CRACM/Orai ion channel expression and function in human lung mast cells

摘要

Background: Influx of extracellular Ca 2+ into human lung mast cells (HLMCs) is essential for the FcεRI-dependent release of preformed granule-derived mediators and newly synthesized autacoids and cytokines. However, the identity of the ion channels underlying this Ca 2+ influx is unknown. The recently discovered members of the CRACM/Orai ion channel family that carries the Ca 2+ release-activated Ca 2+ current are candidates. Objectives: To investigate the expression and function of CRACM channels in HLMCs. Methods: CRACM mRNA, protein, and functional expression were examined in purified HLMCs and isolated human bronchus. Results: CRACM1, -2, and -3 mRNA transcripts and CRACM1 and -2 proteins were detectable in HLMCs. A CRACM-like current was detected following FcεRI-dependent HLMC activation and also in HLMCs dialyzed with 30 μM inositol triphosphate. The Ca 2+-selective current obtained under both conditions was blocked by 10 μM La 3+ and Gd 3+, known blockers of CRACM channels, and 2 distinct and specific CRACM-channel blockers - GSK-7975A and Synta-66. Both blockers reduced FcεRI-dependent Ca 2+ influx, and 3 μM GSK-7975A and Synta-66 reduced the release of histamine, leukotriene C 4, and cytokines (IL-5/-8/-13 and TNFα) by up to 50%. Synta-66 also inhibited allergen-dependent bronchial smooth muscle contraction in ex vivo tissue. Conclusions: The presence of CRACM channels, a CRACM-like current, and functional inhibition of HLMC Ca 2+ influx, mediator release, and allergen-induced bronchial smooth muscle contraction by CRACM-channel blockers supports a role for CRACM channels in FcεRI-dependent HLMC secretion. CRACM channels are therefore a potential therapeutic target in the treatment of asthma and related allergic diseases.