Influence of fibrin structure on the formation and maintenance of capillary-like tubules by human microvascular endothelial cells.
Influence of fibrin structure on the formation and maintenance of capillary-like tubules by human microvascular endothelial cells.
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DOI：
10.1023/a:1009240522808
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发表时间：
1998-01-01
期刊：
影响因子：
9.8
通讯作者：
van Hinsbergh, V W
中科院分区：
文献类型：
--
作者：
Collen, A;Koolwijk, P;van Hinsbergh, V W
纤维蛋白是一种临时基质，不仅覆盖伤口，而且在愈合期间为入侵细胞提供结构。凝块凝胶化之前聚合条件的变化影响纤维蛋白的结构，从而可能影响与侵入细胞的相互作用。因此，我们测试了纤维蛋白结构的变化是否会影响人微血管内皮细胞（hMVEC）在体外血管生成模型中形成毛细血管样管状结构。将在pH 7.0下制备的不透明[125 I]纤维蛋白结构、在pH 7.4下制备的纤维蛋白基质和在pH 7.8下制备的透明[125 I]纤维蛋白结构中和（pH 7.4），然后将hMVEC以汇合密度接种在它们的顶部上。用生长因子[碱性成纤维细胞生长因子（bFGF）和血管内皮生长因子（VEGF）165]和细胞因子[肿瘤坏死因子（TNF）-α]刺激内皮细胞，以诱导u-PA/u-PA受体依赖性毛细血管样管状结构的形成。通过图像分析确定侵入性结构的长度并通过测量伴随的[125 I]纤维蛋白降解来量化这些结构的形成。管状结构的向内生长进行了更快的速度在不透明的基质组成的厚纤维蛋白纤维相比，透明的凝胶与细纤维蛋白纤维。bFGF/TNF-α或VEGF 165/TNF-α诱导的不透明纤维蛋白凝胶中管状结构的更快速向内生长伴随着更大程度的纤维蛋白降解。抑肽酶和ε-氨基己酸抑制这两个过程表明纤溶酶的参与。它们也被抗u-PA或抗u-PA受体IgG抑制，但不被抗t-PA IgG抑制，表明细胞结合u-PA活性的参与。然而，在不透明的纤维蛋白凝胶中，由于过度的纤维蛋白降解，长时间孵育后管状结构溶解。单独用bFGF模拟hMVEC不会诱导管状结构，但会导致高度的t-PA和纤溶酶依赖性纤维蛋白溶解，并且几天后，细胞片部分脱离。通过一系列抑肽酶浓度逐渐抑制过度的纤维蛋白降解，不会导致bFGF处理的细胞中的管形成。这些数据表明，纤维蛋白中hMVEC的管状结构的形成和稳定性伴随着受控的纤维蛋白溶解，并且不仅严重依赖于细胞结合的u-PA依赖性纤溶酶原激活，而且依赖于纤维蛋白结构。由于纤维蛋白结构在很大程度上受到纤维蛋白聚合条件的影响，因此这些条件可能对伤口愈合期间的血管生成和肿瘤基质的血管化产生相当大的影响。
Fibrin is a temporary matrix which not only covers a wound, but also provides a structure for invading cells during healing. Changes in the polymerization conditions before gelation of the clot affect the structure of fibrin and thus might influence the interaction with invading cells. Therefore we tested whether changes in the fibrin structure influence the formation of capillary-like tubular structures by human microvascular endothelial cells (hMVEC) in an in vitro angiogenesis model. Opaque [125I]fibrin structures prepared at pH 7.0, fibrin matrices at pH 7.4 and transparent [125I]fibrin structures prepared at pH 7.8 were neutralized (pH 7.4) before seeding hMVEC on top of them in confluent density. Endothelial cells were stimulated with a growth factor [basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF)165] and a cytokine [tumor necrosis factor (TNF)-alpha] to induce the u-PA/u-PA receptor-dependent formation of capillary-like tubular structures. The formation of these structures was quantified by determining the length of the invasive structures by image analysis and by measuring the accompanying [125I]fibrin degradation. Ingrowth of tubular structures proceeded at a faster rate in opaque matrices consisting of thick fibrin fibers as compared to transparent gels with fine fibrin fibers. The more rapid ingrowth of tubular structures in opaque fibrin gels induced by bFGF/TNF-alpha or VEGF165/TNF-alpha was accompanied by a larger extent of fibrin degradation. Both processes were inhibited by aprotinin and epsilon-aminocaproic acid indicating the involvement of plasmin. They were also inhibited by anti-u-PA or anti-u-PA receptor IgG, but not by anti-t-PA IgG, suggesting the involvement of cell-bound u-PA activity. However, in the opaque fibrin gels, the tubular structures dissolved upon prolonged incubation due to excessive fibrin degradation. Simulation of hMVEC with bFGF alone did not induce tubular structures, but ca used a high degree of t-PA- and plasmin-dependent fibrin lysis, and, after several days, a partial detachment of sheets of cells. Gradual inhibition of the excessive fibrin degradation by a series of aprotinin concentrations did not lead to tube formation in bFGF-treated cells. These data indicate that the formation and stability of tubular structures by hMVEC in fibrin is accompanied by controlled fibrinolysis and depends critically not only on cell-bound u-PA-dependent plasminogen activation, but also on the fibrin structure. Because the fibrin structure is largely influenced by the conditions in which fibrin has been polymerized, these conditions may have considerable impact on angiogenesis during wound healing and vascularization of tumour stroma.