AN ARG-GLY-ASP SEQUENCE WITHIN THROMBIN PROMOTES ENDOTHELIAL-CELL ADHESION
AN ARG-GLY-ASP SEQUENCE WITHIN THROMBIN PROMOTES ENDOTHELIAL-CELL ADHESION
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DOI：
10.1083/jcb.112.2.335
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发表时间：
1991-01-01
影响因子：
7.8
通讯作者：
DEJANA, E
中科院分区：
文献类型：
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作者：
BARSHAVIT, R;SABBAH, V;DEJANA, E
凝血酶除了在止血中发挥核心作用外，还具有与伤口愈合、炎症和动脉粥样硬化有关的多种细胞生物调节功能。在本研究中，我们证明在促凝血或催化位点修饰的凝血酶分子可诱导内皮细胞（EC）粘附、扩散和细胞骨架重组。最有效的粘附性凝血酶类似物（NO2-α-凝血酶）是通过酪氨酸残基的硝化获得的。 NO2-α-凝血酶的细胞粘附促进活性在凝血酶-抗凝血酶 III (ATIII) 复合物形成时被抗凝血酶原抗体阻断，但不受水蛭素影响。含有精氨酸-甘氨酸-天冬氨酸的肽完全抑制ED与NO2-α-凝血酶的粘附，而对应于凝血酶“环B”促有丝分裂位点和凝血酶衍生的趋化片段“CB67-129”的合成肽则无效。免疫荧光研究表明，EC 粘附到 NO2-α-凝血酶后会发生细胞扩散、肌动蛋白微丝组装和焦点接触的形成。通过使用特异性抗体，发现玻连蛋白 (vn) 受体 (alpha-v-beta-3) 在细胞粘附到 NO2-α-凝血酶后以簇形式定位。抗 α-v-β-3 抗体可阻断 EC 粘附和扩散，而抗纤连蛋白 (fn) 受体 (α-5-β-1) 抗体则无效。虽然天然凝血酶表现出非常低的细胞附着活性，但在涂覆塑料表面之前在 37 摄氏度下孵育的凝血酶诱导 EC 附着和扩散。我们建议在某些条件下，凝血酶内自然受阻的 RGD 结构域暴露出来与 EC 上的 alpha-v-beta-3 相互作用。这反过来又促进细胞粘附、扩散和细胞骨架元件的重组，这可能共同有助于受损血管壁的修复机制。这项研究定义了凝血酶的新生物学作用，并表征了 EC 对该分子的新识别机制。
Thrombin, in addition to its central role in hemostasis, possesses diverse cellular bioregulatory functions implicated in wound healing, inflammation, and atherosclerosis. In the present study we demonstrate that thrombin molecules modified either at the procoagulant or catalytic sites induce endothelial cell (EC) adhesion, spreading, and cytoskeletal reorganization. The most potent adhesive thrombin analogue (NO2-alpha-thrombin) was obtained by nitration of tyrosine residues. The cell adhesion promoting activity of NO2-alpha-thrombin was blocked upon the formation of thrombin-antithrombin III (ATIII) complexes and by antiprothrombin antibodies, but was unaffected by hirudin. Arg-Glys-Asp-containing peptides, fully inhibited ED adhesion to NO2-alpha-thrombin, while synthetic peptides corresponding to thrombin "Loop B" mitogenic site and the thrombin-derived chemotactic fragment "CB67-129", were uneffective. Immunofluorescence studies indicated that EC adhesion to NO2-alpha-thrombin was followed by cell spreading, actin microfilament assembly, and formation of focal contacts. By the use of specific antibodies, the vitronectin (vn) receptor (alpha-v-beta-3) was found to be localized in clusters upon cell adhesion to NO2-alpha-thrombin. An anti alpha-v-beta-3 antibody blocked EC adhesion and spreading while antifibronectin (fn) receptor (alpha-5-beta-1) antibodies were uneffective. While native thrombin exhibited a very low cell attachment activity, thrombin that was incubated at 37-degrees-C before coating of plastic surfaces induced EC attachment and spreading. We propose that under certain conditions the naturally hindered RGD domain within thrombin is exposed for interaction with alphia-v-beta-3 on EC. This in turn promotes cell adhesion, spreading, and reorganization of cytoskeletal elements, which may altogether contribute to repair mechanisms in the disturbed vessel wall. This study defines a new biological role of thrombin and characterizes a new recognition mechanism on EC for this molecule.