Orai and TRPC Channel Contribution to Calcium Signaling in Human Mast Cells

Abstract

Mast cells are important targets for the treatment of allergic diseases and anaphylaxis. Inappropriate and chronic activation of mast cells via the IgE receptor leads to the release of a range of pre-stored and newly synthesized inflammatory mediators and the symptoms of disease. Calcium influx is a critical regulator of mast cell signaling, with influx through ion channels absolutely required not only for the exocytosis of preformed mediators but also to direct the synthesis of eicosanoids, cytokines and chemokines.Studies in rodent and human mast cells have identified STIM-regulated Orai channels to be key players in initiating calcium influx and degranulation in antigen-stimulated mast cells; in rodent mast cells, a role for TRPC channels is also emerging. Here we report evidence for a role of TRPC channels in human mast cell signaling. Using gene microarray analysis, we find evidence for expression of multiple TRP family members in primary human lung mast cells and LAD2 cells; the expression of TRPC1 and TRPC6 was further confirmed by immunocytochemistry. Single cell fura-2 imaging experiments, showed that barium could partially substitute for calcium to support influx following antigen- and thapsigargin stimulation, consistent with a contribution of non-selective TRPC-like channels to store operated calcium entry in human mast cells. To investigate the potential involvement of TRPC1 in STIM-regulated store operated calcium signaling in human mast cells, LAD2 cells were transfected with the STIM1 (684KK685) TRPC1 gating mutant. Expression of the mutant, however failed to alter calcium signaling in either thapsigargin or antigen-stimulated cells. Further pharmacological and molecular experiments are currently being performed to further evaluate the putative role of other TRPC channels in human mast cell signaling.