RESUMEN
To ensure normal immune function, mast cells employ different pathways to release mediators. Here, we report a thus far unknown capacity of mast cells to recycle and reuse secretory granules after an antigen-evoked degranulation process under physiological conditions; this phenomenon involves the existence of a recycling secretory granule pool that is available for release in a short time scale. Rapid endocytic modes contributed to the recycling of â¼60% of the total secretory granule population, which involved kiss-and-run and cavicapture mechanisms, causing retention of the intragranular matrix. We found the presence of normal-size granules and giant actomyosin- and dynamin-dependent granules, which were characterized by large quantal content. These large structures allowed the recovered mast cells to release a large amount of 5-HT, compensating for the decrease in the number of exocytosed secretory granules. This work uncovers a new physiological role of the exo-endocytosis cycle in the immunological plasticity of mast cells and reveals a new property of their biological secretion.
Asunto(s)
Degranulación de la Célula , Inmunoglobulina E/metabolismo , Mastocitos/fisiología , Fusión de Membrana , Vesículas Secretoras/metabolismo , Actinas/metabolismo , Animales , Antígenos/metabolismo , Calcimicina/farmacología , Degranulación de la Célula/efectos de los fármacos , Dinaminas/metabolismo , Técnicas Electroquímicas , Endocitosis/efectos de los fármacos , Exocitosis/efectos de los fármacos , Mastocitos/efectos de los fármacos , Fusión de Membrana/efectos de los fármacos , Ratones Endogámicos C57BL , Modelos Biológicos , Miosina Tipo II/metabolismo , Vesículas Secretoras/efectos de los fármacos , Serotonina/metabolismoRESUMEN
5-hydroxytriptamine (5-HT, serotonin) storage and release in mast cell (MC) secretory granules (SG) are dependent on serglycin proteoglycans (PG). This notion is based on the studies of MC of the connective tissue subtype that predominantly contain PG of the heparin type, whereas intestinal mucosal MC, which contain predominantly chondroitin sulfate, have been poorly explored. In the present study, we addressed the possibility that PG contents may differently affect the storage and release of preformed mediators in these two MC subclasses and explain in part their different functional properties. Rat peritoneal (PMC) and intestinal mast cells (IMC) were isolated and purified using a percoll gradient, and the efflux of 5-HT from each SG was measured by amperometric detection. IMC exhibited a â¼34% reduction in the release of 5-HT compared with PMC because of a lower number of exocytotic events, rather than a lower secretion per single exocytotic event. Amperometric spikes from IMC exhibited a slower decay phase and increased half-width but a similar ascending phase and foot parameters, indicating that the fusion pore kinetics are comparable in both MC subclasses. We conclude that both PG subtypes are equally efficient systems, directly involved in serotonin accumulation, and play a crucial role in regulating the kinetics of exocytosis from SG, providing specific secretory properties for the two cellular subtypes.
Asunto(s)
Mucosa Intestinal/metabolismo , Intestinos/citología , Mastocitos/metabolismo , Peritoneo/citología , Peritoneo/metabolismo , Serotonina/metabolismo , Animales , Células Cultivadas , Exocitosis/fisiología , Cinética , Mastocitos/citología , Tasa de Depuración Metabólica , Especificidad de Órganos/fisiología , Ratas , Ratas WistarRESUMEN
The key role of mast cells (MC), either in development of inflammatory pathologies or in response to environmental stress, has been widely reported in recent years. Previous studies have described the effects of corticotropin-releasing hormone (CRH), which is released from inflamed tissues by cellular stress signals, on MC degranulation, a process possibly driven by selective secretion of mediators (piecemeal degranulation). In this study, we introduce a novel granular exo-endocytic pathway induced by CRH on peritoneal MC. We found that CRH triggers substantial exocytosis, which is even stronger than that induced by Ag stimulation and is characterized by large quantal size release events. Membrane fluorescence increases during stimulation in the presence of FM1-43 dye, corroborating the strength of this exocytosis, given that discrete upward fluorescence steps are often observed and suggesting that secretory granules are preferentially released by compound exocytosis. Additionally, the presence of a depot of large tubular organelles in the cytoplasm suggests that the exocytotic process is tightly coupled to a fast compound endocytosis. This CRH-stimulated mechanism is mediated through activation of adenylate cyclase and an increase of cAMP and intracellular Ca(2+), as evidenced by the fact that the effect of CRH is mimicked by forskolin and 8-bromo-cAMP. Thus, these outcomes constitute new evidence for the critical role of MC in pathophysiological conditions within a cellular stress environment and an alternative membrane trafficking route mediated by CRH.