Abstract
Objective and designTo determine whether the neurokinin-1 receptor (NK1R) plays a role in the activation of RBL-2H3 mast cells after FcεRΙ aggregation.Materials and methodsNK1R expression in RBL-2H3 cells was inhibited by small hairpin RNA (shRNA) against NK1R, and determined by western blotting. For activation, both NK1R knockdown and control RBL-2H3 cells were sensitized by dinitrophenol (DNP)-specific IgE and stimulated with the antigen DNP-bovine serum albumin (BSA). Following the activation of RBL-2H3 cells, monocyte chemoattractant protein (MCP-1) production and intracellular calcium flux were monitored by ELISA and confocal microscopy assay, respectively. For investigation of the signaling mechanism, phosphorylation of mitogen-activated protein kinases (MAPKs) after RBL-2H3 cell activation was assessed by western blotting.ResultsshRNA-NK1R mediated an effective inhibition of NK1R expression in RBL-2H3 cells. Protein production of MCP-1 was reduced by more than 55 % in NK1R knockdown RBL-2H3 cells compared with control RBL-2H3 cells. In addition, both calcium mobilization and phosphorylation levels of MAPKs (Erk1/2, JNK, and p38) after DNP-BSA stimulation (via FcεRΙ) were decreased due to the inhibition of NK1R expression.ConclusionNK1R is required for the activation of RBL-2H3 cells following FcεRΙ engagement and involved in the regulation of MAPK signaling pathways.Electronic supplementary materialThe online version of this article (doi:10.1007/s00011-012-0523-x) contains supplementary material, which is available to authorized users.
Highlights
Mast cells are major effectors in allergic diseases
We demonstrate that neurokinin-1 receptor (NK1R) promotes FceRI-evoked MCP-1 mRNA expression and calcium mobilization in RBL-2H3 cells
Our results suggest that NK1R is required for the initiation of rapid, full activation of mitogenactivated protein kinases (MAPKs) upon FceRI aggregation in RBL2H3 cells
Summary
Mast cells are major effectors in allergic diseases. The key receptor distributed on the surface of mast cells and responsible for their activation is the IgE receptor, FceRI [2]. In a typical allergic response, polyvalent antigens cross-link IgE-bearing FceRI molecules on mast cells. This causes the activation of immunoreceptor tyrosinebased activation motifs (ITAMs) in tails of FceRI and initiates intracellular signaling cascades that result in the activation of mast cells [1, 3]. Many signal molecules have been shown to be essential for FceRImediated mast cell activation, such as Src family kinases Lyn and Fyn [4,5,6,7]. The precise mechanism underlying mast cell activation remains far from fully understood
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