RESUMO
Recent developments in the study of host-pathogen interactions have fundamentally altered our understanding of the nature of Staphylococcus aureus infection, and previously held tenets regarding the role of the granulocyte are being cast aside. Novel mechanisms of pathogenesis are becoming evident, revealing the extent to which S. aureus can evade neutrophil responses successfully by resisting microbicides, surviving intracellularly and subverting cell death pathways. Developing a detailed understanding of these complex strategies is especially relevant in light of increasing staphylococcal virulence and antibiotic resistance, and the knowledge that dysfunctional neutrophil responses contribute materially to poor host outcomes. Unravelling the biology of these interactions is a challenging task, but one which may yield new strategies to address this, as yet, defiant organism.
Assuntos
Pele/microbiologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Abscesso/imunologia , Morte Celular , Interações Hospedeiro-Patógeno , Humanos , Hospedeiro Imunocomprometido , Contagem de Leucócitos , Neutrófilos/imunologia , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/microbiologia , VirulênciaRESUMO
There is increasing evidence for a central role in mammalian apoptosis of the interleukin-1 beta-converting enzyme (ICE) family of cysteine proteases, homologues of the product of the nematode "death" gene, ced-3. Ced-3 is thought to act as an executor rather than a regulator of programmed cell death in the nematode. However, it is not known whether mammalian ICE-related proteases (IRPs) are involved in the execution or the regulation of mammalian apoptosis. Moreover, an absolute requirement for one or more IRPs for mammalian apoptosis has yet to be established. We have used two cell-permeable inhibitors of IRPs, Z-Val-Ala-Asp.fluoromethylketone (ZVAD.fmk) and t-butoxy carbonyl-Asp.fluoromethylketone (BD.fmk), to demonstrate a critical role for IRPs in mammalian apoptosis induced by several disparate mechanisms (deregulated oncogene expression, ectopic expression of the Bcl-2 relative Bak, and DNA damage-induced cell death). In all instances, ZVAD.fmk and BD.fmk treatment inhibits characteristic biochemical and morphological events associated with apoptosis, including cleavage of nuclear lamins and poly-(ADP-ribose) polymerase, chromatin condensation and nucleosome laddering, and external display of phosphatidylserine. However, neither ZVAD.fmk nor BD.fmk inhibits the onset of apoptosis, as characterized by the onset of surface blebbing; rather, both act to delay completion of the program once initiated. In complete contrast, IGF-I and Bcl-2 delay the onset of apoptosis but have no effect on the kinetics of the program once initiated. Our data indicate that IRPs constitute part of the execution machinery of mammalian apoptosis induced by deregulated oncogenes, DNA damage, or Bak but that they act after the point at which cells become committed to apoptosis or can be rescued by survival factors. Moreover, all such blocked cells have lost proliferative potential and all eventually die by a process involving cytoplasmic blebbing.
Assuntos
Apoptose/fisiologia , Caspases , Cisteína Endopeptidases/fisiologia , Proteínas de Helminto/fisiologia , Clorometilcetonas de Aminoácidos/farmacologia , Animais , Sangue , Proteínas de Caenorhabditis elegans , Caspase 1 , Linhagem Celular , Membrana Celular , Inibidores de Cisteína Proteinase/farmacologia , Dano ao DNA , Fibroblastos , Expressão Gênica , Genes myc/fisiologia , Proteínas de Helminto/antagonistas & inibidores , Fator de Crescimento Insulin-Like I/fisiologia , Laminas , Proteínas de Membrana/genética , Microscopia de Vídeo , Proteínas Nucleares/metabolismo , Fosfatidilserinas/análise , Poli(ADP-Ribose) Polimerases/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Ratos , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Proteína Killer-Antagonista Homóloga a bcl-2RESUMO
Endothelin-1 (EDN1) has been reported to be implicated in the pathophysiology of asthma. Literature results on the genetic association of EDN1 in asthma are inconsistent. Eleven single nucleotide polymorphisms in EDN1 were genotyped in 342 and 100 families from UK and Norway, respectively. Asthma, bronchial hyperreactivity (BHR) and atopic asthma phenotypes were analyzed for the family-based association. Five single nucleotide polymorphisms (SNPs) were associated with asthma (0.0017Assuntos
Asma/genética
, Endotelina-1/genética
, Genética Populacional
, Polimorfismo de Nucleotídeo Único/genética
, Adolescente
, Adulto
, Alelos
, Estudos de Casos e Controles
, Criança
, Interpretação Estatística de Dados
, Família
, Feminino
, Frequência do Gene
, Marcadores Genéticos
, Haplótipos
, Humanos
, Desequilíbrio de Ligação
, Masculino
, Noruega
, Estatística como Assunto
, Reino Unido
RESUMO
Developing new treatments for chronic obstructive pulmonary disease (COPD) is extremely challenging. This disease, chronic by definition, becomes apparent only after substantial--and probably irreversible--tissue damage has occurred. The observable phenotype is of a stable disease state whose progression is hard to influence and reversal of which appears almost impossible. Identifying key components of the pathological process, targeting of which will result in substantial clinical benefit, is a significant challenge. In this review the nature of the disease is examined and conceptual information and simple tissue models of inflammation are used to explore the pathological network that is COPD. From the concept of COPD as a disease network displaying the features of contiguous immunity (in which many processes of innate and adaptive immunity are in continual dialogue and evolution), refinements are suggested to the strategies aimed at developing effective new treatments for this disease.
RESUMO
Elevation of cytosolic calcium ([Ca2+]i) has been reported to induce apoptosis in a number of cell types. However, in the neutrophil, which undergoes apoptosis constitutively during aging in vitro, activation by inflammatory mediators elevates [Ca2+]i and prolongs lifespan via inhibition of apoptosis. To examine this paradox, we investigated the effects of modulation of [Ca2+]i upon apoptosis of neutrophils in vitro. Calcium ionophores (A23187, ionomycin) retarded apoptosis in neutrophil populations after 20 h (P < 0.001). Conversely, intracellular Ca(2+)-chelation, using bis-(o-aminophenoxy)-N,N,N'N'-tetraacetic acid (BAPTA) acetoxymethyl ester (AM) promoted apoptosis (P < 0.02). W-7 (an inhibitor of calmodulin) also promoted apoptosis (P < 0.05). Measurements of [Ca2+]i, using fura-2, showed (a) increased apoptosis in neutrophil populations was not associated with elevated [Ca2+]i, (b) neutrophils cultured with ionophore at concentrations inhibiting apoptosis exhibited transient (< 1 h) elevations of [Ca2+]i, to levels previously reported with receptor-mediated stimuli, and (c) BAPTA was able to prevent the elevation of [Ca2+]i and the inhibition of apoptosis produced by ionophore. Modulation of apoptosis occurred without alterations in intracellular pH. Thus, in the neutrophil, unlike lymphoid cells, elevation of [Ca2+]i exerts an inhibitory effect upon apoptosis. Furthermore, these data suggest that transient elevation of [Ca2+]i elicits signaling events leading to prolonged inhibition of apoptosis.
Assuntos
Apoptose , Cálcio/metabolismo , Neutrófilos/fisiologia , Apoptose/efeitos dos fármacos , Calcimicina/farmacologia , Calmodulina/antagonistas & inibidores , Citosol/metabolismo , Ácido Egtázico/análogos & derivados , Ácido Egtázico/farmacologia , Endonucleases/metabolismo , Humanos , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Ionomicina/farmacologia , Sulfonamidas/farmacologiaRESUMO
Hypoxia and bacterial infection frequently co-exist, in both acute and chronic clinical settings, and typically result in adverse clinical outcomes. To ameliorate this morbidity, we investigated the interaction between hypoxia and the host response. In the context of acute hypoxia, both S. aureus and S. pneumoniae infections rapidly induced progressive neutrophil mediated morbidity and mortality, with associated hypothermia and cardiovascular compromise. Preconditioning animals through longer exposures to hypoxia, prior to infection, prevented these pathophysiological responses and profoundly dampened the transcriptome of circulating leukocytes. Specifically, perturbation of HIF pathway and glycolysis genes by hypoxic preconditioning was associated with reduced leukocyte glucose utilisation, resulting in systemic rescue from a global negative energy state and myocardial protection. Thus we demonstrate that hypoxia preconditions the innate immune response and determines survival outcomes following bacterial infection through suppression of HIF-1α and neutrophil metabolism. The therapeutic implications of this work are that in the context of systemic or tissue hypoxia therapies that target the host response could improve infection associated morbidity and mortality.
RESUMO
Neutrophil apoptosis leads to macrophage ingestion of intact senescent neutrophils. This may represent a neutrophil removal mechanism that is important both in the control of inflammatory tissue injury and for the normal resolution processes of inflammation. Because apoptosis is likely to be a key control process in cell and tissue homeostasis, a number of inflammatory mediators were tested for their ability to modulate the rate of apoptosis in populations of neutrophils aging in culture. Endotoxic lipopolysaccharide, human recombinant complement factor 5a, and human recombinant granulocyte-macrophage colony-stimulating factor all markedly inhibited the rate of neutrophil apoptosis in a concentration-dependent fashion, without inducing necrosis (as assessed by trypan blue exclusion). This inhibitory effect on the rate of neutrophil apoptosis was shown by morphological criteria and confirmed by gel electrophoresis of extracted DNA. Inhibition of apoptosis of aging neutrophil populations was associated with prolongation of the functional life span of the population as assessed by the ability of neutrophils to spread on glass surfaces, to polarize in response to deliberate stimulation with N-formyl-Met-Leu-Phe (fMLP), and to release the granule enzyme marker myeloperoxidase on fMLP stimulation. These observations show that inflammatory mediators prolong the functional life span of neutrophils through modulation of apoptosis. Further elucidation of these mechanisms will lead to a better understanding of the processes controlling neutrophil residence and function in inflamed tissues and may provide further insights into the molecular mechanisms of apoptosis, which is of widespread importance in tissue biology.
Assuntos
Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Lipopolissacarídeos/farmacologia , N-Formilmetionina Leucil-Fenilalanina/farmacologia , Neutrófilos/citologia , Neutrófilos/fisiologia , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Cromatina/efeitos dos fármacos , Cromatina/ultraestrutura , Complemento C5a/farmacologia , Dano ao DNA , Granulócitos/citologia , Humanos , Inflamação , Neutrófilos/efeitos dos fármacos , Peroxidase/sangue , Proteínas Recombinantes/farmacologiaRESUMO
Onset of apoptosis in many cell types, including the neutrophil granulocyte, leads to recognition and ingestion by macrophages, a key regulatory step in clearance of inflammatory cells from inflamed sites. These studies examined the requirement for protein synthesis in neutrophil apoptosis and in the recognition of apoptotic neutrophils by monocyte-derived macrophages. Treatment with cycloheximide or actinomycin D produced a time- and concentration-dependent acceleration of apoptosis in populations of neutrophils purified from human peripheral blood. Both compounds caused significant promotion of apoptosis after 8 h (apoptosis was 7.7 +/- 2.9%, mean +/- SEM, in control populations, 57.5 +/- 4.9% in cycloheximide-treated, and 73.4 +/- 5.5% in actinomycin D-treated populations, n = 4, P < 0.001), which was associated with loss of neutrophil functional ability (assessed by shape change on N-formyl-methionyl-leucyl-phenylalanine stimulation) and increased macrophage recognition and ingestion of neutrophil populations with accelerated apoptosis. These results support the existence of survival proteins, which act as intracellular suppressors of programmed cell death. However, protein synthesis was not required for the recognition process because macrophage recognition was increased pari passu with the morphology of apoptosis.
Assuntos
Apoptose , Macrófagos/fisiologia , Neutrófilos/citologia , Inibidores da Síntese de Proteínas/farmacologia , Apoptose/efeitos dos fármacos , Tamanho Celular , Sobrevivência Celular , Senescência Celular , Cicloeximida/farmacologia , Fragmentação do DNA/efeitos dos fármacos , Dactinomicina/farmacologia , Humanos , Biossíntese de Proteínas/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacosRESUMO
Apoptosis and clearance of neutrophils is essential for successful resolution of inflammation. Altered signaling via the Fas receptor could explain the observed prolongation of neutrophil lifespan and associated tissue injury at inflammatory sites. We therefore compared inflammatory neutrophils extracted from joints of rheumatoid arthritis patients, with peripheral blood neutrophils. Inflammatory neutrophils underwent constitutive apoptosis in culture more rapidly than peripheral blood neutrophils; this was not explained by changes in surface expression of Fas or by induction of Fas ligand. Inflammatory neutrophils remained sensitive to Fas-induced death, at levels comparable to those seen in peripheral blood neutrophils. Similarly, granulocyte-macrophage colony-stimulating factor reduced apoptosis but did not abolish signaling via Fas. These data provide evidence for the rate of apoptosis in inflammatory neutrophils being continually modulated by death and survival signals in the inflammatory milieu. This allows for rapid resolution of inflammation as levels of survival factors fall, and suggests new strategies for inducing resolution of inflammation.
Assuntos
Apoptose/fisiologia , Artrite Reumatoide/fisiopatologia , Glicoproteínas de Membrana/fisiologia , Neutrófilos/citologia , Neutrófilos/fisiologia , Receptor fas/fisiologia , Artrite Reumatoide/imunologia , Artrite Reumatoide/patologia , Sobrevivência Celular , Proteína Ligante Fas , Humanos , Técnicas In Vitro , Inflamação , Células K562 , Glicoproteínas de Membrana/sangue , Metaloendopeptidases/antagonistas & inibidores , Neutrófilos/imunologia , Inibidores de Proteases/farmacologia , Valores de Referência , Transdução de Sinais , Receptor fas/sangueRESUMO
The majority of patients with intrapulmonary right-to-left shunting due to pulmonary arteriovenous malformations-exhibit good maximum exercise capacity (> 70% predicted) despite profound arterial oxygen desaturation. We studied seven such patients to assess tissue oxygen delivery during steady-state exercise. From rest to exercise [50 +/- 7 (SE) W] arterial saturation fell from 80 +/- 3 to 74 +/- 3%, and mean right-to-left shunt increased slightly from 31 +/- 4 to 34 +/- 5% (P = NS). Minute ventilation was high for oxygen uptake, and the ventilatory equivalent was raised (174 +/- 19% predicted) and was correlated with shunt size (r = 0.93). The majority of the patients maintained pulmonary alveolar blood flow within the predicted range for their power output, but total cardiac output was increased to 142 +/- 11% predicted due to flow through the shunt. Consequently, on exercise, oxygen delivery per unit oxygen consumption [2.3-3.3 (normal range 1.6-2.4)] and calculated mixed venous oxygen tension (27.0 +/- 0.8 Torr) were preserved. Arterial PCO2 rose on exercise by 2.8 +/- 1.2 Torr, in proportion to the ratio of flow through the shunt to total cardiac output (r = 0.73), but remained low (33.1 +/- 1.4 Torr) in absolute terms. The high cardiac output on exercise may be facilitated by a low pulmonary vascular resistance (0.33 +/- 0.08 mmHg.1-1.min, measured at rest), which may explain why exercise performance is better in these patients than in patients with equivalent hypoxemia from other causes.
Assuntos
Malformações Arteriovenosas/fisiopatologia , Exercício Físico/fisiologia , Coração/fisiopatologia , Pulmão/fisiopatologia , Circulação Pulmonar/fisiologia , Adolescente , Adulto , Dióxido de Carbono/sangue , Débito Cardíaco/fisiologia , Feminino , Hemodinâmica/fisiologia , Humanos , Masculino , Pessoa de Meia-Idade , Consumo de Oxigênio/fisiologia , Troca Gasosa Pulmonar/fisiologia , Mecânica Respiratória/fisiologia , Resistência Vascular/fisiologiaRESUMO
Bronchial hyperresponsiveness is widely recognized as a marker of airway inflammation in asthma. The degree of bronchial hyperresponsiveness following acute severe attacks of asthma and the time course of its recovery has not previously been studied. Bronchial responsiveness to histamine was measured in 18 unselected patients admitted to hospital because of acute severe asthma, during their acute admission, and geometric mean PD20 histamine was 0.08 (range 0.02-0.32) mumol. In nine patients, further measurements were performed at 3-4 and 12 weeks following discharge. Geometric mean PD20 histamine was 0.09 mumol acutely, 0.23 mumol at 3-4 weeks (n = 9, p = 0.05 by analysis of variance) and 0.59 mumol at 12 weeks (n = 8, P = 0.04). For the eight patients studied at 12 weeks, a mean 10.3-fold increase in PD20 was shown, with no suggestion of a maximum effect having been achieved. In contrast, spirometry had returned to the normal range by 4 weeks. The dissociation between improvement in bronchial hyperresponsiveness and spirometry is of interest. The delayed reduction in hyperresponsiveness may have important clinical implications for the duration of anti-inflammatory corticosteroid treatment following acute severe asthma.
Assuntos
Asma/fisiopatologia , Brônquios/fisiopatologia , Hiper-Reatividade Brônquica/fisiopatologia , Doença Aguda , Adolescente , Adulto , Idoso , Asma/tratamento farmacológico , Testes de Provocação Brônquica , Quimioterapia Combinada , Feminino , Histamina , Humanos , Hidrocortisona/uso terapêutico , Masculino , Pessoa de Meia-Idade , Prednisolona/uso terapêutico , Espirometria , Fatores de TempoRESUMO
Involvement of the nervous system is an uncommon and underdiagnosed complication of sarcoidosis. We used gallium single photon emission computed tomography (SPECT) to define areas of meningeal involvement in a patient with neurosarcoidosis. This technique may be of value in the assessment of patients with sarcoidosis and suspected central nervous system involvement.
Assuntos
Encefalopatias/diagnóstico por imagem , Sarcoidose/diagnóstico por imagem , Adulto , Isótopos de Gálio , Humanos , Imageamento por Ressonância Magnética , Masculino , Tomografia Computadorizada de Emissão de Fóton ÚnicoAssuntos
Doenças Pulmonares Intersticiais/terapia , Pulmão/patologia , Biópsia/métodos , Lavagem Broncoalveolar , Teste de Esforço , Humanos , Hipertensão Pulmonar/etiologia , Hipertensão Pulmonar/terapia , Doenças Pulmonares Intersticiais/diagnóstico , Transplante de Pulmão , Anamnese , Exame Físico , Testes de Função Respiratória , Insuficiência Respiratória/etiologia , Insuficiência Respiratória/terapia , Medicamentos para o Sistema Respiratório/uso terapêutico , Terminologia como AssuntoRESUMO
The zebrafish is an outstanding model for intravital imaging of inflammation due to its optical clarity and the ability to express fluorescently labelled specific cell types by transgenesis. However, although several transgenic labelling myeloid cells exist, none allow distinction of macrophages from neutrophils. This prevents simultaneous imaging and examination of the individual contributions of these important leukocyte subtypes during inflammation. We therefore used Bacterial Artificial Chromosome (BAC) recombineering to generate a transgenic Tg(fms:GAL4.VP16)i186 , in which expression of the hybrid transcription factor Gal4-VP16 is driven by the fms (CSF1R) promoter. This was then crossed to a second transgenic expressing a mCherry-nitroreductase fusion protein under the control of the Gal4 binding site (the UAS promoter), allowing intravital imaging of mCherry-labelled macrophages. Further crossing this compound transgenic with the neutrophil transgenic Tg(mpx:GFP)i114 allowed clear distinction between macrophages and neutrophils and simultaneous imaging of their recruitment and behaviour during inflammation. Compared with neutrophils, macrophages migrate significantly more slowly to an inflammatory stimulus. Neutrophil number at a site of tissue injury peaked around 6 hours post injury before resolving, while macrophage recruitment increased until at least 48 hours. We show that macrophages were effectively ablated by addition of the prodrug metronidazole, with no effect on neutrophil number. Crossing with Tg(Fli1:GFP)y1 transgenic fish enabled intravital imaging of macrophage interaction with endothelium for the first time, revealing that endothelial contact is associated with faster macrophage migration. Tg(fms:GAL4.VP16)i186 thus provides a powerful tool for intravital imaging and functional manipulation of macrophage behaviour during inflammation.
Assuntos
Macrófagos/patologia , Neutrófilos/patologia , Especificidade de Órgãos , Animais , Animais Geneticamente Modificados , Movimento Celular/efeitos dos fármacos , Movimento Celular/imunologia , Técnicas de Cultura Embrionária , Endotélio Vascular/patologia , Engenharia Genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Inflamação , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Metronidazol/farmacologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/metabolismo , Especificidade de Órgãos/genética , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Transativadores/genética , Transativadores/metabolismo , Peixe-ZebraAssuntos
Imunodeficiência de Variável Comum/complicações , Uveíte/etiologia , Adulto , Imunodeficiência de Variável Comum/tratamento farmacológico , Feminino , Humanos , Imunoglobulinas/uso terapêutico , Trombocitopenia/tratamento farmacológico , Trombocitopenia/etiologia , Uveíte/tratamento farmacológicoRESUMO
The Toll-like receptor family was originally identified in Drosophila, where it provides important developmental and immunological signalling. In mammals, the developmental signal appears to have been lost, but the immunological defence role of these receptors has been expanded to provide broad recognition of bacterial, fungal, viral and parasitic pathogens. There is increasing evidence that these receptors go beyond the recognition of microbial molecules to sense host tissue damage. Recognition of host molecules and commensal microbes is also involved in the restoration of normal tissue architecture after injury and in maintenance of epithelial health. Recent developments in the TLR field highlight the importance of these molecules to human health and disease and demonstrate that their targeting, to boost immunity or inhibit inflammation, is both feasible and also potentially challenging.
Assuntos
Inflamação/imunologia , Receptores Toll-Like/imunologia , Animais , Doenças Transmissíveis/imunologia , Humanos , Ligantes , Transdução de Sinais/imunologia , Especificidade da EspécieRESUMO
The neutrophil is a crucial early defence against microbial infection, but neutrophilic inflammation can result in devastating acute and chronic inflammatory diseases. In the lungs, the neutrophil is a principal part of the pathology of the acute respiratory distress syndrome, and its activation may also be of substantial importance in chronic obstructive pulmonary disease and some forms of asthma. Induction of neutrophil recruitment in response to microbial attack requires activation of TLR (Toll-like receptor)-based signalling pathways and the concerted actions of multiple cell types, including sentinel cells such as monocytes and macrophages acting together with tissue cell types such as the epithelium or smooth-muscle cell. The present review describes some of these networks and the resulting potential for their targeting in respiratory disease.
Assuntos
Infiltração de Neutrófilos/imunologia , Neutrófilos/imunologia , Infecções Respiratórias/imunologia , Receptores Toll-Like/imunologia , Animais , Humanos , Inflamação , Transdução de Sinais/imunologiaRESUMO
BACKGROUND: The relationship between asthma severity and atopy is complex. Many studies have failed to show significant relationships between clinical severity or lung function and markers of atopic sensitisation. AIM: To determine whether increasing asthma severity is related to atopic sensitisation in a population of children with asthma. METHODS: A total of 400 children (7-18 years) with asthma were recruited as part of a multicentre study of the genetics of asthma. Detailed phenotypic data were collected on all participants. Associations between measures of asthma severity and atopic sensitisation were sought using multilevel models allowing variation at the individual and family level. RESULTS: Children recruited to the study had a range of asthma severities, with just over a third having mild persistent asthma. The logarithm of total serum IgE was associated with increased asthma severity score, decreased FEV1, increased airways obstruction, risk of hospital admission, and inhaled steroid use. Increasing skin prick test reactivity to a panel of seven aeroallergens was associated with increased risk of hospital admission, use of an inhaled steroid, and airways obstruction. The results remained highly significant after corrections for age, gender, and birth order. CONCLUSIONS: In children with asthma, increasing atopy is associated with increasing asthma severity. However, the relationships between asthma severity and skin prick tests, and asthma severity and total serum IgE values, appear subtly different.