6-ketoprostaglandin-f1-alpha and ramiprilat

Angiotensin-converting enzyme (ACE) inhibitors elicit outside-in signaling via ACE in endothelial cells. This involves the CK2-mediated phosphorylation of ACE on Ser1270 and the activation of the c-Jun N-terminal kinase (JNK)/c-Jun pathway, resulting in an enhanced endothelial ACE expression. Because cyclooxygenase-2 (COX-2) expression is reported to be increased in subjects treated with ACE inhibitors, we determined the role of ACE signaling in this phenomenon and the transcription factors involved. In lungs from mice treated with the ACE inhibitor ramipril for 5 days, COX-2 expression was increased. A similar (1.5- to 2-fold) increase in COX-2 protein was detected in primary cultures of human endothelial cells treated with ramiprilat. In an endothelial cell line stably expressing human somatic ACE, ramiprilat increased COX-2 promoter activity, an effect not observed in ACE-deficient cells or cells expressing a nonphosphorylatable ACE mutant (S1270A). The ramiprilat-induced, ACE-dependent increase in COX-2 expression and promoter activity (both 1.5- to 2-fold greater than control) was prevented by the inhibition of JNK. Ramiprilat significantly enhanced the DNA binding activity of activator protein-1 in cells expressing ACE but not S1270A ACE. Activator protein-1 decoy oligonucleotides prevented the ACE inhibitor-induced increase in COX-2 promoter activity and protein expression. As a consequence of the ramiprilat-induced increase in COX-2 expression, prostacyclin and prostaglandin E2, but not thromboxane A2, production was increased and was inhibited by the COX-2 inhibitor celecoxib. These results indicate that ACE signaling may underlie the increase in COX-2 and prostacyclin levels in patients treated with ACE inhibitors.

Topics: 6-Ketoprostaglandin F1 alpha; Amino Acid Substitution; Angiotensin-Converting Enzyme Inhibitors; Animals; Anthracenes; Aorta; Binding, Competitive; Celecoxib; Cells, Cultured; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Cyclooxygenase Inhibitors; Dinoprostone; DNA; Endothelial Cells; Endothelium, Vascular; Enzyme Induction; Humans; JNK Mitogen-Activated Protein Kinases; Lung; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Mutation, Missense; Oligodeoxyribonucleotides; Peptidyl-Dipeptidase A; Phosphorylation; Promoter Regions, Genetic; Prostaglandin-Endoperoxide Synthases; Prostaglandins I; Protein Processing, Post-Translational; Pyrazoles; Ramipril; Recombinant Fusion Proteins; Signal Transduction; Sulfonamides; Sus scrofa; Thromboxane A2; Transcription Factor AP-1; Transcription, Genetic; Transfection; Umbilical Veins

We studied whether primary cultured porcine brain capillary endothelial cells (PBCEC) respond to bradykinin with an enhanced intracellular cytosolic calcium concentration [Ca2+]i with subsequent formation of nitric oxide (NO) and prostacyclin (PGI2). In addition we examined whether these cells synthetize and release kinins that may accumulate during angiotensin-converting enzyme (ACE) inhibition. [Ca2+]i was assessed by the fluorescent dye Fura-2, NO formation by determination of intracellular cyclic GMP and PGI2 by a specific radioimmunoassay for 6-ketoprostaglandin F1 alpha. Bradykinin and the ACE inhibitor ramiprilat concentration-dependently increased the formation of cyclic GMP which was completely prevented by the stereospecific inhibitor of NO synthase, NG-nitro-L-arginine. Also the specific B2-kinin receptor antagonist icatibant (Hoe 140) abolished the increase in cyclic GMP as well as the ramiprilat-induced increase in PGI2 formation. The data demonstrate the existence of B2-kinin receptors and ACE activity in PBCEC. Moreover PBCEC are capable of producing and releasing kinins in amounts that lead via stimulation of B2-kinin receptors to an enhanced [Ca2+]i as well as NO and PGI2 synthesis and release, provided that degradation of kinins is prevented by inhibition of endothelial ACE activity.

Topics: 1-Methyl-3-isobutylxanthine; 6-Ketoprostaglandin F1 alpha; Angiotensin-Converting Enzyme Inhibitors; Animals; Arginine; Bradykinin; Calcium; Capillaries; Cells, Cultured; Cerebrovascular Circulation; Cyclic GMP; Cytosol; Dose-Response Relationship, Drug; Endothelium, Vascular; Epoprostenol; Fura-2; Kinetics; Nitric Oxide; Nitroarginine; Ramipril; Swine

The effects of angiotensin-converting enzyme (ACE) inhibitors on endothelial autacoid formation were determined in human cultured endothelial cells and in endothelium-intact bovine coronary arteries under resting conditions and after stimulation with bradykinin. Incubation of cultured human endothelial cells with moexiprilat or ramiprilat (0.3 microM) caused a maintained increase in resting intracellular calcium [Ca2+]i, which was prevented by the selective B2-receptor antagonist Hoe 140 (0.1 microM). Both ACE inhibitors also significantly enhanced the increase in [Ca2+]i elicited by bradykinin (3 nM). In parallel with their effect on resting [Ca2+]i, moexiprilat and ramiprilat both induced an increase in intracellular cyclic GMP (cGMP). This increase was prevented by Hoe 140 (0.1 microM) and was abolished by NG-nitro-L-arginine (30 microM), indicating a kinin-induced nitric oxide (NO) formation in this response. The elevation in [Ca2+]i also led to an enhanced production of prostacyclin (PGI2), as indicated by an increase in the concentration of 6-keto prostaglandin F1 alpha (PGF1 alpha) in the cell supernatant. Similar effects of the ACE inhibitors on endothelial autacoid production were observed in endothelium-intact bovine coronary arteries. Like bradykinin (30 nM), moexiprilat (0.3 microM) elicited a nearly twofold increase in the cGMP content of these arteries, which was abolished by both NG-nitro-L-arginine and removal of the endothelium. The functional consequences of this ACE inhibitor-induced increase in vascular cGMP were reflected by a distinct relaxation of arteries preconstricted with PGF2 alpha.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: 6-Ketoprostaglandin F1 alpha; Angiotensin-Converting Enzyme Inhibitors; Animals; Arginine; Bradykinin; Calcium; Cattle; Cells, Cultured; Coronary Vessels; Cyclic GMP; Endothelium, Vascular; Epoprostenol; Humans; Isoquinolines; Nitric Oxide; Nitroarginine; Ramipril; Tetrahydroisoquinolines; Umbilical Veins; Vasodilation