Corneal Repair: uPA and extracellular matrix

描述：无疤痕的角膜伤口愈合可使角膜恢复透明。愈合后形成疤痕的角膜有混浊和视力障碍。因此，了解影响再生愈合而不是纤维愈合的途径是通过治疗促进健康修复的关键一步。在角膜损伤后，基质中通常静止的细胞分化为成纤维细胞，成纤维细胞在迁移时分泌蛋白酶。这项建议集中在细胞外丝氨酸蛋白酶，尿激酶型纤溶酶原激活物(UPA)的贡献，它是在创伤后由角膜成纤维细胞表达的。当uPA与其细胞表面受体uPAR结合时，uPA在细胞-基质界面产生纤溶酶。纤溶酶降解细胞外基质(ECM)，激活潜在的生长因子，如转化生长因子。转化生长因子刺激成纤维细胞的增殖和迁移，并诱导可运动的成纤维细胞分化为非运动的肌成纤维细胞，这是基质收缩和伤口闭合所必需的。转化生长因子可能产生双重作用的一种方式是通过诱导纤溶酶原激活抑制物(PAI-1)表达。有大量数据表明，PAI-1的相反作用是浓度依赖的：低浓度的PAI-1诱导细胞增殖和迁移，而高浓度则抑制这些过程。在这项建议中被检验的假设是，角膜损伤后，局部PAI-1浓度通过与VN结合而调节。此外，低浓度的PAI-1导致角膜成纤维细胞的增殖和迁移，而高浓度的PAI-1诱导uPA/uPAR下调，并导致肌成纤维细胞分化。其具体目的是确定1)转化生长因子浓度是否调节PAI-1水平、uPA活性和肌成纤维细胞分化。2)VN介导了转化生长因子β和纤溶酶原激活物-1对uPA活性、细胞迁移和肌成纤维细胞分化的影响。3)VN、PAI-1和整合素在创面愈合过程中的表达随基质表型的改变而变化。4)uPAR裂解成非uPA结合形式抑制细胞迁移并诱导肌成纤维细胞分化。这些研究的结果将有助于更好地理解引导创面再生修复的机制。

DESCRIPTION: Corneal wound healing without scarring regenerates corneal transparency. Corneas that heal with scarring have opacities and obstructed vision. Thus, understanding the pathways that influence regenerative rather than fibrotic healing is an essential step towards therapeutically promoting healthy repair. Upon corneal wounding, the normally quiescent cells in the stroma differentiate into fibroblasts, which secrete proteases as they migrate. This proposal is focused on the contributions of the extracellular serine protease, urokinase-type plasminogen activator (uPA), which is expressed by corneal fibroblasts after wounding. When uPA binds to its cell-surface receptor, uPAR, uPA generates plasmin at the cell-matrix interface. Plasmin degrades extracellular matrix (ECM) and activates latent growth factors such as TGF¿. TGF¿ stimulates fibroblast proliferation and migration, and induces the differentiation of motile fibroblasts into non-motile myofibroblasts, which are essential for matrix contraction and wound closure. One way in which TGF¿ may be eliciting its dual effects is through the induction of plasminogen activation inhibitor (PAI-1) expression. There is a significant body of data that the opposing effects of PAI-1 are concentration dependent: low concentrations of PAI-1 induce cell proliferation and migration, whereas, high concentrations inhibit these processes. The hypothesis being tested in this proposal is that after corneal wounding, local PAI-1 concentrations are mediated by binding to Vn. Further, that low concentrations of PAI-1 result in corneal fibroblast proliferation and migration whereas high concentrations of PAI-1 induce uPA/uPAR downregulation and result in myofibroblast differentiation. The specific aims are to determine if 1) TGF¿ concentration regulates PAI-1 levels, uPA activity, and myofibroblast differentiation. 2) Vn mediates the effects of TGF¿ and PAI-1 on uPA activity, cell migration, and myofibroblast differentiation. 3) Vn, PAI-1 and integrin expression change throughout wound healing with changing stromal phenotypes. 4) uPAR cleavage into a non-uPA binding form inhibits cell migration and induces myofibroblast differentiation. The results of these studies will lead to a better understanding of the mechanisms that guide regenerative wound repair.