Inhibitory effect of BQ-123 on endothelin-1-stimulated mitogen-activated protein kinase and cell growth of rat vascular smooth muscle cells.
Studies have suggested that endothelin-1 (ET-1) activates cellular contraction and proliferation in rat vascular smooth muscle cells (VSMC). We examined whether an ET(A) receptor antagonist, BQ-123, inhibits the following cellular actions of ET-1 in rat VSMC: cytosolic free calcium ([Ca2+]i) mobilization, cellular contraction, mitogen-activated protein (MAP) kinase activation, [3H]thymidine incorporation, and MTT reduction. [Ca2+]i was measured by the fluorescent method. MAP kinase activity was measured by the phosphorylation of a synthetic peptide of a specific substrate for MAP kinase. The specificity of BQ-123 for ET-1-activated MAP kinase was checked by Western blotting analysis. [3H]thymidine incorporation and MTT reduction studies were performed using the cells incubated for 12 h with serum-free medium containing effectors. BQ-123 inhibited ET-1 receptor binding and blocked [Ca2+]i mobilization, cellular contraction, MAP kinase activation, [3H]thymidine incorporation, and MTT reduction in response to ET-1. BQ-123 did not affect the arginine vasopressin (AVP)- and angiotensin II (Ang II)-induced increases in [Ca2+]i mobilization or in MAP kinase activity. Preincubation with BQ-123 did not enhance its inhibitory effects but these effects of BQ-123 were diminished by washing after preincubation. Receptor studies revealed that the washing procedure decreased the inhibitory effect of BQ-123 on ET-1 binding to receptors. These results indicate that BQ-123 is a potent and specific ET(A) receptor antagonist that blocks the ET-1-induced cellular contraction and proliferation in rat VSMC.
Guo
X
X
Division of Endocrinology and Metabolism, Jichi Medical School, Tochigi, Japan.
Okada
K
K
Fujita
N
N
Ishikawa
S
S
Komatsu
N
N
Saito
T
T
D000804:Angiotensin II / Q000494:pharmacology; D000818:Animals; D001127:Arginine Vasopressin / Q000494:pharmacology; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D065128:Endothelin Receptor Antagonists; D016232:Endothelins / Q000494:pharmacology*; D066298:In Vitro Techniques; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D010456:Peptides, Cyclic / Q000494:pharmacology*; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley; D013778:Tetrazolium Salts; D013844:Thiazoles; D013936:Thymidine / Q000378:metabolism
D065128:Endothelin Receptor Antagonists; D016232:Endothelins; D010456:Peptides, Cyclic; D013778:Tetrazolium Salts; D013844:Thiazoles; D000804:Angiotensin II; D001127:Arginine Vasopressin; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C022616:thiazolyl blue; C072247:cyclo(Trp-Asp-Pro-Val-Leu); D002118:Calcium; D013936:Thymidine