A LUCIFERASE-ENGINEERED CELL-LINE FOR STUDY OF CAMP REGULATION IN ENDOTHELIAL-CELLS

cAREL is a cAMP-responsive endothelial cell line carrying a luciferase reporter gene introduced by stable transfection of a luciferase enhan cer trap into rabbit aortic endothelial cells. Luciferase gene express ion in cAREL was stimulated 233-fold by 8-BrcAMP. Treatment with the b eta-adrenoceptor agonist isoproterenol induced a 7.0-fold increase in luciferase expression, which was partially blocked by either beta(1)- or beta(2)-adrenoceptor antagonists and totally blocked by propranolol and by a combination of beta(1)- plus beta(2)-adrenoceptor antagonist s. Receptor stimulation was mimicked by cholera toxin, forskolin, 8-Br cAMP, and isobutylmethylxanthine but not by 8-BrcGMP, dexamethasone, o r phorbol 12-myristate 13-acetate. Stimulation by isoproterenol was co mpletely blocked by H-89, a protein kinase A inhibitor. cAREL was also stimulated by A-23187, and this effect was abrogated by EGTA and H-89 . cAREL is the first cAMP-sensitive endothelial cell line described, a nd it can be useful as a positive control, as a model for cAMP regulat ion, as a background to genetic introduction of receptors, as an indic ator of intracellular pathway activation, and as a tool to investigate cAMP effects on other signaling pathways.