RESUMEN

The clinical course of immune or idiopathic thrombocytopenic purpura (ITP) is variable, suggesting different mechanisms for the decreased platelet count. The complement factors C3 and C4 have been detected on platelets, both alone and in association with immunoglobulin G (IgG), and a reduced platelet survival time has been described. Platelets have the capacity to interact with the complement system since they have both complement receptors and complement regulatory proteins on their cell membranes. The membrane attack complex (C5b-9) induced by antiplatelet antibodies generates platelet microparticles in a concentration-dependent manner. A marked variation in resistance to this phenomenon has been demonstrated between individuals and between men and women. These platelet microparticles seem to retain their biological role in haemostasis. Platelets also appear to play a role in the processing of immune complexes. Immunoglobulins and complement factors are found in several clinical situations where circulating immune complexes are expected. Furthermore, human platelets bind immune complexes in vitro and the reaction can be blocked by antireceptor antibodies to immunoglobulins and complement. These findings raise a number of questions about the role of complement in the pathophysiology of ITP.

Asunto(s)

Plaquetas/inmunología , Proteínas del Sistema Complemento/inmunología , Inmunoglobulina G/inmunología , Púrpura Trombocitopénica Idiopática/inmunología , Humanos

RESUMEN

We have previously found that CD9, CD11b, and intracellular ECP (EG2) may be used as activity markers for eosinophils in vitro. The main object of the present study was to determine whether these markers can reflect eosinophil activation in vivo in relation to allergen exposure. for this purpose, six patients with a history of allergic rhinitis and occasional asthma symptoms during the pollen season participated. Blood donors served as controls. Peripheral blood eosinophils were analyzed according to the FOG method and flow cytometry, before and during one birch pollen season with high pollen load (HPL) and one with low pollen load (LPL). The CD9 expression on peripheral eosinophils from the patients was significantly increased both before (P < 0.05) and during (P < 0.01) HPL season, and CD11b expression solely during HPL season (P = 0.01) as compared to controls. The intracellular expression of the EG2 epitope was increased before (P < 0.01) and during (P < 0.05) HPL season, and increased significantly (P < 0.05) during season as compared to before. No changes were observed before and during LPL season. The proportion of eosinophils was increased both before (P < 0.05) and during (P < 0.001) the HPL season as compared to controls. The markers CD9, EG2, and, to a lesser extent, CD11b seem to detect activated eosinophils in the circulation, whereas EG2 may also reflect increased antigen exposure during season.

Asunto(s)

Alérgenos/efectos adversos , Antígenos CD/inmunología , Asma/inmunología , Proteínas Sanguíneas/inmunología , Eosinófilos/inmunología , Mediadores de Inflamación/inmunología , Antígeno de Macrófago-1/inmunología , Glicoproteínas de Membrana , Polen , Ribonucleasas , Adulto , Asma/sangre , Biomarcadores/sangre , Estudios de Casos y Controles , Proteínas en los Gránulos del Eosinófilo , Femenino , Humanos , Recuento de Leucocitos , Masculino , Persona de Mediana Edad , Estaciones del Año , Índice de Severidad de la Enfermedad , Tetraspanina 29

RESUMEN

The objective of the present study was to investigate the potential of the long-acting beta 2-agonist salmeterol as an inhibitor of various components of IgE-mediated inflammatory in man. For this purpose, we measured gross skin reactivity (diameters of wheal and flare reaction [WFR] and late cutaneous reaction [LCR] as well as inflammatory cells, mediators, and protein in cutaneous suction blister chambers in eight subjects with allergic rhinitis. Blisters were induced, two on each forearm, by gentle suction and heating, and were unroofed 12 h later, after which plastic chambers were placed over the denuded area. The chambers were challenged for 2 h with antihuman IgE (titer 1:10) in the presence and absence of salmeterol or terbutaline. Normal goat IgG served as negative control. Chamber fluids were removed hourly for the first 4 h, and this followed by a 4-h incubation before final collection. Salmeterol (10(-6)M) and terbutaline (10(-5)M) injected intradermally 30 min before, aw well as together with anti-IgE (titer 1:100), inhibited the WFRs by up to 30%. The effect of salmeterol on the ensuing LCR (75% inhibition at 24 h) tended to be more pronounced than the corresponding inhibition by terbutaline. Both salmeterol and terbutaline very effectively inhibited the anti-IgE-induced extravasation of alpha 2-macroglobulin into skin chambers, with a significantly more sustained effect by salmeterol. Interestingly, only terbutaline reduced the histamine release evoked by anti-IgE. With the present experimental design, where both drugs were washed out from the chambers after 2 h, neither drug inhibited recruitment of leukocytes (including eosinophils). Taken together, salmeterol had a more sustained inhibitory effect than terbutaline on indices of IgE-mediated edema formation (late induration and plasma protein extravasation). On the other hand, under the present experimental conditions, salmeterol failed to reduce the histamine release (in contrast to terbutaline), and neither salmeterol nor terbutaline affected the recruitment of leukocytes.

Asunto(s)

Agonistas Adrenérgicos beta/uso terapéutico , Albuterol/análogos & derivados , Dermatitis Atópica/tratamiento farmacológico , Premedicación , Rinitis Alérgica Estacional/complicaciones , Terbutalina/uso terapéutico , Administración Cutánea , Adulto , Albuterol/uso terapéutico , Vesícula/tratamiento farmacológico , Dermatitis Atópica/complicaciones , Dermatitis Atópica/inmunología , Humanos , Persona de Mediana Edad , Xinafoato de Salmeterol

RESUMEN

The present study aimed to evaluate the predictive value of eosinophils and markers of their activity for bronchial hyperreactivity (BHR) in a population of patients with recently developed clinical symptoms of asthma. The activation of eosinophils was estimated by measuring eosinophil cationic protein (ECP) in serum. In addition, flow cytometry was used to measure the expression of the EG2-epitope on intracellular ECP in eosinophils from peripheral blood. Twenty-eight consecutive patients with clinical history of asthma were studied. Of the 28 patients, 18 had a positive bronchial challenge test measured as PD20 < or = 1600 micrograms histamine. A significantly higher concentration of eosinophils and a trend to higher ECP in the peripheral blood was found in the hyperreactive group than in the nonreactive group. However, the intracellular expression of ECP did not correlate with the PD20 value, and no significant difference between the groups was found. With one eosinophil activity marker, either serum ECP or EG2, BHR could be predicted in 70% of the patients. If we combined any two of the activity markers (serum ECP, EG2, or the percentage of eosinophils), the predictive value increased to 100%. We conclude that the blood eosinophil concentration, as well as, to some extent, serum ECP, has a high specificity for BHR in patients with recently developed clinical symptoms of asthma. Despite normal bronchial reactivity, some patients had signs of activated eosinophils, i.e., high serum ECP and increased EG2 expression. Thus, these markers may reflect early stages in the development of BHR. Our results also indicate that a combined evaluation of percentage of eosinophils and of eosinophil activity markers is of clinical value to predict BHR.

Asunto(s)

Asma/fisiopatología , Proteínas Sanguíneas/análisis , Hiperreactividad Bronquial/diagnóstico , Ribonucleasas , Adolescente , Adulto , Proteínas Sanguíneas/inmunología , Hiperreactividad Bronquial/epidemiología , Pruebas de Provocación Bronquial , Proteínas en los Gránulos del Eosinófilo , Eosinófilos/química , Eosinófilos/citología , Epítopos/fisiología , Femenino , Humanos , Mediadores de Inflamación/análisis , Recuento de Leucocitos , Masculino , Persona de Mediana Edad , Factores de Riesgo

RESUMEN

The aim of this study was to examine potential differences between healthy and atopic subjects with regard to IgE-mediated cutaneous inflammation. For this purpose, we analyzed histamine, tryptase, leukotriene B4, albumin, eosinophils, and total leukocytes in skin chamber fluid after challenge with anti-human IgE. We also measured gross skin reactivity (wheal, flare, and late-phase reactions), circulating IgE, and eosinophils, as well as the state of eosinophil activation. It was found that despite having more circulating IgE, the skin responsiveness of the atopic subjects did not differ significantly from that of the nonatopic subjects with respect to mediator release, albumin extravasation, or total recruitment of leukocytes. Moreover, the sizes of anti-IgE-induced wheal, flare, and late-phase reactions were very similar in the two groups. On the other hand, significant recruitment of eosinophils during the IgE-mediated reaction was more or less restricted to the atopic group. Yet the recruited eosinophils, of which the majority was in an early state of activation before degranulation, did not seem to contribute significantly to the IgE-mediated delayed skin edema. Furthermore, the eosinophil count in anti-IgE chambers of the atopic subjects did not correlate with any of the other parameters monitored. Thus because the anti-IgE-induced recruitment of eosinophils appeared to be unrelated to factors such as the number of peripheral blood eosinophils, the degree of mast cell activation, the intensity of inflammatory skin changes, and the level of circulating IgE, it is apparent that the mechanisms for and pathophysiologic role of IgE-mediated dermal eosinophil accumulation in atopic subjects require further investigation.

Asunto(s)

Anticuerpos Antiidiotipos/farmacología , Quimiotaxis de Leucocito/inmunología , Hipersensibilidad Inmediata/inmunología , Hipersensibilidad Inmediata/metabolismo , Inmunoglobulina E/inmunología , Mediadores de Inflamación/metabolismo , Albúmina Sérica/metabolismo , Piel/inmunología , Cámaras de Difusión de Cultivos , Eosinófilos/inmunología , Humanos , Piel/metabolismo , Piel/patología

RESUMEN

We studied the generation of CD11b/CD18 mobilizing factors in serum after incubation with dialysis membrane fragments of different chemical composition. We also evaluated the relative importance of the alternative and classical pathways of the complement system in the generation of such factors. Monocytes and granulocytes from healthy blood donors were incubated in normal human serum (NHS) and in NHS that had been preincubated with Cuprophan (CU) membrane (NHS-CU), Hemophan (HE) (NHS-HE) or polysulfone (PS) (NHS-PS). NHS-CU caused the highest up-regulation of the CD11b/CD18 receptor on monocytes and granulocytes. The rank in capacity to mobilize CD11b/CD18 on granulocytes was CU > HE > PS (p < 0.001), CU > HE (p < 0.05) and HE > PS (p < 0.001). The rank in capacity to mobilize CD11b/CD18 on monocytes was CU > HE > PS (p < 0.001), CU > HE (p < 0.05) and HE > PS (p < 0.01). NHS-PS induced a lower up-regulation of CD11b/CD18 compared to NHS which indicates that serum factors with the ability to mobilize the CD11b/CD18 receptor on monocytes and granulocytes are deposited on or absorbed by PS. In order to study the relative contribution of the alternative and classical pathways of the complement system in the generation of CD11b/CD18 mobilizing factors in serum, three different serum preparations (1. both pathways intact. 2. only the alternative intact and 3. only the classical pathway intact) were used. The CU membrane activated the classical pathway to a larger extent than the PS membrane (p < 0.01). When only the alternative pathway was intact no difference in the generation of CD11b/CD18 mobilizing factors between the CU and PS membranes was observed. These studies show that CD11b/CD18 mobilizing serum factors are generated after incubation with CU membranes and that such factors are probably adsorbed by PS. The classical pathway of complement activation seems to contribute to the generation of CD11b/CD18 mobilizing factors in serum.

Asunto(s)

Materiales Biocompatibles/efectos adversos , Antígenos CD11/sangre , Antígenos CD18/sangre , Granulocitos/inmunología , Membranas Artificiales , Monocitos/inmunología , Materiales Biocompatibles/metabolismo , Moléculas de Adhesión Celular/metabolismo , Celulosa/efectos adversos , Celulosa/análogos & derivados , Celulosa/sangre , Activación de Complemento/efectos de los fármacos , Citometría de Flujo , Granulocitos/citología , Granulocitos/efectos de los fármacos , Humanos , Monocitos/citología , Monocitos/efectos de los fármacos , Polímeros/efectos adversos , Polímeros/metabolismo , Diálisis Renal , Coloración y Etiquetado , Relación Estructura-Actividad , Sulfonas/efectos adversos , Sulfonas/metabolismo

RESUMEN

The hampered inflammation and host defense seen in alcoholics may be due to impairment of functional responses of neutrophil polymorphonuclear leukocytes (PMN). We have shown that ethanol inhibits the oxidative metabolism of PMN induced by surface receptor dependent stimuli, such as N-formyl-methionyl-leucyl-phenylalanine (fMLP) and opsonized zymosan. Because the unresponsiveness might be due to reduced numbers of surface receptors, we assessed the expression of CR1, Fc-gamma, and fMLP receptors as well as membrane fluidity after treatment of PMN with ethanol in vitro. Ethanol impaired the induced expression of CR1 and fMLP receptors to 71% and 51% of control, respectively, but did not affect the resting level of CR1 nor Fc-gamma receptor expression. Furthermore, the mobility of cell membrane glycoconjugates was increased by ethanol. However, phagocytosis, a functional response dependent on membrane rheology, was unaffected. Because the results indicated an effect of ethanol on mobilization of receptors from intracellular stores, we assessed lactoferrin release, which was reduced to 59%. Thus, ethanol appeared to hamper the upregulation of PMN surface receptors or functional subsets of those stored in granules. Ethanol also increased the mobility of the cell membrane. These reactions were accompanied by reductions in the functional responses mediated by either class of receptors.

Asunto(s)

Etanol/farmacología , Neutrófilos/efectos de los fármacos , Receptores de Droga/metabolismo , Células Cultivadas , Humanos , Lactoferrina/metabolismo , Mediciones Luminiscentes , Fluidez de la Membrana , Neutrófilos/metabolismo , Fagocitosis , Receptores de Complemento 3b/metabolismo , Receptores Fc/metabolismo , Receptores de Formil Péptido , Receptores Inmunológicos/metabolismo , Receptores de Péptidos/metabolismo

Asunto(s)

Enfermedad Celíaca/etiología , Enfermedad Celíaca/inmunología , Hipersensibilidad a los Alimentos/complicaciones , Hipersensibilidad a los Alimentos/inmunología , Enfermedad Celíaca/fisiopatología , Hipersensibilidad a los Alimentos/fisiopatología , Humanos , Enfermedades del Complejo Inmune/inmunología

RESUMEN

Expression of CD9 is a feature of both eosinophils and platelets. We have investigated the CD9 expression on resting and activated eosinophils with regard to possibly interacting platelets. Mixed leukocytes were obtained from the platelet-containing (PC) and platelet-depleted (PD) peripheral blood of healthy donors. A cell membrane permeabilization technique, the FOG method, enabled us to detect the eosinophils as a separate population and permitted flow cytometric analysis of both surface and intracellular antigens. Monoclonal antibodies against CD61 were used to identify platelets. The CD9/CD61 ratio indicated that CD9 on resting eosinophils originates mainly from eosinophils and not from adhered platelets. No difference in CD9 expression was obtained between resting eosinophils in PC and PD blood. However, the expression of CD9 was decreased (p < 0.05) on eosinophils in PMA-activated PD blood but increased (p = 0.001) in PC blood, probably due to interacting platelets since CD61 increased simultaneously. In addition, we were able to detect an intracellularly stored pool of CD9 in eosinophils which decreased after activation with PMA. Together these results indicate a translocation of intracellularly stored CD9 to the cell membrane upon activation, probably followed by a subsequent shedding.

Asunto(s)

Antígenos CD/análisis , Eosinófilos/inmunología , Glicoproteínas de Membrana , Membrana Celular/inmunología , Células Cultivadas , Citoplasma/inmunología , Eosinófilos/citología , Citometría de Flujo , Humanos , Inmunohistoquímica , Tetraspanina 29

RESUMEN

We have investigated the effect of gradual degranulation on the expression of functional receptors (CR1 and CR3) on human neutrophils. Incubation with increasing concentrations of fMLP (10(-10) - 10(-7) M) translocated CR1 and CR3 to the cell surface in a similar kinetic pattern. When reaching maximal expression of receptors (10(-7) M fMLP), 78 +/- 10% and 87 +/- 9% of the total pool of CR1 and CR3, respectively, were translocated to the cell surface. To drive the mobilization process further, cytochalasin B was introduced to increase the stimulatory effect of fMLP. No further increase in CR1 surface expression was obtained. However, we found a characteristic time course of surface appearance of CR1 and CR3 with a maximal surface expression within 1 minute, followed by a time-related down-regulation of CR1 but not CR3. In addition, the total pool of CR1 in cytochalasin B treated neutrophils was reduced after 15 minutes stimulation with fMLP measured by flow cytometry and immunoblotting, indicating degradation of CR1. The down-regulation of CR1 was concomitant with a translocation of azurophil granules, in terms of upregulation of CD63. Azurophil, but not specific nor secretory, granule fractions caused a down-regulation of CR1 on fMLP activated neutrophils. The presence of human sera and serine protease inhibitor protected CR1 from down-regulation. Together, these findings indicate that intracellular stored proteases, released in the late part of the sequential mobilization process, alters the expression of functional receptors mobilized in the early part of the mobilization process. The findings also focus on the importance of the microenvironment for the net outcome of neutrophil activation in terms of functional receptor expression.

Asunto(s)

Gránulos Citoplasmáticos/fisiología , Regulación hacia Abajo/fisiología , Neutrófilos/metabolismo , Receptores de Complemento/biosíntesis , Adolescente , Adulto , Anciano , Fosfatasa Alcalina/metabolismo , Degranulación de la Célula/efectos de los fármacos , Degranulación de la Célula/fisiología , Complemento C1/metabolismo , Complemento C3/metabolismo , Citocalasina B/farmacología , Gránulos Citoplasmáticos/efectos de los fármacos , Gránulos Citoplasmáticos/enzimología , Regulación hacia Abajo/efectos de los fármacos , Humanos , Inmunohistoquímica , Técnicas In Vitro , Persona de Mediana Edad , N-Formilmetionina Leucil-Fenilalanina/farmacología , Neutrófilos/enzimología , Neutrófilos/ultraestructura , Receptores de Complemento/efectos de los fármacos , Inhibidores de Serina Proteinasa/farmacología , Inhibidores de Serina Proteinasa/fisiología , Fracciones Subcelulares/enzimología , Fracciones Subcelulares/metabolismo , Fracciones Subcelulares/ultraestructura

RESUMEN

BACKGROUND: Both overtreating and undertreating asthma can be harmful, either because of side effects or development of uncontrolled asthma. As the eosinophil granulocyte is of importance in asthma, it is logical to study this cell in the search for a marker for the right level of treatment. OBJECTIVE: To study the changes in eosinophil activity and the correlation to clinical status in asthmatic patients who have deteriorated and during prednisolone treatment. METHODS: Nine patients were studied on two occasions: with exacerbation (visit 1) and during prednisolone treatment (visit 2). Clinical evaluation was performed as well as analysis of percentage of eosinophils, eosinophilic cationic protein (S-ECP) in serum and expression of intracellular ECP, measured by cell flow cytometry using a monoclonal antibody (EG2) against the activated form of ECP. RESULTS: Visit 1: four patients had eosinophils above normal, mean 6.5%, range 1.5 to 13.8. S-ECP was measured in seven patients of whom five had values above normal (16 micrograms/L), mean 29.5, range 1.5 to 13.8. Intracellular expression of ECP was above normal (mean 26.7 +/- 10.8) in seven out of nine (mean 48.1, range 28.8 to 69.6). Visit 2: all patients improved and all eosinophil parameters decreased. The eosinophil concentration fell to a mean of 3.0%, range 1.0 to 6.6, S-ECP to a mean of 10.5 micrograms/L, range 3.2 to 17 and intracellular expression of ECP to a mean of 33.4, range 19.4 to 40.9. CONCLUSIONS: Reduction in eosinophil activity followed improvement in clinical condition. Measuring intracellular expression of ECP may be of value in addition to eosinophil count and S-ECP in monitoring asthma.

Asunto(s)

Antiinflamatorios/uso terapéutico , Asma/tratamiento farmacológico , Asma/inmunología , Eosinófilos/inmunología , Prednisolona/uso terapéutico , Ribonucleasas , Adulto , Biomarcadores , Proteínas Sanguíneas/inmunología , Proteínas en los Gránulos del Eosinófilo , Femenino , Citometría de Flujo , Humanos , Mediadores de Inflamación/inmunología , Recuento de Leucocitos , Masculino , Persona de Mediana Edad

RESUMEN

After severe burns, a wound revision is often done to remove devitalized tissue and minimize bacterial growth. After such revision, the patient may show signs of sepsis. In a group of burned patients we found a transient endotoxemia, and a subsequent leukocyte activation, monitored as increased expression of the beta 2-integrin CD11b, after such wound revision. In most patients we could detect elevated levels of plasma TNF-alpha before the operation, with no increases in these levels after the operation. Plasma levels of IL-6 were elevated in all patients and increased after the wound revision in all patients. They also had elevated plasma levels of soluble E-selectin, indicating systemic inflammation. The close relation between endotoxin levels and CD11b expression, and lack of evidence for additional production of TNF-alpha, suggests that up-regulation of the beta 2 adhesion protein during wound revision is mainly caused by endotoxin interaction with the leukocyte.

Asunto(s)

Quemaduras/complicaciones , Antígenos CD18/biosíntesis , Quimiocinas/metabolismo , Desbridamiento/efectos adversos , Selectina E/sangre , Endotoxinas/sangre , Leucocitos/metabolismo , Antígeno de Macrófago-1/biosíntesis , Síndrome de Respuesta Inflamatoria Sistémica/etiología , Adolescente , Adulto , Anciano , Quemaduras/cirugía , Adhesión Celular , Activación de Complemento , Femenino , Humanos , Masculino , Persona de Mediana Edad , Síndrome de Respuesta Inflamatoria Sistémica/metabolismo , Factor de Necrosis Tumoral alfa/análisis , Regulación hacia Arriba

RESUMEN

The aim of this study was to assess the ability of the H1-receptor antagonist loratadine to modify anti-IgE-induced cutaneous wheal-and-flare and late-phase reactions (WFR and LPR), as well as histamine release and leukocyte accumulation in skin chambers. For this purpose, 10 atopics with allergic rhinitis were entered into a double-blind crossover study in which they received either placebo or loratadine (20 mg/day orally) for 8 days separated by a 7-day washout period. Blisters were induced on both forearms on day 7 of each treatment period, and were unroofed on day 8 and covered with plastic skin chambers. Chamber fluids were collected during 7 h after 1-h incubation with anti-IgE or control IgG. Intradermal challenge with histamine and anti-IgE was performed at the same occasion. As compared to placebo treatment, loratadine inhibited the immediate WFRs to anti-IgE by 35% (wheal) and 65% (flare), respectively (P < 0.01), and corresponding reactions to histamine challenge by 50% and 70% (P < 0.001), respectively. Moreover, the initial phase (0-2 h) of the LPR induced by anti-IgE was attenuated by up to approximately 60% (P < 0.001) during loratadine treatment. Thereafter, no inhibition of the LPR was observed. The magnitude and time course of histamine release into skin chambers was virtually the same after loratadine and placebo treatment, with a peak during 0-1 h and a progressive decline during the following 2 h. Accumulation of alpha 2-macroglobulin, reflecting extravasation of large plasma proteins, also peaked during the first hour and was unaffected by loratadine during the 8-h observation period.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

Anticuerpos Antiidiotipos/inmunología , Dermatitis Atópica/tratamiento farmacológico , Inmunoglobulina E/inmunología , Leucocitos/fisiología , Loratadina/uso terapéutico , Piel/inmunología , Adulto , Estudios Cruzados , Dermatitis Atópica/inmunología , Método Doble Ciego , Femenino , Liberación de Histamina/efectos de los fármacos , Humanos , Leucocitos/efectos de los fármacos , Masculino , Piel/metabolismo , Piel/patología , Pruebas Cutáneas , alfa-Macroglobulinas/metabolismo

RESUMEN

This study was designed to identify differences in the immunological reactions in adenoid tissue between children suffering from chronic secretory otitis media (SOM) and control children without ear disease. Cell populations were identified using monoclonal antibodies and flow cytofluorometry to facilitate quantitative comparisons. A modification of the FOG method was developed to quantify lymphocytes with intracellular IgG and IgA. Immunological screening was done in the first part of the study. No significant differences were found between the groups regarding cells positive for CD3, CD4, CD8, CD20 or CD25. A significantly higher number of PCA-1 positive cells (presumably plasma cells) were found in the SOM group. The second part of the study concentrated specifically on cells containing IgG or IgA. No statistically significant differences in number of positive cells were found between the groups. When we related the percentage of positive cells to age, a statistically significant decrease with age for IgA-positive cells was found in the SOM group but not in the control group. This result supports the hypothesis that SOM is associated with an immunological reaction that influences immunoglobulin production in adenoid tissue.

Asunto(s)

Tonsila Faríngea/inmunología , Inmunoglobulina A/análisis , Inmunoglobulina G/análisis , Subgrupos Linfocitarios , Otitis Media con Derrame/inmunología , Adolescente , Factores de Edad , Antígenos CD/análisis , Antígenos de Diferenciación de Linfocitos B/análisis , Niño , Preescolar , Femenino , Citometría de Flujo , Humanos , Lactante , Recuento de Linfocitos , Masculino

RESUMEN

We have investigated the effect of different cell preparation procedures on the surface expression of CD11b/CD18 and CD62L on human monocytes. Both EDTA and heparin anticoagulated blood were used as sources for leukocytes. The monocytes were analysed by flow cytometry in a mixed leukocyte suspensions obtained after ammonium chloride mediated lysis and in mononuclear cell suspension prepared by density gradient centrifugation (Ficoll-Paque) performed both at 4 degrees C and at 20 degrees C. Monocytes from heparinized blood had a higher expression of CD11b/CD18 and a more pronounced inter-individual variation than monocytes from EDTA blood. Monocytes isolated by Ficoll-Paque had a higher degree of ex vivo activation by means of altered expression of CD11b/CD18 and CD62L compared to monocytes prepared by ammonium chloride mediated lysis. This was more pronounced when the isolation procedure was performed at 20 degrees C. Our findings indicate that monocytes prepared by ammonium chloride mediated lysis of EDTA blood and with the cell handling temperature kept at 4 degrees C are exposed to the smallest ex vivo modulation by means of receptor alteration. An awareness of ex vivo modulation by different cell preparation procedures is of importance especially when comparing the expression of functional receptors on leukocytes of disparate origin.

Asunto(s)

Antígenos CD11/biosíntesis , Antígenos CD18/biosíntesis , Separación Celular/métodos , Monocitos/inmunología , Glicoproteínas de Membrana Plaquetaria/biosíntesis , Adolescente , Adulto , Anciano , Antígenos CD/biosíntesis , Centrifugación por Gradiente de Densidad , Citometría de Flujo , Humanos , Persona de Mediana Edad , Monocitos/citología , Selectina-P

RESUMEN

We studied the modulation of cell surface receptors related to cell adhesion (L-selectin and Mac-1) on monocytes and granulocytes during clinical (7 patients treated with cuprophan, Cu, and polysulfone, PS, membranes, n = 14) and experimental Cu and PS hemodialysis (n = 14). The objective was to compare cell surface receptor modulation in vivo when large subpopulations of cells are withdrawn from the circulating pool with the experimental model when cells are not sequestrated. The expression of Mac-1 and L-selectin on monocytes increased during clinical Cu dialysis (p = 0.024 and p = 0.0096, respectively) but remained stable during PS dialysis. On granulocytes, an inverse receptor modulation of Mac-1 and L-selectin was observed during clinical Cu dialysis but not during PS dialysis. Mac-1 was significantly higher and L-selectin lower on granulocytes after 15 min of clinical Cu as compared to PS dialysis (p = 0.001 and p = 0.0093, respectively). During experimental Cu dialysis, Mac-1 expression increased and L-selectin decreased markedly and continuously on both monocytes and granulocytes. The L-selectin/Mac-1 ratio on monocytes and granulocytes may be used as an index of the ability of leukocytes to adhere and to be recruited to an inflammatory focus. This ratio was significantly lower during clinical Cu as compared to PS dialysis (p = 0.0008 and p = 0.0015 respectively) indicating that the recruitment of leukocytes to infection foci may be precluded in patients on Cu membranes. Both monocytes and granulocytes showed significantly lower L-selectin/Mac-1 ratio during and after experimental Cu as compared to PS dialysis.(ABSTRACT TRUNCATED AT 250 WORDS)

Asunto(s)

Granulocitos/inmunología , Selectina L/sangre , Antígeno de Macrófago-1/sangre , Monocitos/inmunología , Diálisis Renal , Anciano , Adhesión Celular , Celulosa/análogos & derivados , Femenino , Citometría de Flujo , Granulocitos/metabolismo , Humanos , Fallo Renal Crónico/inmunología , Fallo Renal Crónico/metabolismo , Fallo Renal Crónico/terapia , Masculino , Membranas Artificiales , Persona de Mediana Edad , Monocitos/metabolismo , Polímeros , Sulfonas

RESUMEN

Exposure to sawdust and its contaminants, e.g., terpenes, may cause respiratory tract and lung parenchymal inflammation. To monitor these changes over time. Sprague-Dawley rats were exposed at one occasion to 2.5 mg sawdust or saline by intratracheal instillation. Flow cytometry analyses were done on bronchoalveolar lavage (BAL) cells. Lung tissue specimens were analyzed histologically and immunohistochemically. After one week, the number of BAL polymorphonuclear leukocytes was increased (P < 0.05, N = 8), followed at six weeks by increases of macrophages and lymphocytes (both P < 0.01, N = 8). Enhanced expressions of class II antigens and complement receptors on macrophages after one week were even more pronounced at six weeks, indicating cellular activation. The BAL findings, also including increased (P < 0.001, N = 8) concentrations of hyaluronan with progressing changes over time, confirmed the signs of inflammation, as did the histological analysis of the lung tissue specimens with an accumulation of polymorphonuclears, macrophages, and hyaluronan in the interstitium.

Asunto(s)

Tejido Conectivo/patología , Neumoconiosis/patología , Alveolos Pulmonares/patología , Madera , Animales , Líquido del Lavado Bronquioalveolar , Tejido Conectivo/microbiología , Endotoxinas/análisis , Femenino , Citometría de Flujo , Macrófagos Alveolares/patología , Tamaño de la Partícula , Neumoconiosis/etiología , Alveolos Pulmonares/microbiología , Ratas , Ratas Sprague-Dawley , Esporas Bacterianas/aislamiento & purificación , Esporas Fúngicas/aislamiento & purificación , Terpenos/análisis

RESUMEN

Many attempts have been made to find screening tests for celiac disease to reduce the need for biopsy, or to achieve better selection criteria before intestinal biopsy. We have recently analyzed apparently healthy blood donors for antigliadin antibodies (AGA) to select subjects for further gastrointestinal investigation. A prevalence of gluten enteropathy of at least 1/256 was found in this population. The positive predictive value (+PV), however, was only 20%. In the present study we have analyzed IgA antiendomysium antibodies (IgA-EmA) to estimate the sensitivity and specificity of the test, and determine whether or not the +PV of the assay increases when screening for adult celiac disease in an asymptomatic population. We found that asymptomatic persons with celiac disease may have IgA-EmA. We found a 100% specificity of IgA-EmA in the tested population of blood donors, whereas the sensitivity was about the same as that of IgA-AGA. This result of a +PV of 100% indicates that a positive IgA-EmA could replace biopsy in diagnosing celiac disease. However, further extended studies are needed to determine whether this is applicable in other populations. To screen patients for celiac disease, we recommend the easy and cheap IgA-AGA assay as a preliminary test and the IgA-EmA to verify the diagnosis and avoid unnecessary biopsies.

Asunto(s)

Autoanticuerpos/inmunología , Enfermedad Celíaca/diagnóstico , Inmunoglobulina A/análisis , Músculos/inmunología , Adulto , Anticuerpos/análisis , Enfermedad Celíaca/inmunología , Técnica del Anticuerpo Fluorescente , Gliadina/inmunología , Humanos , Valor Predictivo de las Pruebas , Sensibilidad y Especificidad

RESUMEN

We have previously reported that polymorphonuclear granulocyte (PMN) chemiluminescence (CL) and superoxide anion production are abnormally low in patients with polycythaemia vera (PV) after simulation with n-formyl-methionyl-leucyl-phenylalanine (fMLP), but normal when elicited by phorbol myristate acetate (PMA). This study documents that both fMLP and PMA induced CL was normal in PMN from patients with chronic myelogenous leukaemia (CML) and essential thrombocythaemia (ET). Furthermore, we monitored intracellular hydrogen peroxide (H2O2) production in PMN and monocytes from patients with PV, CML and ET by flow cytometry. H2O2 production in resting and PMA-stimulated cells was normal in all diseases. So also was fMLP induced H2O2 generation in ET PMN and monocytes. In contrast, fMLP-induced H2O2 production was significantly lower both in PV PMN (1.8 +/- 0.7 mean fluorescence intensity units in PV compared to 8.4 +/- 3.4 in healthy controls; P < 0.02), and in PV monocytes (0.3 +/- 0.5 compared to 2.5 +/- 0.7 in controls; P < 0.02). A less pronounced reduction of fMLP stimulated H2O2 production was noted in CML PMN (3.8 +/- 3.1 compared to 8.4 +/- 3.4 in controls; P < 0.05), and monocytes (1.3 +/- 0.6 compared to 2.5 +/- 0.7 in controls; P < 0.05). The reduction of H2O2 generation in PV and CML PMN was not attributed to subpopulations of less responsive cells. However, one ET and one CML patient showed a subpopulation of less responsive PMN. Thus intracellular H2O2 (as well as extracellular release of superoxide ions) is reduced in fMLP-stimulated PV PMN and monocytes but normal after PMA stimulation, a phenomenon that is not consistently found in other myeloproliferative disorders.

Asunto(s)

Peróxido de Hidrógeno/metabolismo , Monocitos/metabolismo , Trastornos Mieloproliferativos/metabolismo , Neutrófilos/metabolismo , Policitemia Vera/metabolismo , Adulto , Anciano , Humanos , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Mediciones Luminiscentes , Persona de Mediana Edad , N-Formilmetionina Leucil-Fenilalanina/metabolismo , Acetato de Tetradecanoilforbol/metabolismo , Trombocitemia Esencial/metabolismo

RESUMEN

We have measured the altered expression of the adhesion-promoting glucoproteins L-selectin and Mac-1 on monocytes and neutrophils after incubation with different concentrations of the chemotactic factor FMLP. We found a time- and dose-related down-regulation of L-selectin on both monocytes and neutrophils, which was more pronounced on neutrophils as compared to monocytes at both low (10(-12) M) and high (10(-7) M) concentrations of the chemotactic factor FMLP. Presence of human albumin reduced the down-regulation of L-selectin on both monocytes and neutrophils but to a greater extent on monocytes. Mac-1 expression increased in a time and dose-dependent manner on both monocytes and neutrophils when they were incubated with both low (10(-12) M) and high (10(-7) M) concentrations of FMLP, but the expression was more pronounced on neutrophils. The up-regulation of Mac-1 was equally reduced by albumin on monocytes and neutrophils. Presence of EDTA in RPMI medium reduced the FMLP-induced down-regulation of L-selectin on both monocytes and neutrophils, indicating that the down-regulation of L-selectin is partly divalent cation dependent. Differences in the altered expression of L-selectin and Mac-1 in monocytes and neutrophils with sustained expression of L-selectin on monocytes can contribute to the sequential recruitment of these cells into an inflammatory site.

Asunto(s)

Moléculas de Adhesión Celular/biosíntesis , Antígeno de Macrófago-1/biosíntesis , Monocitos/metabolismo , Neutrófilos/metabolismo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Albúminas/metabolismo , Supervivencia Celular , Regulación hacia Abajo , Humanos , Inflamación/metabolismo , Inflamación/patología , Selectina L , Persona de Mediana Edad , N-Formilmetionina Leucil-Fenilalanina/farmacología , Regulación hacia Arriba