Epidermal growth factor induces intracellular Ca2+ oscillations in microvascular endothelial cells.
An increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) may play a role in the proliferative effect of several growth factors. In this study, the changes in [Ca(2+)](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 Ca(2+) 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 Ca(2+) oscillations. On average, the fraction of responsive cells, the total number of oscillations and the duration of the Ca(2+) 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 [Ca(2+)](i) increase. EGF-induced Ca(2+) 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 Ca(2+)-free solution, while re-addition of extracellular Ca(2+) recovered the spiking activity. The oscillatory signal was prevented by the SERCA inhibitor thapsigargin and abolished by the calcium entry blockers Ni(2+) and La(3+). Moreover, EGF-induced Ca(2+) transients were abolished by the InsP(3) receptor blocker caffeine, while ryanodine was without effect. Confocal imaging microscopy showed that the Ca(2+) response to EGF was localized both in the cytoplasm and in the nucleus. We suggest that EGF-induced [Ca(2+)](i) increase may play a role in the proliferative action of EGF on endothelial cells.
Moccia
Francesco
F
Department of Physiological and Pharmacological Sciences, University of Pavia, Pavia, Italy. moccia@alpha.szn.it
Berra-Romani
Roberto
R
Tritto
Simona
S
Signorelli
Silvia
S
Taglietti
Vanni
V
Tanzi
Franco
F
D000200:Action Potentials / Q000187:drug effects; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism* / Q000502:physiology; D004815:Epidermal Growth Factor / Q000008:administration & dosage / Q000494:pharmacology*; D005110:Extracellular Space / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism*; D008833:Microcirculation; D009991:Oscillometry; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D012433:Ryanodine / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D013873:Thionucleotides / Q000494:pharmacology; D010738:Type C Phospholipases / Q000502:physiology; D014544:Uridine Triphosphate / Q000494:pharmacology
D013873:Thionucleotides; D012433:Ryanodine; D002110:Caffeine; D004815:Epidermal Growth Factor; D019284:Thapsigargin; D000255:Adenosine Triphosphate; D011505:Protein-Tyrosine Kinases; D010738:Type C Phospholipases; D002118:Calcium; D014544:Uridine Triphosphate; C008501:2-methylthio-ATP