Morinelli T A, Zhang L M, Newman W H, Meier K E
Department of Medicine, Medical University of South Carolina, Charleston 29425.
J Biol Chem. 1994 Feb 25;269(8):5693-8.
Prostaglandin H2 (PGH2) and thromboxane A2 (TXA2) are potent activators of platelets and vascular smooth muscle whose responses are mediated through a common G-protein coupled receptor (TXA2/PGH2 receptor). Despite the many studies describing their ability to aggregate platelets and contract vascular smooth muscle, little is known concerning the potential mitogenic capabilities of these autocoids. Mitogen-activated protein kinases (MAP kinases) and ribosomal S6 kinases are well characterized intracellular mediators involved in proliferation of cells. The present study was designed to examine the activation of MAP kinase and S6 kinase in guinea pig coronary artery smooth muscle cells (CASMC) in response to stimulation by a TXA2/PGH2 mimetic, I-BOP ([1S-(1 alpha,2 beta(5Z),3 alpha(1E,3R*),4 alpha)]-7-[3-(3-hydroxy-4-(4'- iodophenoxy)-1-butenyl)-7-oxabicyclo-[2.2.1]heptan-2-yl]-5-h eptenoic acid). Equilibrium radioligand binding assays using [125I]BOP defined a single class of high affinity TXA2/PGH2 receptors on monolayers of guinea pig CASMC (Kd = 0.18 +/- 0.03 nM; 26,476 +/- 3,600 sites/cell; 0.08 +/- 0.01 pmol/mg of protein; n = 12). I-BOP produced a concentration-dependent increase in [3H]thymidine incorporation in these cells (EC50 = 0.3 nM) which was inhibited by a series of TXA2/PGH2 receptor antagonists as well as by verapamil and staurosporine. I-BOP also produced a time-dependent increase in the activation of kinases phosphorylating myelin basic protein (MBP; a substrate for MAP kinase) and RRLSSLRA (S6 peptide; a substrate for pp85rsk kinase), reaching a peak activation between 5 and 10 min. Stimulated MBP kinases were identified as ERK1 and ERK2. The activation of these kinases by I-BOP was inhibited by the TXA2/PGH2 receptor antagonist SQ29548 and also by staurosporine. These results indicate that I-BOP, a TXA2/PGH2 mimetic, produces growth of coronary artery vascular smooth muscle cells, which is preceded by activation of MAP kinase and S6 kinase.
前列腺素H2（PGH2）和血栓素A2（TXA2）是血小板和血管平滑肌的强效激活剂，它们的反应是通过一种常见的G蛋白偶联受体（TXA2/PGH2受体）介导的。尽管有许多研究描述了它们聚集血小板和收缩血管平滑肌的能力，但对于这些自体活性物质的潜在促有丝分裂能力却知之甚少。丝裂原活化蛋白激酶（MAP激酶）和核糖体S6激酶是参与细胞增殖的特征明确的细胞内介质。本研究旨在检测豚鼠冠状动脉平滑肌细胞（CASMC）中MAP激酶和S6激酶在受到TXA2/PGH2模拟物I-BOP（[1S-(1α,2β(5Z),3α(1E,3R*),4α)]-7-[3-(3-羟基-4-(4'-碘苯氧基)-1-丁烯基)-7-氧杂双环-[2.2.1]庚烷-2-基]-5-庚烯酸）刺激后的激活情况。使用[125I]BOP进行的平衡放射性配体结合试验确定了豚鼠CASMC单层上的一类单一高亲和力TXA2/PGH2受体（Kd = 0.18 +/- 0.03 nM；26,476 +/- 3,600个位点/细胞；0.08 +/- 0.01 pmol/mg蛋白质；n = 12）。I-BOP使这些细胞中的[3H]胸苷掺入量呈浓度依赖性增加（EC50 = 0.3 nM），这被一系列TXA2/PGH2受体拮抗剂以及维拉帕米和星形孢菌素所抑制。I-BOP还使磷酸化髓鞘碱性蛋白（MBP；MAP激酶的底物）和RRLSSLRA（S6肽；pp85rsk激酶的底物）的激酶活性呈时间依赖性增加，在5至10分钟之间达到峰值激活。受刺激的MBP激酶被鉴定为ERK1和ERK2。I-BOP对这些激酶的激活被TXA2/PGH2受体拮抗剂SQ29548以及星形孢菌素所抑制。这些结果表明，TXA2/PGH2模拟物I-BOP可使冠状动脉血管平滑肌细胞生长，这之前会有MAP激酶和S6激酶的激活。
