Fibroblast migration in fibrin gel matrices.
Fibroblast migration in fibrin gel matrices.
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发表时间：
1993
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通讯作者：
L. Brown;Naomi Lanir;Jan McDonagh;K. Tognazzi;Ann M. Dvorak;Harold F. Dvorak
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作者：
L. Brown;Naomi Lanir;Jan McDonagh;K. Tognazzi;Ann M. Dvorak;Harold F. Dvorak
在愈合的伤口和许多实体瘤中，局部增加的微血管通透性导致纤维蛋白原外渗及其血管外凝血形成纤维蛋白凝胶，伴随着因子XIIIa对纤维蛋白的共价交联。随后，炎性细胞、成纤维细胞和内皮细胞迁移到凝胶中，并将其组织成肉芽组织，随后组织成成熟的胶原结缔组织。为了深入了解与这些过程相关的一些细胞迁移事件，我们开发了一种定量的体外试验，可以研究纤维蛋白凝胶中的成纤维细胞迁移。使早期传代的人或大鼠成纤维细胞附着在组织培养皿上，然后覆盖一薄层用凝血酶凝结的纤维蛋白原。成纤维细胞在24小时内开始向上迁移到纤维蛋白中，并在几天内定量其数量和迁移距离。成纤维细胞迁移的程度受到纤维蛋白凝块性质的重要影响。成纤维细胞最佳地迁移到由浓度为~ 3 mg/ml的纤维蛋白原制备的凝胶中;即，接近正常血浆纤维蛋白原水平。当体内发生凝血时，通过因子XIIIa广泛交联纤维蛋白α链，可大大增强迁移。当纤维蛋白原在Dulbecco改良Eagle培养基中凝结时，γ链交联，但α链交联被显著抑制，成纤维细胞迁移较差。从因子XIII-耗尽纤维蛋白原制备的凝胶既没有表现出α-也没有表现出γ-链交联，并且不支持成纤维细胞迁移。通过阴离子交换高压液相色谱法进一步纯化纤维蛋白原，耗尽纤维蛋白原中的纤连蛋白、纤溶酶原和其他杂质;这种纯化的纤维蛋白原凝结形成纤维蛋白凝胶，支持可再现的成纤维细胞迁移。
In healing wounds and many solid tumors, locally increased microvascular permeability results in extravasation of fibrinogen and its extravascular coagulation to form a fibrin gel, with concomitant covalent cross-linking of fibrin by factor XIIIa. Subsequently, inflammatory cells, fibroblasts, and endothelial cells migrate into the gel and organize it into granulation tissue and later into mature collagenous connective tissue. To gain insight into some of the cell migration events associated with these processes, we developed a quantitative in vitro assay that permits the study of fibroblast migration in fibrin gels. Early passage human or rat fibroblasts were allowed to attach to tissue culture dishes and then were overlaid with a thin layer of fibrinogen that was clotted with thrombin. Fibroblasts began to migrate upwards into the fibrin within 24 hours and their numbers and the distance migrated were quantified over several days. The extent of fibroblast migration was affected importantly by the nature of the fibrin clot. Fibroblasts migrated optimally into gels prepared from fibrinogen at concentrations of -3 mg/ml; ie, near normal plasma fibrinogen levels. Migration was greatly enhanced by extensive cross-linking of the fibrin alpha-chains by factor XIIIa, as occurs when clotting takes place in vivo. When fibrinogen was clotted in Dulbecco's modified Eagle's medium, gamma-chains were cross-linked, but alpha-chain cross-linking was strikingly inhibited, and fibroblasts migrated poorly. Gels prepared from factor XIII-depleted fibrinogen exhibited neither alpha-nor gamma-chain cross-linking and did not support fibroblast migration. Further purification of fibrinogen by anion exchange high pressure liquid chromatography depleted fibrinogen of fibronectin, plasminogen, and other impurities; this purified fibrinogen clotted to form fibrin gels that supported reproducible fibroblast migration.