Role of cyclic nucleotides in the regulation of endothelin-1 production by human endothelial cells.
The regulation of endothelin-1 (ET-1) production by endothelial cells is likely of crucial physiological importance in the maintenance of vascular homeostasis. The aim of the present study was to explore the possible role of cyclic nucleotides in the control of ET-1 production in human umbilical vein endothelial cells (HUVEC). ET-1 release was determined by measuring levels of immunoreactive ET-1 in HUVEC-conditioned media after 6-h incubations. In the presence of 10% fetal calf serum (FCS) there was a threefold increase in ET-1 release compared with serum-free conditions (1.96 +/- 0.17 vs. 0.56 +/- 0.06 pg/micrograms protein), respectively. Inhibition of protein kinase (PK) C using staurosporine (10 nM) reduced basal ET-1 release by approximately 50% and completely prevented the response to FCS. In contrast, the addition of other PK inhibitors had little effect on basal or serum-stimulated ET-1 release at the concentrations used. N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (DBcAMP) produced significant alterations in ET-1 release depending on the basal level of production. Under serum-free conditions of low basal ET-1 production, DBcAMP increased ET-1 release by 68 +/- 22% but only at the highest concentration studied (1 mM). The dose-response relationship for DBcAMP was potentiated by KT-5720 (0.1 microM), an inhibitor of PKA, with a significant shift to 10-fold lower concentrations, whereas it was blocked by KT-5823 (4 microM), which can inhibit PKG.(ABSTRACT TRUNCATED AT 250 WORDS)
Stewart
D J
DJ
Department of Medicine, McGill University, Montreal, Quebec, Canada.
Cernacek
P
P
Mohamed
F
F
Blais
D
D
Cianflone
K
K
Monge
J C
JC
D000818:Animals; D001483:Base Sequence; D001920:Bradykinin / Q000494:pharmacology; D000242:Cyclic AMP / Q000494:pharmacology; D004075:Diglycerides / Q000378:metabolism; D019332:Endothelin-1; D016232:Endothelins / Q000096:biosynthesis* / Q000235:genetics; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D009712:Nucleotides, Cyclic / Q000494:pharmacology / Q000502:physiology*; D015345:Oligonucleotide Probes / Q000235:genetics; D047428:Protein Kinase Inhibitors; D011498:Protein Precursors / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013917:Thrombin / Q000494:pharmacology
D004075:Diglycerides; D019332:Endothelin-1; D016232:Endothelins; D009712:Nucleotides, Cyclic; D015345:Oligonucleotide Probes; D047428:Protein Kinase Inhibitors; D011498:Protein Precursors; D012333:RNA, Messenger; D000242:Cyclic AMP; D013917:Thrombin; D001920:Bradykinin