Barnathan E S, Kuo A, Van der Keyl H, McCrae K R, Larsen G R, Cines D B
Hematology/Oncology Section, Hospital of the University of Pennsylvania, University of Pennsylvania School of Medicine, Philadelphia 19104.
J Biol Chem. 1988 Jun 5;263(16):7792-9.
The endothelium may contribute to fibrinolysis through the binding of plasminogen activators or plasminogen activator inhibitors to the cell surface. Using a solid-phase radioimmunoassay, we observed that antibodies to recombinant tissue-type plasminogen activator (rt-PA) and plasminogen activator inhibitor type 1 (PAI-1) bound to the surface of cultured human umbilical vein endothelial cells (HUVEC). HUVEC also specifically bound added radiolabeled rt-PA with apparent steady-state binding being reached by 1 h at 4 degrees C. When added at low concentrations (less than 5 nM), rt-PA bound with high affinity mainly via the catalytic site, forming a sodium dodecyl sulfate-stable 105-kDa complex which dissociates from the cell surface over time and which could be immunoprecipitated by a monoclonal antibody to PAI-1. rt-PA bound to this high affinity site retained less than 5% of its expected plasminogen activator activity. At higher concentrations, binding did not require the catalytic site and was rapidly reversible. rt-PA initially bound to this site retained plasminogen activator activity. These studies suggest that tissue-type plasminogen activator and PAI-1 are expressed on the surface of cultured HUVEC. HUVEC also express unoccupied binding sites for exogenous tissue-type plasminogen activator. The balance between the expression of plasminogen activator inhibitors and these unoccupied binding sites for plasminogen activators on the endothelial surface may contribute to the regulation of fibrinolysis.
内皮细胞可能通过纤溶酶原激活剂或纤溶酶原激活剂抑制剂与细胞表面的结合来促进纤维蛋白溶解。使用固相放射免疫测定法，我们观察到针对重组组织型纤溶酶原激活剂（rt-PA）和1型纤溶酶原激活剂抑制剂（PAI-1）的抗体与培养的人脐静脉内皮细胞（HUVEC）表面结合。HUVEC还特异性结合添加的放射性标记rt-PA，在4℃下1小时达到明显的稳态结合。当以低浓度（小于5 nM）添加时，rt-PA主要通过催化位点以高亲和力结合，形成十二烷基硫酸钠稳定的105-kDa复合物，该复合物随时间从细胞表面解离，并且可以被抗PAI-1单克隆抗体免疫沉淀。与该高亲和力位点结合的rt-PA保留的纤溶酶原激活剂活性不到其预期活性的5%。在较高浓度下，结合不需要催化位点且迅速可逆。最初与该位点结合并保留纤溶酶原激活剂活性的rt-PA。这些研究表明，组织型纤溶酶原激活剂和PAI-1在培养的HUVEC表面表达。HUVEC还表达外源性组织型纤溶酶原激活剂的未占据结合位点。内皮表面纤溶酶原激活剂抑制剂的表达与这些纤溶酶原激活剂的未占据结合位点之间的平衡可能有助于纤维蛋白溶解的调节。
