capilar/fisiologia , Linfa/fisiologia , NF-kappa B/metabolismo , Artéria Pulmonar/fisiologia , Circulação Pulmonar/fisiologia , Ovinos , Tromboxanos/sangue
20.
Chest ; 117(5): 1482-7, 2000 May.
Artigo em Inglês | MEDLINE | ID: mdl-10807839

RESUMO

Nuclear factor-kappa B (NF-kappaB) is a family of DNA-binding protein factors that are required for transcription of most proinflammatory molecules, including adhesion molecules, enzymes, cytokines, and chemokines. NF-kappaB activation seems to be a key early event in a variety of cell and animal model systems developed to elucidate the pathobiology of lung diseases. The purpose of this short review is to describe what is known about the molecular biology of NF-kappaB and to review information that implicates NF-kappaB in the pathogenesis of lung disease, including ARDS, systemic inflammatory response syndrome, asthma, respiratory viral infections, occupational and environmental lung disease, and cystic fibrosis.


Assuntos
Pneumopatias/fisiopatologia , NF-kappa B/fisiologia , Animais , Humanos , Mediadores da Inflamação/metabolismo , Síndrome de Resposta Inflamatória Sistêmica/fisiopatologia
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