ATP-evoked Ca2±waves Stimulate Gene Expression In Human Airway Fibroblasts

We sought to investigate the effects of a variety of autacoids on Ca2+-handling in human airway fibroblasts. Primary cultured fibroblasts were loaded with the Ca2+-indicator dye fluo-4 and studied using confocal fluorimetric microscopy. ATP (10−5 M) evoked recurring Ca2+-waves . This fluorimetric change was greater and longer lasting within the nucleus of the cell than in the non-nuclear portion of the cytosol, and was only sometimes accompanied by a contraction. These responses were completely occluded by cyclopiazonic acid (10−5 M; depletes the internal Ca2+-store) or the phospholipase C inhibitor U73122 (10−6 M). Pretreatment of the cells with ryanodine (10−5 M), on the other hand, had no effect on the ATP-evoked responses. With respect to the receptor through which this response was exerted, we found it to be mimicked by UTP or ADP but not by adenosine or α,β-methylene-ATP, and to be blocked by the purinergic receptor blocker PPADS; interestingly, PPADS itself appeared to sometimes evoke a rise in [Ca2+]i on its own. ATP also evoked a membrane conductance change with characteristics of a non-selective cation current, markedly enhanced synthesis of the cytokine TGFβ and the matrix proteins fibronectin and collagen I; these changes in protein synthesis were blocked by PPADs and were partially reduced by ryanodine. We conclude that, in human pulmonary fibroblasts, ATP acts upon P2Y receptors to liberate internal Ca2+ through ryanodine-insensitive channels, leading to a Ca2+-wave which courses throughout the cell and triggers protein synthesis.This work was supported by the Canadian Institutes of Health Research