β-Defensins Activate Human Mast Cells via Mas-Related Gene X2

Abstract

Human β-defensins (hBDs) stimulate degranulation in rat peritoneal mast cells in vitro and cause increased vascular permeability in rats in vivo. Here, we sought to determine if hBDs activate murine and human mast cells and to delineate the mechanisms of their regulation. hBD2 and hBD3 did not induce degranulation in murine peritoneal or bone marrow-derived mast cells (BMMC) in vitro and had no effect on vascular permeability in vivo. By contrast, these peptides induced sustained Ca2+ mobilization and substantial degranulation in human mast cells with hBD3 being more potent. Pertussis toxin (PTx) had no effect on hBD-induced Ca2+ mobilization but La3+ and 2-Aminoethoxydiphenyl borate (2-APB; a dual inhibitor of IP3 receptor and transient receptor potential (TRP) channels caused substantial inhibition of this response. Interestingly, degranulation induced by hBDs was substantially inhibited by PTx, La3+ or 2-APB. While human mast cells endogenously express G protein coupled receptor (GPCR); Mas-related gene X2 (MrgX2), rat basophilic leukemia, RBL-2H3 cells and murine BMMCs do not. Silencing the expression of MrgX2 in human mast cells inhibited hBD-induced degranulation but had no effect on anaphylatoxin C3a-induced response. Furthermore, ectopic expression of MrgX2 in RBL-2H3 and murine BMMCs rendered these cells responsive to hBDs for degranulation. This study demonstrates that hBDs activate human mast cells via MrgX2, which couples to both PTx sensitive and insensitive signaling pathways likely involving Gαq and Gαi to induce degranulation. Furthermore, murine mast cells are resistant to hBDs for degranulation and this reflect the absence of MrgX2 in these cells.

Pharmacology and Signaling of MAS-Related G Protein–Coupled Receptors

“…In line with this observation, proadrenomedullin peptides that endogenously occur as side products during adrenomedullin synthesis in the adrenal medulla have also been shown to activate MRGPRX2 (Kamohara et al, 2005). MRGPRX2 were also shown to be involved in mast cell activation by a set of structurally similar, endogenous (Tatemoto et al, 2006;Subramanian et al, 2011aSubramanian et al, , 2013 or exogenous (Kashem et al, 2011;Subramanian et al, 2011b) basic secretagogues (see section III.G.2.d). Finally, in concordance with murine MRGPRC, the human PAR2-derived "SLIGKV"-peptide was shown to activate MRGPRX2 after expression in CHO cells at a concentration of 20 mM .…”

“…Different groups agree that activation of G q/11 proteins is involved in this process (Lembo et al, 2002;Robas et al, 2003;Kamohara et al, 2005;Breit et al, 2006;Burstein et al, 2006). However, apart from its G q/11 coupling, additional coupling to PTX-sensitive G i/o proteins was described for MRGPRX1 (Chen and Ikeda, 2004;Gales et al, 2005;Burstein et al, 2006) or -2 (Kamohara et al, 2005;Subramanian et al, 2013). According to their expression pattern, MRGPRX1 and -2 may be involved in somatosensory detector functions and plasticity of primary sensory neurons (see sections III.G.2.b and III.G.2.c) and in mast cell biology (see section III.G.2.d).…”

“…Although MRGPRX2 are not involved in mast cell activation provoked by endogenous anaphylatoxins, mast cell degranulation induced by a synthetic C5a receptor antagonist (PMX-53) or a C3a receptor superagonist (E7) required MRGPRX2 expression (Kashem et al, 2011;Subramanian et al, 2011b). Later on during infection, epithelial cells can secrete b-defensins or cathelicidins, both inducing mast cell degranulation via MRGPRX2 (Subramanian et al, 2011a(Subramanian et al, , 2013. Hence, MRGPRX2 can integrate paracrine input from various cell types by detecting alterations of the local milieu and inducing mast cell degranulation.…”

“…However, MRGPRX2 activation by LL-37 or human b-defensin-3 involves two different signaling cascades that act synergistically on degranulation, but only one of which depends on G i/o -induced PKC activation (Subramanian et al, 2011a(Subramanian et al, , 2013. The second PTX-insensitive pathway involved calcium influx and release, as suggested by the inhibitory actions of the calcium release-activated calcium channel blocker, lathanium (La 3+ ), and the inositol 1,4,5-trisphosphate receptor and canonical TRP blocker, 2-aminoethoxydiphenyl borate (Subramanian et al, 2011a(Subramanian et al, , 2013.…”

Targeting IL-33 in Autoimmunity and Inflammation

“…Despite IL-33 sharing the IL-1 receptor accessory protein with other members of the IL-1 family that are known to upregulate an inducible antimicrobial peptide human b-defensin 2 (hBD2), IL-33 downregulates serum-induced hBD2 in human primary keratinocytes; this finding may explain the increased colonization rate of Staphylococcus aureus seen in atopic dermatitis (AD) patients (Alase et al, 2012). Interestingly, hBD2 has been reported to activate human MCs (Subramanian et al, 2013).…”