Suchard S J, Burton M J, Dixit V M, Boxer L A
Department of Pediatrics, University of Michigan School of Medicine, Ann Arbor 48109.
J Immunol. 1991 Jun 1;146(11):3945-52.
Thrombospondin (TSP), a 450-kDa trimeric glycoprotein secreted by platelets and endothelial cells at sites of tissue injury or inflammation, may play an important role in polymorphonuclear leukocyte (PMN) adherence to blood vessel walls before diapedesis. We have examined the adherence of PMN to TSP and compared it to adherence to other extracellular matrix proteins. PMN adherence to TSP-coated plastic was complete by 60 min with spreading completed by 2 h. The kinetics of adhesion and spreading on TSP were similar to that of vitronectin (VN), laminin (LN), and fibronectin (FN). Activation of PMN with the calcium ionophore A23187 or the chemotactic peptide FMLP increased PMN adherence to LN and FN, but not to TSP or VN, suggesting that PMN activation may differentially regulate expression of TSP and VN receptors as compared to LN and FN receptors. The specificity of PMN adherence to TSP was confirmed by competition with saturating amounts of TSP and inhibition with anti-TSP antibodies. mAb A6.1, which binds to the protease-resistant core of TSP, was the most effective in blocking PMN adherence to TSP. Using TSP proteolytic fragments, we demonstrated that the primary interaction of PMN with TSP was mediated through the 140-kDa COOH-terminal domain. Inasmuch as the 140-kDa fragment of TSP contains an Arg-Gly-Asp sequence similar to the cell recognition site of FN and VN, we determined whether RGDS peptides would inhibit PMN adhesion. RGDS did not significantly inhibit PMN adhesion to TSP, VN, or LN, but reduced PMN adhesion to FN by 50%. To determine if PMN adhesion to TSP was mediated by a beta 2 integrin receptor such as LFA-1, MO-1, or p150,95, we performed adhesion assays using PMN isolated from patients with leukocyte adhesion deficiency that lack beta 2 receptors. Leukocyte adhesion deficiency PMN exhibited normal adherence to TSP. In contrast, adherence to VN, LN, and FN was reduced by 95%. Therefore, adherence to TSP is probably not mediated by a beta 2 integrin receptor. These data contribute to the accumulating evidence that PMN can interact with extracellular matrix proteins through a CD11/CD18-independent process.
血小板反应蛋白（TSP）是一种由血小板和内皮细胞在组织损伤或炎症部位分泌的450 kDa三聚体糖蛋白，它可能在多形核白细胞（PMN）在渗出前黏附于血管壁的过程中发挥重要作用。我们研究了PMN对TSP的黏附，并将其与对其他细胞外基质蛋白的黏附进行了比较。PMN对包被有TSP的塑料的黏附在60分钟时完成，铺展在2小时时完成。PMN在TSP上的黏附和铺展动力学与玻连蛋白（VN）、层粘连蛋白（LN）和纤连蛋白（FN）相似。用钙离子载体A23187或趋化肽FMLP激活PMN会增加PMN对LN和FN的黏附，但不会增加对TSP或VN的黏附，这表明与LN和FN受体相比，PMN激活可能会差异性地调节TSP和VN受体的表达。通过用饱和量的TSP进行竞争以及用抗TSP抗体进行抑制，证实了PMN对TSP黏附的特异性。与TSP的蛋白酶抗性核心结合的单克隆抗体A6.1在阻断PMN对TSP的黏附方面最有效。使用TSP蛋白水解片段，我们证明了PMN与TSP的主要相互作用是通过140 kDa的COOH末端结构域介导的。由于TSP的140 kDa片段包含一个与FN和VN的细胞识别位点相似的Arg-Gly-Asp序列，我们确定RGDS肽是否会抑制PMN的黏附。RGDS并没有显著抑制PMN对TSP、VN或LN的黏附，但使PMN对FN的黏附减少了50%。为了确定PMN对TSP的黏附是否由β2整合素受体（如LFA-1、MO-1或p150,95）介导，我们使用从缺乏β2受体的白细胞黏附缺陷患者中分离出的PMN进行了黏附试验。白细胞黏附缺陷的PMN对TSP表现出正常的黏附。相比之下，对VN、LN和FN的黏附减少了95%。因此，对TSP的黏附可能不是由β2整合素受体介导的。这些数据为PMN可以通过不依赖CD11/CD18的过程与细胞外基质蛋白相互作用这一越来越多的证据做出了贡献。
