Methiothepin downregulates SNAP‐23 and inhibits degranulation of rat basophilic leukemia cells and mouse bone marrow‐derived mast cells

Abstract

AbstractIn the present study, we found that methiothepin (a nonselective 5‐hydroxytryptamine [5‐HT] receptor antagonist) inhibited antigen‐induced degranulation in rat basophilic leukemia cells and mouse bone marrow‐derived mast cells. Although antigen stimulation induces release of histamine and serotonin (5‐HT) by exocytosis and mast cells express several types of 5‐HT receptor, the detailed role of these receptors remains unclear. Here, pretreatment of cells with methiothepin attenuated increased intracellular Ca2+ concentration, phosphorylated critical upstream signaling components (Src family tyrosine kinases, Syk, and PLCγ1), and suppressed TNF‐α secretion via inhibition of Akt (a Ser/Thr kinase activated by PI3K)and ERK phosphorylation. Furthermore, it inhibited PMA/ionomycin‐induced degranulation; this finding suggested that methiothepin affected downstream signaling. IκB kinase β phosphorylates synaptosomal associated protein 23, which regulates the fusion events of the secretory granule/plasma membrane after mast cell activation, resulting in degranulation. We showed that methiothepin blocked PMA/ionomycin‐induced phosphorylation of synaptosomal associated protein 23 by inhibiting its interaction with IκB kinase β. Together with the results of selective 5‐HT antagonists, it is suggested that methiothepin inhibits mast cell degranulation by downregulating upstream signaling pathways and exocytotic fusion machinery through mainly 5‐HT1A receptor. Our findings provide that 5‐HT antagonists may be used to relieve allergic reactions.