Identification of a New Exo-Endocytic Mechanism Triggered by Corticotropin-Releasing Hormone in Mast Cells

Abstract

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 Ca2+, 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.

Transient fusion ensures granule replenishment to enable repeated release after IgE-mediated mast cell degranulation

“…Cultures were maintained in glutamine (4 mM) and FBS (5% v/v) (BioWest) Advanced RPMI 1640 medium (Gibco) containing penicillin (50 U/ml) and streptomycin (50 µg/ml) (Gibco), stem cell factor (30 ng/ml) and IL-3 (10 ng/ml) (PeproTech) at 37°C under a 5% CO 2 humidified atmosphere. To induce antigen stimulation, mast cells were sensitized with mouse monoclonal anti-DNP IgE (1 µg/ml) (Sigma) in medium for 24 h (Balseiro-Gomez et al, 2015). For FM1-43 (Invitrogen) loading through spontaneous activity, mast cells were incubated in FM1-43 (4 μM) in basal Locke solution for 60 min (at 37°C under a 5% CO 2 humidified atmosphere).…”

“…Detection of 5-HT release from secretory granules was performed as previously described elsewhere (Alvarez de Toledo et al, 1993;Balseiro-Gomez et al, 2015). Before amperometry recording, cells were incubated in basal Locke solution containing 5-HT (1 mM) for 30 min at 37°C.…”

Frontline Science: Corticotropin-releasing factor receptor subtype 1 is a critical modulator of mast cell degranulation and stress-induced pathophysiology

“…3D-F). The range of concentrations used for CRF were based on the affinity of CRF for CRF 1 and CRF 2 [35,36] and the ranges used previously in cultured MCs [22,24,37] and in tissue physiology experiments in humans and animals [38,39].…”

High glucocorticoid levels during gestation activate the inflammasome in hippocampal oligodendrocytes of the offspring

“…The biological actions of CRH and Ucn II (urocortin II, a member of the CRH peptide family) are mediated by CRH receptor 1 (CRHR1) and CRHR2, which are expressed in mast cells, microglia, astrocytes, and neurons (Stevens et al, ; Wang et al, ; Alysandratos et al, ; Tillinger et al, ). These receptors are involved in the increase of proinflammatory cytokine production (Stevens et al, ) and mediate rapid mast cell degranulation induced by CRH (Theoharides et al, ; Balseiro‐Gomez et al, ). An alternative source of CRH are stress‐ or CRH/urocortin‐activated mast cells, which have been proposed to be potential sources of CRH and urocortin that may act as autocrine and paracrine signals (Kempuraj et al, ).…”

“…CRH secreted by the hypothalamus is a key modulator of ACTH release in the HPA axis (Evanson and Herman, 2015), and is critically involved in neuroinflammatory responses to stress (Balseiro-Gomez et al, 2015). The biological actions of CRH and Ucn II (urocortin II, a member of the CRH peptide family) are mediated by CRH receptor 1 (CRHR1) and CRHR2, which are expressed in mast cells, microglia, astrocytes, and neurons (Stevens et al, 2003;Wang et al, 2007;Alysandratos et al, 2012;Tillinger et al, 2013).…”