RESUMEN
BACKGROUND: We previously demonstrated in mice that airway eosinophils traffic from the airway lumen into lung-draining paratracheal lymph nodes. However, mechanisms whereby eosinophils traverse from the lungs and home to paratracheal lymph nodes remain unclear. We investigated roles of cysteinyl leukotrienes in mediating eosinophil trafficking from lungs to paratracheal lymph nodes. METHODS: The expression of CCR7 was determined by flow cytometry. Transwell assays were used to test chemotactic responses of leukotriene C4 synthase-deficient and control airway eosinophils to the chemokine CCL19 ex vivo. Eosinophils from the spleens of IL-5 transgenic mice, fluorescently labeled ex vivo, were intratracheally injected into ovalbumin-sensitized and ovalbumin aerosol-challenged leukotriene C4 synthase-deficient and control mice. Eosinophils were identified by microscopy and flow cytometry in the lungs and paratracheal lymph nodes. RESULTS: Mouse eosinophils expressed CCR7, the receptor for CCL19, and responded chemotactically to CCL19. Leukotriene C4 synthase-deficient eosinophils exhibited impaired chemotaxis to CCL19 that was restored by exogenous leukotriene C4 . The migration of intratracheally injected eosinophils into paratracheal lymph nodes from distal alveolar lung was diminished in leukotriene C4 synthase-deficient mice compared with wild-type mice, with increased retention of eosinophils in the lungs of leukotriene C4 synthase-deficient mice. Exogenous administration of leukotriene C4 restored trafficking of eosinophils to paratracheal lymph nodes in leukotriene C4 synthase-deficient mice. CONCLUSIONS: Our findings that cysteinyl leukotrienes are involved in regulating airway and lung eosinophil migration into paratracheal lymph nodes identify previously unrecognized roles for the cysteinyl leukotrienes in regulating the pulmonary trafficking of eosinophils in experimental allergic asthma.
Asunto(s)
Quimiotaxis , Eosinófilos/citología , Leucotrieno C4/inmunología , Ganglios Linfáticos/citología , Animales , Asma/patología , Quimiocina CCL19/fisiología , Eosinófilos/metabolismo , Leucotrieno C4/administración & dosificación , Leucotrieno C4/deficiencia , Pulmón/citología , Ganglios Linfáticos/metabolismo , Ratones , Receptores CCR7/fisiologíaRESUMEN
BACKGROUND: The salvage laryngectomy (SLE) is very often the only curative option in recurrent laryngeal or hypopharyngeal carcinomas. But the SLE is associated with an increased risk of complications such as the formation of salivary fistulas. To reduce the rate of fistulas a simultaneous elevation of the myofascial pectoralis major flap (PMML) is described. The aim of this study was to compare the SLE with and without the use of the PMML for prophylaxis of salivary fistulas. PATIENTS AND METHOD: 9 patients were included, suffering from a T4a larynx or hypopharynx carcinoma recurrence after RCT in the years 2012 and 2013 and subsequently treated by a SLE. An additional elevation of PMML was indicated due to the following criteria: end of RCT less than one year ago, tumor localization outside the glottis, infiltration of thyroid cartilage and prelaryngeal muscles. After PMML elevation the flap was sewed onto a primary closed pharynx. RESULTS: 6 out of 9 patients (2/3) received an additional covering of the pharynx by the PMML during SLE. In no case a postoperative salivary fistula was seen. In the remaining 3 patients (1/3) the pharynx was primarily closed without an additional covering by the PMML. In this group of patients one postoperative salivary fistula was seen. CONCLUSION: Due to the simultaneous application of the PMML in the context of SLE the rate of postoperative salivary fistula could be effectively reduced in our own patients. The PMML is suitable due to its safe elevation technique, the missing secondary thoracal cutaneous defect, and a good modelling possibility in the recipient area.
Asunto(s)
Neoplasias Hipofaríngeas/cirugía , Neoplasias Laríngeas/cirugía , Laringectomía/métodos , Colgajo Miocutáneo/cirugía , Recurrencia Local de Neoplasia/cirugía , Complicaciones Posoperatorias/etiología , Terapia Recuperativa/métodos , Anciano , Fístula Cutánea/etiología , Fístula Cutánea/prevención & control , Fístula/etiología , Fístula/prevención & control , Humanos , Neoplasias Hipofaríngeas/patología , Neoplasias Laríngeas/patología , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/patología , Estadificación de Neoplasias , Enfermedades Faríngeas/etiología , Enfermedades Faríngeas/prevención & control , Complicaciones Posoperatorias/prevención & control , Fístula de las Glándulas Salivales/etiología , Fístula de las Glándulas Salivales/prevención & controlRESUMEN
UNLABELLED: Treatment of Recurrent Epistaxis by Artery Ligation: Up to Date or Old Fashioned? BACKGROUND: Despite the ongoing development in the field of endoscopic treatment techniques, recurrent epistaxis remains a challenge for otolaryngologists. The aim of the present study was to compare our own results of various interventions for the treatment of recurrent epistaxis. MATERIALS AND METHODS: From 2007 to 2013 we performed surgical treatment of recurrent epistaxis under general anaesthesia in 148 cases. While the majority of causes were idiopathic (n=98), epistaxis also occurred postoperatively (n=30), post-traumatically (n=7) or as a result of M. Osler (n=12). In 141/148 cases the treatment was performed by mono- or bipolar coagulation in the area of the bleeding source - this required an ethmoidectomy in 17 cases. In 19 cases the intervention was combined with a septoplasty. In 4 patients with recurrent bleeding of unknown origin, where electrocoagulation under general anaesthesia failed, we performed a clipping of the ethmoid- and/or the maxillary arteries in the pterygopalatine fossa. Following this intervention no further bleeding episodes occured. In further 3 patients, neuroradiological embolization was successfully performed. CONCLUSION: If conservative measures fail in the treatment of epistaxis, surgical treatment by electrocoagulation of the bleeding site under general anaesthesia is an effective intervention in 95% of cases. However for the remaining 5% where these measures have been proven to be ineffective, clipping of the ipsilateral anterior and posterior ethmoid- and/or the maxillar artery provides a treatment option being equally efficient as neuroradiological interventions.
Asunto(s)
Epistaxis/cirugía , Senos Etmoidales/irrigación sanguínea , Arteria Maxilar/cirugía , Adulto , Anciano , Arterias/cirugía , Electrocoagulación , Epistaxis/etiología , Femenino , Humanos , Ligadura , Masculino , Persona de Mediana Edad , Nariz , Recurrencia , Reoperación , Estudios RetrospectivosRESUMEN
BACKGROUND: Eosinophils have the capacity to secrete varied cytotoxic proteins. Among the proteins are the eosinophil-associated RNases (EARs): the human eosinophil-derived neurotoxin and eosinophilic cationic protein, and their murine ortholog EARs, which have been shown to be involved in host defense, tissue remodeling, and immunity regulation. However, the signal transduction that regulates EARs secretion in response to physiological stimuli, such as chemokines, has been little studied in human and scarcely in mouse eosinophils, the foremost animal model for eosinophil-associated human diseases. OBJECTIVE: In this study, we aimed to understand the signal transduction involved in the secretion of enzymatically active EARs following chemokine stimulation. METHODS: Fresh mouse and human eosinophils were stimulated with CCL11 and CCL24, and the secretion of enzymatically active EARs was detected using an RNase activity assay. The involvement of signaling factors or integrins was probed using specific inhibitors and blocking antibodies. Adhesion was evaluated by microscopy. RESULTS: We found that secretion of mouse EARs in response to CCL11 and CCL24 was Gαi -dependent. Both mouse and human eosinophils required the activation of PI3K, ERK, and p38 MAPK. In addition, the adhesion molecules ß1 and ß2 integrins were found to be crucial for EAR secretion, and we suggest a mechanism in which spreading is obligatory for EAR secretion. CONCLUSIONS: Collectively, these data suggest a common CCR3-mediated signaling pathway that leads to EAR secretion in both mouse and human eosinophils. These findings are applicable for eosinophil-mediated host defense and eosinophil-associated diseases.
Asunto(s)
Eosinófilos/inmunología , Quinasas MAP Reguladas por Señal Extracelular/inmunología , Integrinas/inmunología , Fosfatidilinositol 3-Quinasas/inmunología , Ribonucleasas/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/inmunología , Animales , Western Blotting , Células Cultivadas , Quimiocina CCL11/inmunología , Quimiocina CCL11/metabolismo , Eosinófilos/citología , Eosinófilos/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Citometría de Flujo , Humanos , Integrinas/metabolismo , Sistema de Señalización de MAP Quinasas , Ratones , Fosfatidilinositol 3-Quinasas/metabolismo , Receptores CCR3/inmunología , Receptores CCR3/metabolismo , Ribonucleasas/inmunología , Sensibilidad y Especificidad , Transducción de Señal , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Several lines of evidence suggest that deficiency of eosinophils is not associated with any characteristic abnormality. Patients lacking eosinophils, in the setting of immunodeficiency or as a consequence of IgG-mediated eosinophil precursor destruction, do not display any distinguishing abnormalities related to eosinophil reduction. The observation that eosinophil-deficient mice do not display any distinctive syndrome or failure of their health is evidence that, under ordinary laboratory conditions, the eosinophil does not play a critical role in the well-being of mammals. Observations that monoclonal antibodies to interleukin-5 (IL-5) are well tolerated appear unsurprising in light of these findings. For example, patients with the hypereosinophilic syndrome have received mepolizumab, an anti-IL-5 monoclonal antibody, for as long as 6 years and have not developed any characteristic set of adverse events. Safety data for reslizumab, another anti-IL-5 monoclonal antibody, and benralizumab, a monoclonal antibody to the IL-5 receptor α-chain, are comparatively limited, especially for benralizumab, although reports of administration of these antibodies to humans suggest that they are well tolerated. Thus, data to the present suggest that reduction of eosinophils appears to have no characteristic ill effects on normal health, and monoclonal antibodies that deplete eosinophils have the potential to be widely employed in the treatment of eosinophil-associated diseases.
Asunto(s)
Anticuerpos Monoclonales Humanizados/uso terapéutico , Eosinófilos/inmunología , Síndromes de Inmunodeficiencia/sangre , Síndromes de Inmunodeficiencia/inmunología , Timoma/inmunología , Neoplasias del Timo/inmunología , Animales , Anticuerpos Monoclonales Humanizados/efectos adversos , Estudios de Cohortes , Modelos Animales de Enfermedad , Eosinófilos/metabolismo , Femenino , Humanos , Síndromes de Inmunodeficiencia/tratamiento farmacológico , Incidencia , Interleucina-5/efectos adversos , Interleucina-5/inmunología , Interleucina-5/uso terapéutico , Recuento de Leucocitos , Masculino , Ratones , Pronóstico , Medición de Riesgo , Timoma/sangre , Timoma/epidemiología , Neoplasias del Timo/sangre , Neoplasias del Timo/epidemiologíaRESUMEN
The pseudoautosomal boundaries are the interface between pseudoautosomal and sex chromosome-specific DNA sequences. We have isolated a gene, PBDX, from the human pseudoautosomal boundary region of Xp. The three exons at the 5' end of PBDX are situated in the pseudoautosomal region immediately downstream of MIC2, whereas the other seven exons are in the X-specific region. Hence, PBDX is inherited in two modes: its 5' end is pseudoautosomally inherited and its 3' end is X-linked. The predicted amino acid sequence of the 540 bp coding region is 48% homologous to 12E7, the product of MIC2. By virtue of its position, PBDX becomes an excellent candidate for the XG blood group gene.
Asunto(s)
Antígenos CD , Antígenos de Grupos Sanguíneos/genética , Moléculas de Adhesión Celular/genética , Genes , Glicoproteínas de Membrana/genética , Cromosoma X , Antígeno 12E7 , Secuencia de Aminoácidos , Secuencia de Bases , Mapeo Cromosómico , Intercambio Genético , ADN Complementario/genética , Compensación de Dosificación (Genética) , Exones , Femenino , Regulación de la Expresión Génica , Humanos , Masculino , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Recombinación Genética , Alineación de Secuencia , Homología de Secuencia de AminoácidoRESUMEN
We have identified the Xga antigen, encoded by the XG blood group gene, by employing rabbit polyclonal and mouse monoclonal antibodies raised against a peptide derived from the N-terminal domain of a candidate gene, referred to earlier as PBDX. In indirect haemagglutination assays, these anti-peptide antibodies react with Xg(a+) but not Xg(a-) erythrocytes. In antibody-specific immobilization of antigen (ASIA) and immunoblot assays, the anti-peptide antibodies react with the same molecule as does human anti-Xga. Therefore, by its identity with PBDX, Xga is identified as a cell-surface protein that is 48% homologous to CD99 (previously designated the 12E7 antigen), the product of MIC2 which is tightly linked to XG. PBDX is renamed here XG.
Asunto(s)
Antígenos CD , Antígenos de Grupos Sanguíneos/genética , Moléculas de Adhesión Celular/genética , Genes , Antígeno 12E7 , Secuencia de Aminoácidos , Animales , Anticuerpos Monoclonales/inmunología , Secuencia de Bases , Western Blotting , Moléculas de Adhesión Celular/inmunología , Moléculas de Adhesión Celular/fisiología , Reacciones Cruzadas , Epítopos/inmunología , Fibroblastos/metabolismo , Ligamiento Genético , Pruebas de Hemaglutinación , Células Madre Hematopoyéticas/metabolismo , Humanos , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/inmunología , Ratones , Datos de Secuencia Molecular , Especificidad de Órganos , Reacción en Cadena de la Polimerasa , ARN Mensajero/biosíntesis , Conejos , Homología de Secuencia de Aminoácido , Células Tumorales CultivadasRESUMEN
BACKGROUND: The successful closure of a nasal septal perforation is a surgical challenge, which applies especially to large defects with a diameter exceeding 2.0 × 2.0 cm. This retrospective study presents results using open septoplasty via a transcolumellar approach and bipedicled flaps according to Schultz-Coulon for closure. RESULTS: In 45/50 patients the perforation was closed successfully (90%) (CI 0.82-0.98). The survey of patients showed significant improvement rates for all questioned parameters. CONCLUSIONS: In our hands the bipedicled flap technique combined with open access to the nasal septum yields good success rates for closure and consecutive improvement of life quality, also for large defects exceeding 2.0 × 3.0 cm. The open approach provides cosmetically excellent results based on the appropriate suture techniques and is advantageous concerning intraoperative overview and suture of the mucosal flaps. Additionally it can be easily combined with an open septorhinoplasty.
Asunto(s)
Perforación del Tabique Nasal/cirugía , Rinoplastia/métodos , Colgajos Quirúrgicos/cirugía , Adolescente , Adulto , Cartílago/trasplante , Terapia Combinada , Estética , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Satisfacción del Paciente , Complicaciones Posoperatorias/etiología , Complicaciones Posoperatorias/cirugía , Calidad de Vida , Recurrencia , Reoperación , Estudios Retrospectivos , Técnicas de Sutura , Adulto JovenRESUMEN
The clinical development of a candidate p38 kinase inhibitor was terminated because of its unexpectedly rapid clearance in human subjects. Its short half-life and metabolic profile in human beings were vastly different from that in rats, dogs, and monkeys characterized during routine pre-clinical studies. Mice generated the predominant drug (4-hydroxylated) metabolite produced in human beings, which was not found in other species. The data from a murine in vitro drug biotransformation assay that used liver extracts from 14 inbred mouse strains were analyzed by haplotype-based computational genetic analysis. This led to the identification of aldehyde oxidase-1 (AOX1) as the enzyme responsible for the rapid metabolism of this drug. Specific enzyme inhibitors and expressed recombinant enzymes were used to confirm that AOX catalyzed the formation of the 4-hydroxylated drug metabolite in mouse and man. Genetic variation within Aox1 regulated the level of hepatic Aox1 mRNA, AOX1 protein, and enzyme activity among the inbred strains. Thus, computational murine pharmacogenetic analysis can facilitate the identification and characterization of drug metabolism pathways that are differentially utilized by humans and other species.
Asunto(s)
Aldehído Oxidasa/metabolismo , Farmacogenética/métodos , Inhibidores de Proteínas Quinasas/farmacocinética , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Adolescente , Adulto , Aldehído Oxidasa/biosíntesis , Aldehído Oxidasa/genética , Animales , Artritis Reumatoide/tratamiento farmacológico , Perros , Femenino , Semivida , Haplorrinos , Humanos , Hígado/enzimología , Hígado/metabolismo , Masculino , Ratones , Ratones Endogámicos , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Pirimidinonas/farmacología , Ratas , Ratas Endogámicas BB , Método Simple Ciego , Especificidad de la Especie , Adulto Joven , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
The nematode parasites that cause human lymphatic filariasis survive for long periods in their vascular habitats despite continual exposure to host cells. Since prostanoids formed from arachidonic acid can modulate interactions among platelets, leukocytes, and endothelial cells, we examined whether intravascular nematode parasites can elaborate prostanoids. Microfilariae of Brugia malayi utilize exogenous and endogenous arachidonic acid to generate and release two predominant prostanoids, prostacyclin and prostaglandin E2. Filarial metabolism of host fatty acids to form these vasodilatory, antiaggregatory, and immunomodulatory eicosanoids provides a means by which these helminthic parasites may influence host immune and other cellular responses.
Asunto(s)
Brugia/metabolismo , Eicosanoides/biosíntesis , Prostaglandina-Endoperóxido Sintasas/metabolismo , Animales , Ácido Araquidónico , Ácidos Araquidónicos/metabolismo , Brugia/aislamiento & purificación , Cromatografía en Capa Delgada , Prostaglandinas/biosíntesis , Prostaglandinas/aislamiento & purificación , Tromboxano B2/biosíntesis , Tromboxano B2/aislamiento & purificaciónRESUMEN
The CD4 glycoprotein, expressed on leukocytes belonging to subsets of T lymphocytes and to cells of monocyte/macrophage lineage, participates in the functioning of T cells and serves as a receptor for HIV-1 and HIV-2. Human eosinophils, a class of granulocytic leukocytes, have been found to express CD4. With anti-CD4 mAbs CD4 was demonstrable on eosinophils from both normal and eosinophilic donors. Eosinophils synthesized a 55-kD CD4 polypeptide immunoprecipitable with two anti-CD4 mAbs. Eosinophil CD4 bound HIV-1 gp120 as assessed by competition for anti-OKT4A, but not anti-OKT4, mAb binding. Eosinophils, normally rich in gastrointestinal and genitourinary tract tissues, increase in numbers in patients with metazoan parasitic infections. In these sites and diseases, CD4 expression by eosinophils may be pertinent to their immunologic functions and could make these cells susceptible to HIV infection.
Asunto(s)
Antígenos de Diferenciación de Linfocitos T/metabolismo , Eosinófilos/metabolismo , VIH/metabolismo , Proteínas de los Retroviridae/metabolismo , Proteínas del Envoltorio Viral/metabolismo , Citometría de Flujo , HumanosRESUMEN
Lymphocyte chemoattractant factor (LCF) is a tetrameric glycoprotein of 56,000 relative molecular mass produced by activated T lymphocytes. LCF binds to CD4 and has previously been found to stimulate migration of CD4+ lymphocytes and monocytes. Because human eosinophils, like T cells and monocytes, express CD4, we examined functional responses of eosinophils to LCF. Recombinant LCF (rLCF) expressed in COS cells was purified on a CD4 affinity column. Migration of eosinophils was elicited by rLCF at low concentrations: the 50% effective dose (ED50) was 10(-12) to 10(-11) M, concentrations 100- to 1,000-fold lower than the ED50s for the recognized eosinophil chemoattractants C5a and platelet-activating factor. Two other ligands which bound to CD4, human immunodeficiency virus-1 envelope glycoprotein gp120 and monoclonal antibody OKT4, also stimulated eosinophil migration. Monovalent OKT4 Fab competitively inhibited eosinophil responses to rLCF. rLCF did not influence other functional responses of eosinophils tested, including degranulation, superoxide generation, leukotriene C4 production, in vitro survival, or surface expression of the adherence receptor CR3 (CD11b), human histocompatibility leukocyte antigen DR, or interleukin 2 receptor p55 (CD25). We conclude that CD4 on eosinophils is capable of transducing a migratory stimulus and serves as a receptor for a chemoattractant lymphokine LCF. T cell-derived LCF may contribute to recruitment of eosinophils and CD4+ mononuclear cells concomitantly at inflammatory reactions.
Asunto(s)
Antígenos CD4/fisiología , Factores Quimiotácticos/fisiología , Quimiotaxis de Leucocito , Eosinófilos/fisiología , Linfocinas/fisiología , Eosinófilos/inmunología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Antígenos HLA-DR/análisis , Humanos , Técnicas In Vitro , Interleucina-16 , Neutrófilos/fisiología , Receptores de Interleucina-2/análisis , Superóxidos/metabolismoRESUMEN
Lipid bodies, lipid rich cytoplasmic inclusions, are characteristically abundant in vivo in leukocytes associated with inflammation. Because lipid bodies are potential reservoirs of esterified arachidonate and sites at which eicosanoid-forming enzymes may localize, we evaluated mechanisms of lipid body formation in neutrophils (PMN). Among receptor-mediated agonists, platelet activating factor (PAF), but not C5a, formyl-methyl-phenylalanine, interleukin 8, or leukotriene (LT) B4, induced the rapid formation of lipid bodies in PMN. This action of PAF was receptor mediated, as it was dose dependently inhibited by the PAF receptor antagonist WEB 2086 and blocked by pertussis toxin. Lipid body induction by PAF required 5-lipoxygenase (LO) activity and was inhibited by the 5-lipoxygenase-activating protein antagonist MK 886 and the 5-LO inhibitor zileuton, but not by cyclooxygenase inhibitors. Corroborating the dependency of PAF-induced lipid body formation on 5-LO, PMN and macrophages from wild-type mice, but not from 5-LO genetically deficient mice, formed lipid bodies on exposure to PAF both in vitro and in vivo within the pleural cavity. The 5-LO product inducing lipid body formation was not LTB4 but was 5(S)-hydroxyeicosatetraenoic acid [5(S)-HETE], which was active at 10-fold lower concentrations than PAF and was also inhibited by pertussis toxin but not by zileuton or WEB 2086. Furthermore, 5-HETE was equally effective in inducing lipid body formation in both wild-type and 5-LO genetically deficient mice. Both PAF- and 5(S)-HETE-induced lipid body formation were inhibited by protein kinase C (PKC) inhibitors staurosporine and chelerythrine, the phospholipase C (PLC) inhibitors D609 and U-73122, and by actinomycin D and cycloheximide. Prior stimulation of human PMN with PAF to form lipid bodies enhanced eicosanoid production in response to submaximal stimulation with the calcium ionophore A23187; and the levels of both prostaglandin (PG) E2 and LTB4 correlated with the number of lipid bodies. Furthermore, pretreatment of cells with actinomycin D or cycloheximide inhibited not only the induction of lipid body formation by PAF, but also the PAF-induced "priming" for enhanced PGE2 and LTB4 in PMN. Thus, the compartmentalization of lipids to form lipid bodies in PMN is dependent on specific cellular responses that can be PAF receptor mediated, involves signaling through 5-LO to form 5-HETE and then through PKC and PLC, and requires new protein synthesis. Since increases in lipid body numbers correlated with priming for enhanced PGE2 and LTB4 production in PMN, the induction of lipid bodies may have a role in the formation of eicosanoid mediators by leukocytes involved in inflammation.
Asunto(s)
Araquidonato 5-Lipooxigenasa/metabolismo , Compartimento Celular , Cuerpos de Inclusión/efectos de los fármacos , Metabolismo de los Lípidos , Neutrófilos/efectos de los fármacos , Factor de Activación Plaquetaria/farmacología , Receptores de Superficie Celular , Receptores Acoplados a Proteínas G , Animales , Araquidonato 5-Lipooxigenasa/genética , Relación Dosis-Respuesta a Droga , Proteínas de Unión al GTP/metabolismo , Humanos , Inhibidores de la Lipooxigenasa/farmacología , Ratones , Ratones Noqueados , Neutrófilos/enzimología , Glicoproteínas de Membrana Plaquetaria/metabolismo , Proteína Quinasa C/metabolismo , Inhibidores de la Síntesis de la Proteína/farmacología , Transducción de Señal , Fosfolipasas de Tipo C/metabolismoRESUMEN
The specific intracellular sites at which enzymes act to generate arachidonate-derived eicosanoid mediators of inflammation are uncertain. We evaluated the formation and function of cytoplasmic lipid bodies. Lipid body formation in eosinophils was a rapidly (<1 h) inducible response which was platelet-activating factor (PAF) receptor-mediated, involved signaling through protein kinase C, and required new protein synthesis. In intact and enucleated eosinophils, the PAF-induced increases in lipid body numbers correlated with enhanced production of both lipoxygenase- and cyclooxygenase-derived eicosanoids. All principal eosinophil eicosanoid-forming enzymes, 5-lipoxygenase, leukotriene C4 synthase, and cyclooxygenase, were immunolocalized to native as well as newly induced lipid bodies in intact and enucleated eosinophils. Thus, lipid bodies are structurally distinct, inducible, nonnuclear sites for enhanced synthesis of paracrine eicosanoid mediators of inflammation.
Asunto(s)
Eicosanoides/biosíntesis , Eosinófilos/metabolismo , Eosinófilos/ultraestructura , Cuerpos de Inclusión/metabolismo , Cuerpos de Inclusión/ultraestructura , Metabolismo de los Lípidos , Adulto , Araquidonato 5-Lipooxigenasa/metabolismo , Sitios de Unión , Núcleo Celular/metabolismo , Inhibidores Enzimáticos/farmacología , Eosinófilos/efectos de los fármacos , Glutatión Transferasa/metabolismo , Humanos , Inmunohistoquímica , Técnicas In Vitro , Cuerpos de Inclusión/efectos de los fármacos , Mediadores de Inflamación/metabolismo , Factor de Activación Plaquetaria/farmacología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/metabolismo , Inhibidores de la Síntesis de la Proteína/farmacología , Transducción de SeñalRESUMEN
Transforming growth factor alpha (TGF-alpha) is a pleuripotential cytokine with diverse biological effects, including the ability to influence the proliferation of normal cells or neoplastic epithelial cells. Eosinophils are a subset of granulocytes that normally enter the peripheral tissues, particularly those beneath gastrointestinal, respiratory, and urogenital epithelium, where they reside in close proximity to the epithelial elements. In this study, we demonstrate that the great majority of eosinophils infiltrating the interstitial tissues adjacent to two colonic adenocarcinomas and two oral squamous cell carcinomas labeled specifically by in situ hybridization with a 35S-riboprobe for human TGF-alpha (hTGF-alpha). No other identifiable leukocytes in these lesions contained detectable hTGF-alpha mRNA. We also examined leukocytes purified from a patient with the idiopathic hypereosinophilic syndrome. 80% of these eosinophils, but none of the patient's neutrophils or mononuclear cells, were positive for hTGF-alpha mRNA by in situ hybridization, and 55% of these eosinophils were positive by immunohistochemistry with a monoclonal antibody directed against the COOH terminus of the mature hTGF-alpha peptide. Finally, the identification of the purified eosinophil-associated transcript as hTGF-alpha was confirmed by polymerase chain reaction product restriction enzyme analysis followed by Southern blot hybridization. In contrast to eosinophils from the patient with hypereosinophilic syndrome, the peripheral blood eosinophils from only two of seven normal donors had detectable TGF-alpha mRNA and none of these eosinophils contained immunohistochemically detectable TGF-alpha product. Taken together, these findings establish that human eosinophils can express TGF-alpha, but suggest that the expression of TGF-alpha by eosinophils may be under microenvironmental regulation. Demonstration of TGF-alpha production by tissue-infiltrating eosinophils and the eosinophils in the hypereosinophilic syndrome identifies a novel mechanism by which eosinophils might contribute to physiological, immunological, and pathological responses.
Asunto(s)
Adenocarcinoma/genética , Neoplasias del Colon/genética , Eosinófilos/metabolismo , Neoplasias de la Boca/genética , Factores de Crecimiento Transformadores/genética , Secuencia de Bases , Northern Blotting , Línea Celular , Eosinofilia/sangre , Eosinofilia/genética , Humanos , Datos de Secuencia Molecular , Sondas de Oligonucleótidos , ARN/genética , ARN/aislamiento & purificación , ARN Mensajero/genética , ARN Neoplásico/genética , ARN Neoplásico/aislamiento & purificación , Síndrome , Factores de Crecimiento Transformadores/sangreRESUMEN
We report that the hypodense eosinophil population in three patients with corticosteroid-unresponsive IHES was uniquely long lived ex vivo in the absence of exogenous cytokines. Serum or plasma from these patients conferred prolonged viability ex vivo to normodense eosinophils from reference donors and converted them to a functionally activated hypodense phenotype. In that antibody against IL-5 neutralized this activity in IHES serum, excessive quantities of this cytokine may account for the characteristic eosinophilia and long-lived, functionally augmented eosinophil phenotype in this disorder.
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Eosinofilia/sangre , Eosinófilos/patología , Interleucinas/sangre , Adulto , Supervivencia Celular , Eosinofilia/inmunología , Eosinófilos/citología , Humanos , Técnicas In Vitro , Interleucina-5 , Masculino , Persona de Mediana Edad , Fenotipo , Valores de Referencia , SíndromeAsunto(s)
Suelo de la Boca/cirugía , Neoplasias de la Boca/cirugía , Disección del Cuello , Picibanil/administración & dosificación , Complicaciones Posoperatorias/tratamiento farmacológico , Ránula/tratamiento farmacológico , Adulto , Diagnóstico Diferencial , Estudios de Seguimiento , Humanos , Inyecciones Intralesiones , Imagen por Resonancia Magnética , Masculino , Complicaciones Posoperatorias/diagnóstico , Ránula/diagnóstico , Succión , Resultado del TratamientoRESUMEN
Lipid bodies, nonmembrane-bound cytoplasmic inclusions, serve as repositories of esterified arachidonate and are increased in cells associated with inflammatory reactions. We have evaluated stimuli and mechanisms responsible for lipid body formation within human polymorphonuclear leukocytes (PMNs). Arachidonic acid and oleic acid stimulated dose-dependent formation of lipid bodies over 0.5-1 h. Other C20 and C18 fatty acids were less active and demonstrated rank orders as follows: cis-unsaturated fatty acids were much more active than trans-fatty acids, and activity diminished with decreasing numbers of double bonds. Lipid bodies elicited in vitro with cis-fatty acids were ultrastructurally identical to lipid bodies present in PMNs in vivo. Lipid body induction was not because of fatty acid-elicited oxidants or fatty acid-induced ATP depletion. Cis-fatty acid-induced activation of protein kinase C (PKC) was involved in lipid body formation as evidenced by the capacity of other PKC activators, 1-oleoyl-2-acetyl-glycerol and two active phorbol esters, phorbol myristate acetate, and phorbol 12,13 dibutyrate, but not an inactive phorbol, to induce lipid body formation. The PKC inhibitor, 1-O-hexadecyl-2-O-methyl-glycerol, inhibited PMN lipid body formation induced by oleic and arachidonic acids and by 1-oleoyl-2-acetyl-glycerol and phorbol myristate acetate. Other PKC inhibitors (staurosporine, H-7) also inhibited lipid body formation. Formation of lipid bodies in PMNs is a specific cellular response, stimulated by cis-fatty acids and diglycerides and apparently mediated by PKC, which results in the mobilization and deposition of lipids within discrete, ultrastructurally defined cytoplasmic domains.