title: Thromboxane binding and signal transduction in rat glomerular mesangial cells.
abstract: Thromboxane A2 (TxA2) stimulates contraction of glomerular mesangial cells. However, mesangial cell TxA2 receptors have not been previously characterized. We therefore investigated TxA2 binding and TxA2-associated signal transduction pathways in rat glomerular mesangial cells using the specific thromboxane receptor agonist (1S-[1 alpha,2 beta(5Z),3 alpha-(1E,3S)4 alpha])-7-(3-[3-hydroxy-4-(p- iodophenoxy)-1-butenyl]7-oxabicyclo[2.2.1]hept-2-yl)-5-heptenoic acid (IBOP). In these cells, [125I]BOP binding was saturable, displaceable, and of high affinity. Scatchard analysis revealed a single class of binding sites with a dissociation constant (Kd) of 293 pM and a maximal density of binding sites (Bmax) of 33 fmol/mg protein. Specific binding was inhibited by the thromboxane agonist (15S)-hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5Z,13E-dienoic acid (U-46619) [inhibitor dissociation constant (Ki) = 297 nM] and the TxA2 receptor antagonists SQ 29548 (Ki = 1 nM) and (1R-[1 alpha(Z),2 beta,3 beta,5 alpha])-(+)-7-(5-[(1,1'-biphenyl)- 4-yl-methoxy]-3-hydroxy-2-(1-piperidinyl)cyclopentyl]-4-heptenoic acid (GR 32191) (Ki = 92 nM). Binding was also highly specific for thromboxane because prostaglandin E2 (Ki = 16 microM) and the inactive thromboxane metabolite, TxB2 (Ki = 41 microM), were approximately 1,000-fold less potent at inhibiting binding. IBOP stimulated phosphatidylinositol hydrolysis with an effective concentration of drug that produces 50% of the maximal response of 229 pM, which correlated well with the equilibrium Kd and enhanced phosphorylation of an acidic 80-kDa protein substrate for protein kinase C.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Medicine, Duke University Medical Center, Durham, North Carolina.
authors.forename: R F
authors.initials: RF
authors.lastname: Spurney
authors.forename: J J
authors.initials: JJ
authors.lastname: Onorato
authors.forename: F J
authors.initials: FJ
authors.lastname: Albers
authors.forename: T M
authors.initials: TM
authors.lastname: Coffman
mesh_terms: D000818:Animals; D001667:Binding, Competitive; D001713:Biphenyl Compounds / Q000378:metabolism / Q000494:pharmacology; D001643:Bridged Bicyclo Compounds / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D005231:Fatty Acids, Unsaturated / Q000378:metabolism; D006538:Heptanoic Acids / Q000378:metabolism / Q000494:pharmacology; D006834:Hydrazines / Q000378:metabolism / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins; D007678:Kidney Glomerulus / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D007700:Kinetics; D008565:Membrane Proteins; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D013931:Thromboxanes / Q000378:metabolism*
chemical_list: D001713:Biphenyl Compounds; D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D005231:Fatty Acids, Unsaturated; D006538:Heptanoic Acids; D006834:Hydrazines; D007295:Inositol Phosphates; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D011506:Proteins; D013931:Thromboxanes; C061750:7-(3-(3-hydroxy-4-(4'-iodophenoxy)-1-butenyl)-7-oxabicyclo(2.2.1)heptan-2-yl)-5-heptenoic acid; D000076250:Myristoylated Alanine-Rich C Kinase Substrate; C045749:SQ 29548; D011493:Protein Kinase C; C060013:vapiprost