Epidermal growth factor induces intracellular Ca2+ oscillations in microvascular endothelial cells

An increase in intracellular Ca2+ concentration ([Ca2+]) may play a role in the proliferative effect of several growth factors. In this study, the changes in [Ca2+](i) elicited by epidermal growth factor (EGF) in rat cardiac microvascular endothelial cells (CMEC) have been investigated by using fura-2 conventional and confocal microscopy. A large heterogeneity in the latency and in the pattern of the Ca2+ response was found at each dose of EGF (2.5-100 ng/ml), whereas some cells displayed a non-oscillatory behavior and others exhibited a variable number of Ca2+ oscillations. On average, the fraction of responsive cells, the total number of oscillations and the duration of the Ca2+ signal were higher at around 10 ng/ml EGF, while there was no dose-dependence in the lag time and in the amplitude of the [Ca2+](i) increase. EGF-induced Ca2+ spikes were abolished by the tyrosine kinase inhibitor genistein, but not by its inactive analogue daidzein, and by the phospholipase C blocker NCDC. Only 1-2 transients could be elicited in Ca2+-free solution, while re-addition of extracellular Ca2+ recovered the spiking activity. The oscillatory signal was prevented by the SERCA inhibitor thapsigargin and abolished by the calcium entry blockers Ni2+ and La3+. Moreover, EGF-induced Ca2+ transients were abolished by the InsP(3) receptor blocker caffeine, while ryanodine was without effect. Confocal imaging microscopy showed that the Ca2+ response to EGF was localized both in the cytoplasm and in the nucleus. We suggest that EGF-induced [Ca2+](i) increase may play a role in the proliferative action of EGF on endothelial cells. (C) 2002 Wiley-Liss, Inc.