RESUMEN

Although human exposure to Gram-negative Vibrio vulnificus (V. vulnificus) lipopolysaccharide (LPS) has been reported to result in septic shock, its impact on the central nervous system's innate immunity remains undetermined. The purpose of this study was to determine whether V. vulnificus MO6-24/O LPS might activate rat microglia in vitro and stimulate the release of superoxide anion (O2⁻), a reactive oxygen species known to cause oxidative stress and neuronal injury in vivo. Brain microglia were isolated from neonatal rats, and then treated with either V. vulnificus MO6-24/O LPS or Escherichia coli O26:B6 LPS for 17 hours in vitro. O2⁻ was determined by cytochrome C reduction, and matrix metalloproteinase-2 (MMP-2) and MMP-9 by gelatinase zymography. Generation of cytokines tumor necrosis factor alpha (TNF-α), interleukin-1 alpha (IL-1α), IL-6, and transforming growth factor-beta 1 (TGF-ß1), chemokines macrophage inflammatory protein (MIP-1α)/chemokine (C-C motif) ligand 3 (CCL3), MIP-2/chemokine (C-X-C motif) ligand 2 (CXCL2), monocyte chemotactic protein-1 (MCP-1)/CCL2, and cytokine-induced neutrophil chemoattractant-2alpha/beta (CINC-2α/ß)/CXCL3, and brain-derived neurotrophic factor (BDNF), were determined by specific immunoassays. Priming of rat microglia by V. vulnificus MO6-24/O LPS in vitro yielded a bell-shaped dose-response curve for PMA (phorbol 12-myristate 13-acetate)-stimulated O2⁻ generation: (1) 0.1-1 ng/mL V. vulnificus LPS enhanced O2⁻ generation significantly but with limited inflammatory mediator generation; (2) 10-100 ng/mL V. vulnificus LPS maximized O2⁻ generation with concomitant release of thromboxane B2 (TXB2), matrix metalloproteinase-9 (MMP-9), and several cytokines and chemokines; (3) 1000-100,000 ng/mL V. vulnificus LPS, with the exception of TXB2, yielded both attenuated O2⁻ production, and a progressive decrease in MMP-9, cytokines and chemokines investigated. Thus concentration-dependent treatment of neonatal brain microglia with V. vulnificus MO6-24/O LPS resulted in a significant rise in O2⁻ production, followed by a progressive decrease in O2⁻ release, with concomitant release of lactic dehydrogenase (LDH), and generation of TXB2, MMP-9, cytokines and chemokines. We hypothesize that the inflammatory mediators investigated may be cytotoxic to microglia in vitro, by an as yet undetermined autocrine mechanism. Although V. vulnificus LPS was less potent than E. coli LPS in vitro, inflammatory mediator release by the former was clearly more efficacious. Finally, we hypothesize that should V. vulnificus LPS gain entry into the CNS, it would be possible that microglia might become activated, resulting in high levels of O2⁻ as well as neuroinflammatory TXB2, MMP-9, cytokines and chemokines.

Asunto(s)

Escherichia coli/patogenicidad , Lipopolisacáridos/administración & dosificación , Microglía/metabolismo , Vibrio vulnificus/patogenicidad , Animales , Animales Recién Nacidos , Encéfalo/inmunología , Encéfalo/metabolismo , Encéfalo/microbiología , Quimiocinas/metabolismo , Citocinas/metabolismo , Inmunidad Innata , Lipopolisacáridos/aislamiento & purificación , Metaloproteinasa 9 de la Matriz/efectos de los fármacos , Metaloproteinasa 9 de la Matriz/metabolismo , Microglía/inmunología , Microglía/microbiología , Ratas , Ratas Sprague-Dawley , Superóxidos/metabolismo , Tromboxano B2/metabolismo

RESUMEN

The epidemiological association between exposure to fine particulate matter (PM2.5) and adverse health effects is well-known. Here we report the size distribution, metals content, endotoxin content, and biological activity of National Institute of Standards and Technology (NIST) Interim Reference Material (RM) PM2.5. Biological activity was measured in vitro by effects on cell viability and the release of four inflammatory immune mediators, from human A549 alveolar epithelial cells or murine RAW264.7 monocytes. A dose range covering three orders of magnitude (1-1000µg/mL) was tested, and biological activity was compared to an existing Standard Reference Material (SRM) for urban PM (NIST SRM 1648). Robust release of IL-8 and MCP-1 from A549 cells was observed in response to IRM PM2.5 exposures. Significant TNF-α, but not IL-6, secretion from RAW264.7 cells was observed in response to both IRM PM2.5 and SRM 1648 particle types. Cytokine or chemokine release at high doses often occurred in the presence of cytotoxicity, likely as a result of externalization of preformed mediator. Our results are consistent with a local cytotoxic and pro-inflammatory mechanism of response to exposure to inhaled ambient PM2.5 and reinforce the continued relevance of in vitro assays for mechanistic research in PM toxicology. Our study furthers the goal of developing reference samples of environmentally relevant particulate matter of various sizes that can be used for hypothesis testing by multiple investigators.

Asunto(s)

Contaminantes Atmosféricos/normas , Material Particulado/normas , Contaminantes Atmosféricos/química , Contaminantes Atmosféricos/toxicidad , Animales , Línea Celular , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Citocinas/metabolismo , Endotoxinas/análisis , Humanos , Metales/análisis , Ratones , Tamaño de la Partícula , Material Particulado/química , Material Particulado/toxicidad , Estándares de Referencia

RESUMEN

A major challenge for the treatment of many central nervous system (CNS) disorders is the lack of convenient and effective methods for delivering biological agents to the brain. Mucopolysaccharidosis II (Hunter syndrome) is a rare inherited lysosomal storage disorder resulting from a deficiency of iduronate-2-sulfatase (I2S). I2S is a large, highly glycosylated enzyme. Intravenous administration is not likely to be an effective therapy for disease-related neurological outcomes that require enzyme access to the brain cells, in particular neurons and oligodendrocytes. We demonstrate that intracerebroventricular and lumbar intrathecal administration of recombinant I2S in dogs and nonhuman primates resulted in widespread enzyme distribution in the brain parenchyma, including remarkable deposition in the lysosomes of both neurons and oligodendrocytes. Lumbar intrathecal administration also resulted in enzyme delivery to the spinal cord, whereas little enzyme was detected there after intraventricular administration. Mucopolysaccharidosis II model is available in mice. Lumbar administration of recombinant I2S to enzyme deficient animals reduced the storage of glycosaminoglycans in both superficial and deep brain tissues, with concurrent morphological improvements. The observed patterns of enzyme transport from cerebrospinal fluid to the CNS tissues and the resultant biological activity (a) warrant further investigation of intrathecal delivery of I2S via lumbar catheter as an experimental treatment for the neurological symptoms of Hunter syndrome and (b) may have broader implications for CNS treatment with biopharmaceuticals.

Asunto(s)

Sistema Nervioso Central/efectos de los fármacos , Terapia de Reemplazo Enzimático/métodos , Iduronato Sulfatasa/uso terapéutico , Mucopolisacaridosis II/tratamiento farmacológico , Animales , Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/patología , Perros , Humanos , Iduronato Sulfatasa/administración & dosificación , Iduronato Sulfatasa/genética , Inmunohistoquímica , Inyecciones Espinales , Radioisótopos de Yodo , Lisosomas/metabolismo , Macaca fascicularis , Ratones , Ratones Noqueados , Mucopolisacaridosis II/genética , Mucopolisacaridosis II/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Oligodendroglía/efectos de los fármacos , Oligodendroglía/metabolismo , Oligodendroglía/patología , Tomografía de Emisión de Positrones , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/farmacocinética , Proteínas Recombinantes/uso terapéutico , Especificidad de la Especie , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Médula Espinal/patología , Distribución Tisular

RESUMEN

BACKGROUND: Complement C2 deficiency is the most common genetically determined complete complement deficiency and is associated with a number of diseases. Most prominent are the associations with recurrent serious infections in young children and the development of systemic lupus erythematosus (SLE) in adults. The links with these diseases reflect the important role complement C2 plays in both innate immunity and immune tolerance. Infusions with normal fresh frozen plasma for the treatment of associated disease have demonstrated therapeutic effects but so far protein replacement therapy has not been evaluated. RESULTS: Human complement C2 was cloned and expressed in a mammalian cell line. The purity of recombinant human C2 (rhC2) was greater than 95% and it was characterized for stability and activity. It was sensitive to C1s cleavage and restored classical complement pathway activity in C2-deficient serum both in a complement activation ELISA and a hemolytic assay. Furthermore, rhC2 could increase C3 fragment deposition on the human pathogen Streptococcus pneumoniae in C2-deficient serum to levels equal to those with normal serum. CONCLUSIONS: Taken together these data suggest that recombinant human C2 can restore classical complement pathway activity and may serve as a potential therapeutic for recurring bacterial infections or SLE in C2-deficient patients.

Asunto(s)

Complemento C2/metabolismo , Síndromes de Inmunodeficiencia/genética , Lupus Eritematoso Sistémico/genética , Proteínas Recombinantes/metabolismo , Infecciones Estreptocócicas/genética , Streptococcus pneumoniae/inmunología , Adulto , Línea Celular Transformada , Niño , Complemento C1/inmunología , Complemento C1/metabolismo , Complemento C2/genética , Complemento C2/uso terapéutico , Complemento C3/inmunología , Complemento C3/metabolismo , Vía Clásica del Complemento/efectos de los fármacos , Humanos , Síndromes de Inmunodeficiencia/complicaciones , Síndromes de Inmunodeficiencia/tratamiento farmacológico , Lupus Eritematoso Sistémico/complicaciones , Lupus Eritematoso Sistémico/tratamiento farmacológico , Unión Proteica/efectos de los fármacos , Proteínas Recombinantes/genética , Proteínas Recombinantes/uso terapéutico , Recurrencia , Infecciones Estreptocócicas/complicaciones , Infecciones Estreptocócicas/tratamiento farmacológico

RESUMEN

SHG2210, a fusion protein containing the N-terminus of human nicotinic acetylcholine receptor α (AchR-α; aa1-210) and human transferrin (TF), was characterized as a potential therapeutic for myasthenia gravis (MG) caused predominately by α subunit autoantibodies. SHG2210 was shown to be able to bind to α subunit autoantibodies and the TF receptor (TFR). SHG2210 and SHG2210-anti-AchR antibody complex are internalized through TFR-mediated endocytosis. The SHG2210 and SHG2210-anti-AchR antibody complex is present in Lamp1-positive lysosomal compartments after internalization; however, neither SHG2210 nor SHG2210-antibody complex is present in Rab11-positive recycling endosomes. SHG2210 bound to α subunit of AChR autoantibodies may be cleared by the lysosome, resulting in short cellular half-life relative to SHG2210. SHG2210 is shown to have a protective effect on antigenic modulation of the AChR induced by serum from select patients with MG, suggesting that a fusion protein approach may be an effective therapeutic for treating MG.

Asunto(s)

Miastenia Gravis/inmunología , Receptores Nicotínicos/inmunología , Receptores de Transferrina/inmunología , Proteínas Recombinantes/farmacología , Transferrina/inmunología , Unión Competitiva/inmunología , Células HeLa , Humanos , Microscopía Confocal , Miastenia Gravis/tratamiento farmacológico , Receptores Nicotínicos/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/uso terapéutico , Transferrina/genética

RESUMEN

Induction of proinflammatory mediators by alveolar macrophages exposed to ambient air particulate matter has been suggested to be a key factor in the pathogenesis of inflammatory and allergic diseases in the lungs. However, receptors and mechanisms underlying these responses have not been fully elucidated. In this study, we examined whether TLR2, TLR4, and the key adaptor protein, MyD88, mediate the expression of proinflammatory cytokines and chemokines by mouse peritoneal macrophages exposed to fine and coarse PM. TLR2 deficiency blunted macrophage TNF-alpha and IL-6 expression in response to fine (PM2.5), while not affecting cytokine-inducing ability of coarse NIST Standard Reference Material (SRM 1648) particles. In contrast, TLR4(-/-) macrophages showed inhibited cytokine expression upon stimulation with NIST SRM 1648 but exhibited normal responses to PM2.5. Preincubation with polymyxin B markedly suppressed the capacity of NIST SRM 1648 to elicit TNF-alpha and IL-6, indicating endotoxin as a principal inducer of cytokine responses. Overexpression of TLR2 in TLR2/4-deficient human embryonic kidney 293 cells imparted PM2.5 sensitivity, as judged by IL-8 gene expression, whereas NIST SRM 1648, but not PM2.5 elicited IL-8 expression in 293/TLR4/MD-2 transfectants. Engagement of TLR4 by NIST SRM 1648 induced MyD88-independent expression of the chemokine RANTES, while TLR2-reactive NIST IRM PM2.5 failed to up-regulate this response. Consistent with the shared use of MyD88 by TLR2 and TLR4, cytokine responses of MyD88(-/-) macrophages to both types of air PM were significantly reduced. These data indicate differential utilization of TLR2 and TLR4 but shared use of MyD88 by fine and coarse air pollution particles.

Asunto(s)

Macrófagos/inmunología , Factor 88 de Diferenciación Mieloide/metabolismo , Material Particulado/efectos adversos , Neumonía/inmunología , Receptor Toll-Like 2/metabolismo , Receptor Toll-Like 4/metabolismo , Animales , Antibacterianos/farmacología , Línea Celular , Células Cultivadas , Quimiocina CCL5/metabolismo , Humanos , Hipersensibilidad/inmunología , Hipersensibilidad/fisiopatología , Mediadores de Inflamación/metabolismo , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Ratones , Ratones Noqueados , Factor 88 de Diferenciación Mieloide/genética , Neumonía/genética , Neumonía/fisiopatología , Polimixina B/farmacología , Transducción de Señal/inmunología , Receptor Toll-Like 2/genética , Receptor Toll-Like 4/genética , Factor de Necrosis Tumoral alfa/metabolismo

RESUMEN

BACKGROUND: Myasthenia gravis (MG) is an autoimmune disease affecting approximately 40,000 patients in the United States. One of the major mechanisms of disease pathology in MG is the binding, internalization, and eventual destruction of acetylcholine receptors (AChR) at the neuromuscular junction by cross-linking AChR-specific autoantibodies. This process, known as antigenic modulation, ultimately attenuates the ability of muscle cells to contract in response to signals from neurons, leading to muscle weakness and fatigue. For this reason, antigenic modulation of the AChR on cultured cells has become an important diagnostic tool for assessing the pathogenicity of AChR-specific autoantibodies. Traditionally, these assays have been done using radiolabeled AChR ligands such as (125)I alpha-bungarotoxin to determine relative AChR number. Here, we present a high-throughput immunofluorescent flow cytometry-based assay that can be used to quantify AChR levels on the cell surface and assess the efficacy of molecules designed to rescue antigenic modulation. METHODS: AChR levels were quantified on human muscle cells before and after treatment with AChR antibodies via immunofluorescent labeling with the AChR monoclonal antibodies, mAb210 and mAb B3, followed by flow cytometry of EDTA-treated cells. RESULTS: Using a novel, flow cytometry-based assay, antigenic modulation of the AChR was demonstrated on human cells using both AChR-specific monoclonal antibody and MG patient serum. The degree of antigenic modulation was dose responsive to antibody levels and could be reversed by preincubating antibodies with soluble AChR alpha subunit extracellular domain. SUMMARY: A rapid, nonradioactive assay was developed to determine the potential of AChR-specific antibodies in the serum of MG patients to bind and down-regulate the AChR. This assay can be used to assess the ability of putative therapeutics that rescue antigenic modulation and could be developed for the treatment of MG.

Asunto(s)

Anticuerpos Monoclonales/inmunología , Autoanticuerpos/inmunología , Regulación hacia Abajo , Citometría de Flujo/métodos , Fluorescencia , Receptores Colinérgicos/análisis , Receptores Colinérgicos/inmunología , Anticuerpos Monoclonales/farmacología , Anticuerpos Monoclonales/uso terapéutico , Reacciones Antígeno-Anticuerpo/efectos de los fármacos , Reacciones Antígeno-Anticuerpo/inmunología , Línea Celular , Regulación hacia Abajo/efectos de los fármacos , Humanos , Miastenia Gravis/inmunología , Miastenia Gravis/metabolismo , Sensibilidad y Especificidad , Factores de Tiempo

Asunto(s)

Sistemas de Registros Médicos Computarizados , Planes de Incentivos para los Médicos/normas , Indicadores de Calidad de la Atención de Salud/normas , Mecanismo de Reembolso , Centers for Medicare and Medicaid Services, U.S. , Humanos , Planes de Incentivos para los Médicos/economía , Estados Unidos

RESUMEN

BACKGROUND: Recent studies indicate that the composition of fine particulate matter [PM

Asunto(s)

Contaminantes Atmosféricos/análisis , Asma/epidemiología , Zinc/análisis , Adolescente , Niño , Preescolar , Femenino , Geografía , Humanos , Lactante , Recién Nacido , Masculino , Maryland/epidemiología , Modelos Teóricos , Salud Urbana/estadística & datos numéricos

RESUMEN

Cryptosporidiosis in young children prompts local inflammation in the intestinal tract. We studied a cohort of young children with cryptosporidiosis to determine whether systemic inflammatory responses occur and, if so, to evaluate whether inflammation persists after infection. Cryptosporidiosis was associated with increased levels of interleukin-8 and tumor necrosis factor- alpha systemically, which persisted at 6 months after enrollment. The level of intestinal tumor necrosis factor- alpha was elevated at enrollment, but elevated levels did not persist. Worsening of malnutrition, particularly stunting, was observed after infection. The association of cryptosporidiosis, inflammation, and stunting in children with cryptosporidiosis warrants further evaluation.

Asunto(s)

Criptosporidiosis/metabolismo , Interferón gamma/metabolismo , Interleucinas/metabolismo , Receptores del Factor de Necrosis Tumoral/metabolismo , Antígenos de Protozoos/sangre , Estudios de Cohortes , Femenino , Humanos , Lactante , Interferón gamma/sangre , Masculino , Receptores del Factor de Necrosis Tumoral/sangre

RESUMEN

Enterotoxigenic Bacteroides fragilis (ETBF) secretes a 20-kDa metalloprotease toxin termed B. fragilis toxin (BFT). ETBF disease in animals is associated with an acute inflammatory response in the intestinal mucosa, and lethal hemorrhagic colitis may occur in rabbits. In this study, we confirmed recent reports (J. M. Kim, Y. K. Oh, Y. J. Kim, H. B. Oh, and Y. J. Cho, Clin. Exp. Immunol. 123:421-427, 2001; L. Sanfilippo, C. K. Li, R. Seth, T. J. Balwin, M. J. Menozzi, and Y. R. Mahida, Clin. Exp. Immunol. 119:456-463, 2000) that purified BFT stimulates interleukin-8 (IL-8) secretion by human intestinal epithelial cells (HT29/C1 cells) and demonstrate that stimulation of IL-8 production is dependent on biologically active BFT and independent of serum. Induction of IL-8 mRNA expression occurs rapidly and ceases by 6 h after BFT treatment, whereas IL-8 secretion continues to increase for at least 18 h. Our data suggest that BFT-stimulated IL-8 secretion involves tyrosine kinase-dependent activation of nuclear factor-kappaB (NF-kappaB) as well as activation of the mitogen-activated protein kinases (MAPKs), p38 and extracellular signal-related kinase. Simultaneous activation of NF-kappaB and MAPKs appears necessary for secretion of IL-8 by HT29/C1 cells treated with BFT.

Asunto(s)

Interleucina-8/metabolismo , Mucosa Intestinal/efectos de los fármacos , Metaloendopeptidasas/farmacología , Proteínas Quinasas Activadas por Mitógenos/metabolismo , FN-kappa B/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos , Bacteroides fragilis , Línea Celular Tumoral , Activación Enzimática/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Células Epiteliales/enzimología , Células Epiteliales/metabolismo , Genisteína/farmacología , Humanos , Interleucina-8/biosíntesis , Mucosa Intestinal/enzimología , Mucosa Intestinal/metabolismo , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Suero , Factores de Tiempo , Proteínas Quinasas p38 Activadas por Mitógenos

RESUMEN

Vibrio vulnificus is the leading cause of death in the United States associated with the consumption of raw seafood, particularly oysters. In epidemiological studies, primary septicemia and inflammation-mediated septic shock caused by V. vulnificus is strongly associated with liver disease, often in the context of chronic alcohol abuse. The present study was undertaken to determine whether clinical biomarkers of liver function or cellular oxidative stress are associated with peripheral blood mononuclear cell inflammatory cytokine responses to V. vulnificus. Levels of interleukin-1 beta (IL-1 beta), IL-6, IL-8, and tumor necrosis factor alpha elicited in response to V. vulnificus and measured in cell supernatants were not associated with the liver biomarkers aspartate aminotransferase (AST) or alanine aminotransferase (ALT) or the AST/ALT ratio. In contrast, reduced glutathione (GSH) levels were associated with the release of all four cytokines (IL-1 beta [R(2) = 0.382; P = 0.006], IL-6 [R(2) = 0.393; P = 0.005], IL-8 [R(2) = 0.487; P = 0.001], and TNF-alpha [R(2) = 0.292; P = 0.021]). Those individuals with below-normal GSH levels produced significantly less proinflammatory cytokines in response to V. vulnificus. We hypothesize that persons with markers for cellular oxidative stress have increased susceptibility to V. vulnificus septicemia.

Asunto(s)

Alcoholismo/inmunología , Citocinas/biosíntesis , Leucocitos Mononucleares/inmunología , Hígado/fisiopatología , Estrés Oxidativo , Vibrio vulnificus/inmunología , Adulto , Alanina Transaminasa/sangre , Alcoholismo/fisiopatología , Aspartato Aminotransferasas/sangre , Biomarcadores , Glutatión/análisis , Humanos , Persona de Mediana Edad

RESUMEN

Molecular methods, including DNA probes, were used to identify and enumerate pathogenic Vibrio species in the Chesapeake Bay; our data indicated that Vibrio vulnificus exhibits seasonal fluctuations in number. Our work included a characterization of total microbial communities from the Bay; development of microarrays that identify and quantify the diversity of those communities; and observation of temporal changes in those communities. To identify members of the microbial community, we amplified the 16S rDNA gene from community DNA isolated from a biofilm sample collected from the Chesapeake Bay in February, 2000. The resultant 75 sequences were 95% or more similar to 7 species including two recently described Shewanella species, baltica and frigidimarina, that have not been previously isolated from the Chesapeake. When the genera of bacteria from biofilm after culturing are compared to those detected by subcloning amplified 16S fragments from community DNA, the cultured sample exhibited a strong bias. In oysters collected in February, the most common bacteria were previously unknown. Based on our 16S findings, we are developing microarrays to detect these and other microbial species in these estuarine communities. The microarrays will detect each species using four distinct loci, with the multiple loci serving as an internal control. The accuracy of the microarray will be measured using sentinel species such as Aeromonas species, Escherichia coli, and Vibrio vulnificus. Using microarrays, it should be possible to determine the annual fluctuations of bacterial species (culturable and non-culturable, pathogenic and non-pathogenic). The data may be applied to understanding patterns of environmental change; assessing the "health" of the Bay; and evaluating the risk of human illness associated with exposure to and ingestion of water and shellfish.

Asunto(s)

ADN Bacteriano/genética , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Ribosómico 16S/genética , Vibrio/genética , Animales , Monitoreo del Ambiente/métodos , Humanos , Maryland , Ostreidae/microbiología , Dinámica Poblacional , Salud Pública , Medición de Riesgo , Mariscos , Virginia , Microbiología del Agua

RESUMEN

Upon activation with various noncytokine stimuli, polymorphonuclear leukocytes (PMNs) mobilize intracellular sialidase to the plasma membrane, where the sialidase releases sialic acid from the cell surface. This desialylation enhances PMN adherence, spreading, deformability, and motility, functions critical to diapedesis. We now have examined the role of sialidase activity in PMN adhesion to and migration across the endothelium in vivo. A polyclonal antibody prepared against Clostridium perfringens neuraminidase 1) detected surface expression of sialidase on human PMNs stimulated with IL-8 in vitro and on murine PMNs stimulated in vivo, but not on that of unstimulated cells, 2) recognized proteins in human PMN lysates and granule preparations that were not detected by preimmune antibody, 3) inhibited bacterial neuraminidase and human PMN sialidase activities in vitro, and 4) inhibited both pulmonary leukostasis in mice systemically infused with cobra venom factor and intrapulmonary transendothelial migration of PMNs into the bronchoalveolar compartment of mice intranasally challenged with interleukin-8. We conclude that the chemokine interleukin-8, like other PMN agonists, induces the translocation of sialidase to the PMN surface and that surface expression of this sialidase is a prerequisite to PMN recruitment in vivo. The ability of antibodies raised against a prokaryotic neuraminidase to recognize eukaryotic sialidase extends the concept of the neuraminidase superfamily to mammalian enzymes. Inhibition of mobilized endogenous sialidase may provide a novel strategy for limiting the inflammatory response.

Asunto(s)

Neuraminidasa/metabolismo , Neutrófilos/citología , Animales , Western Blotting , Bronquios/citología , Bronquios/metabolismo , Calcimicina/farmacología , Clostridium perfringens/enzimología , Humanos , Inmunoglobulina G/inmunología , Interleucina-8/farmacología , Ratones , Modelos Animales , N-Formilmetionina Leucil-Fenilalanina/farmacología , Neuraminidasa/antagonistas & inhibidores , Neuraminidasa/inmunología , Neutrófilos/efectos de los fármacos , Neutrófilos/metabolismo , Conejos , Acetato de Tetradecanoilforbol/farmacología