Detection of Low-Molecular-Weight Mast Cell-Activating Factors in Serum From Patients With Chronic Spontaneous Urticaria.

BACKGROUND AND OBJECTIVES: Functionally active autoantibodies to IgE and to the high-affinity IgE receptor (FcεRI) can be detected in serum in about 40% of patients with chronic spontaneous urticaria (CSU). Recent studies showed that serum from patients with CSU can induce activation of mast cells, irrespective of whether they carry high-affinity IgE receptors. To evaluate mast cell activation induced by factors in the serum of CSU patients with a molecular weight lower than that of autoantibodies. METHODS: Eight CSU patients and 5 healthy controls were evaluated. Whole serum and serum fractionated at 100, 50, and 30 kDa were used to stimulate in vitro LAD2 mast cells. The enzymatic activity of ß-hexosaminidase was evaluated in supernatants and cell pellets as a measure of mast cell degranulation. RESULTS: Mean (SEM) release of mast cell ß-hexosaminidase induced by whole serum from CSU patients was higher than that induced by serum from the healthy controls (14.4 [2.7%] vs 5.1 [2.4%]; P=.027). In addition, serum fractions below 100 kDa and below 50 kDa from CSU patients induced mast cell degranulation that was significantly higher than that induced by the corresponding fractions in sera from healthy controls (10.2% [1.4%] vs 3.8% [1.9%] [P=.024] and 10.1% [1.2%] vs 3.9% [1.7%] [P=.012], respectively). In 4 CSU patients, we evaluated serum fractions <30 kDa, which retained their capacity to activate mast cells (11.0% [0.7%]). CONCLUSIONS: This study shows that sera from CSU patients may contain low-molecular-weight mast cell-activating factors other than autoantibodies. These factors could be an additional mechanism contributing to the pathogenesis of CSU.

Detection of low-molecular-weight mast cell-activating factors in serum from patients with chronic spontaneous urticaria

Background: Functionally active autoantibodies to IgE and to the high-affinity IgE receptor (FcεRI) can be detected in serum in about 40% of patients with chronic spontaneous urticaria (CSU). Recent studies showed that serum from patients with CSU can induce activation of mast cells, irrespective of whether they carry high-affinity IgE receptors. Objective: To evaluate mast cell activation induced by factors in the serum of CSU patients with a molecular weight lower than that of autoantibodies. Methods: Eight CSU patients and 5 healthy controls were evaluated. Whole serum and serum fractionated at 100, 50, and 30 kDa were used to stimulate in vitro LAD2 mast cells. The enzymatic activity of β-hexosaminidase was evaluated in supernatants and cell pellets as a measure of mast cell degranulation. Results: Mean (SEM) release of mast cell β-hexosaminidase induced by whole serum from CSU patients was higher than that induced by serum from the healthy controls (14.4 [2.7%] vs 5.1 [2.4%]; P =.027). In addition, serum fractions below 100 kDa and below 50 kDa from CSU patients induced mast cell degranulation that was significantly higher than that induced by the corresponding fractions in sera from healthy controls (10.2% [1.4%] vs 3.8% [1.9%] [P=.024] and 10.1% [1.2%] vs 3.9% [1.7%] [P=.012], respectively). In 4 CSU patients, we evaluated serum fractions <30 kDa, which retained their capacity to activate mast cells (11.0% [0.7%]). Conclusions: This study shows that sera from CSU patients may contain low-molecular-weight mast cell-activating factors other than autoantibodies. These factors could be an additional mechanism contributing to the pathogenesis of CSU (AU)

Introducción: Los autoanticuerpos IgE funcionalmente activos y los receptores de alta afinidad para la IgE (FcεRI) pueden ser detectados n el suero de aproximadamente un 40% de los pacientes con urticaria crónica espontánea (UCE). Estudios recientes muestran que el uero de estos pacientes puede inducir activación de mastocitos con o sin receptores de alta afinidad para la IgE. Objetivo: El objetivo de este estudio fue evaluar la actividad de los factores séricos de los sueros de pacientes con UCE con un peso molecular inferior al de los autoanticuerpos. Métodos: Para ello se estudiaron 8 pacientes con UCE y 5 controles sanos. El suero completo de cada uno de ellos y el fraccionado a 100,50 y 30 kDA se utilizó para estimular in vitro mastocitos LAD2. La actividad enzimática de la β-hexosaminidasa se determinó en los sobrenadantes y en el botón celular con el fin de cuantificar la degranulación mastocitaria. Resultados: En cuanto a los resultados obtenidos se observó una liberación de β-hexosaminidasa mastocitaria inducida por los sueros completos de los pacientes con UCE (14,4±2,7%, media ± EE de la media) significativamente mayor que la inducida por sueros de controles sanos (5,1±2,4%; p=0,027). Así mismo, las fracciones séricas inferiores a 100 kDa e inferiores a 50 kDa de los pacientes con UCE indujeron degranulación mastocitaria significativamente superior a la inducida por las fracciones correspondientes de sueros controles (10,2±1,4% vs 3,8±1,9% [p=0,024] and 10,1±1,2% vs 3,9±1,7% [p=0,012], respectivamente). En 4 pacientes con UCE observamos que las fracciones inferiores a 30 kDa mantenían la capacidad de activar a los mastocitos (11,0±0,7%). Conclusiones: En conclusión, este estudio muestra que el suero de los pacientes con UCE puede contener factores de bajo peso molecular diferentes a los autoanticuerpos que son capaces de activar a los mastocitos. Este hallazgo podría contribuir a conocer los mecanismos de la patogénesis de la UCE (AU)

Mast cells are critically involved in serum-mediated vascular leakage in chronic urticaria beyond high-affinity IgE receptor stimulation.

BACKGROUND: Chronic urticaria (CU) is one of the most common skin disorders whose pathogenic mechanisms are not fully clarified. Autoimmune aetiology can be ascribed to 45% of patients with CU, and basophil histamine release is positive in 40% of cases. Our aim was to use a novel approach to evaluate the serum permeabilizing effect to identify the mediators of endothelial cell (EC) leakage and to define the role of mast cells (MCs) in the process. METHODS: Permeabilizing activity of sera from 19 patients with CU and 11 healthy blood donors was evaluated by measuring serum-induced degranulation of two MC lines, expressing (LAD2) or lacking (HMC-1) the IgE receptor. Mast cell supernatant (SN) was then incubated with an EC monolayer, and endothelial permeability was evaluated by Fluorescein isothiocyanate-bovine serum albumin leakage in a transwell system. RESULTS: All 19 patient sera failed to induce direct EC leakage, but 15/19 and 17/19 promoted degranulation of HMC-1 and LAD2, respectively. Interestingly, 85% of autologous serum skin test-negative sera were able to cause MC degranulation. Also, 17/19 SNs from HMC-1 and all SNs from LAD2 incubated with CU sera increased endothelial permeability. Endothelial cell leakage remained unchanged after Ig depletion and was prevented by antihistamine, platelet-activating factor or leukotriene antagonist. CONCLUSIONS: Our study shows that CU sera are able to degranulate MCs through an IgE- and IgG-independent mechanism. The nature of histamine-releasing factors involved is still unclear, but our finding opens new ways to the understanding of the pathogenesis of CU, particularly in patients not showing circulating autoantibodies to FcεRI or IgE.