The Role of Extracelluar Enzymes in Regulating Corneal Repair

描述（由申请人提供）：角膜混浊影响数百万人，是世界上第二大致盲原因。显然需要开发非手术策略来预防和治疗角膜纤维化以及随后的角膜混浊的发展。先前和初步的研究表明，细胞外酶包括6-O-硫酸酯酶Sulf-1和基质金属蛋白酶（MMP），包括MMP-9和MMP-12，只在角膜损伤后表达，暗示它们可能在角膜修复中的作用。这些酶由上皮细胞、基质细胞和炎症细胞产生，它们在角膜修复中的作用尚未得到充分研究。在上皮中，MMP-9似乎通过其对TGF途径的作用减少再上皮化，而在基质中，MMP-12已显示在各种其他疾病过程中通过TGF-β保护免于纤维化。此外，MMP-12比其他MMP更有效地产生血管抑素的能力表明它可能在预防角膜血管生成中发挥作用。已发现Sulf-1在体外调节生长因子和趋化因子的结合。各种生长因子的刺激，特别是TGF-β 2和FGF，对角膜纤维化表型和角膜新生血管形成至关重要。总之，这些胞外酶可以调节修复过程的各个方面，积极和消极的。在本申请中，我们建议研究细胞外酶MMP-9、MMP-12和Sulf-1在上皮、基质和炎症性角膜对损伤的反应中的作用。具体目标1将确定MMP-9或Sulf-1活性对上皮细胞对损伤的增殖反应的影响。具体目标2将使用MMP-12敲除小鼠来检查MMP- 12在角膜基质对损伤的反应期间在纤维化和血管生成中的作用。具体目标3将检查这些细胞外酶招募炎症细胞到角膜的能力，并将使用一种基于旋转圆盘共聚焦显微镜的新型成像技术来研究角膜中的炎症细胞动态。了解细胞外酶在受伤角膜中的作用，将进一步加深我们对维持角膜透明性的重要机制的理解，并提供基本的见解，从长远来看，可以更好地治疗角膜混浊。

DESCRIPTION (provided by applicant): Corneal opacification affects millions of people and is the second leading cause of blindness in the world. A clear need exists for the development of non-surgical strategies to prevent and treat corneal fibrosis and the subsequent development of corneal opacification. Previous and preliminary studies show that extracellular enymes including the 6-O-sulfatase Sulf-1 and matrix metalloproteinases (MMPs) including MMP-9 and MMP-12 are expressed in the cornea only upon injury, implicating their possible roles in corneal repair. These enzymes are produced by epithelial, stromal and inflammatory cells, and their roles in corneal repair c have not been well-studied. In the epithelium, MMP-9 appears to decrease re-epithelialization through its effects on the TGF pathway, while in the stroma MMP-12 has been shown to be protective against fibrosis via TGF-p in various other disease processes. In addition, MMP-12's ability to produce angiostatin more efficiently than other MMPs suggests that it might play a role in preventing corneal angiogenesis. Sulf-1 has been found to modulate the binding of growth factors and chemokines in vitro. Stimulation of various growth factors, particularly TGF-p2 and FGF, are critical to the corneal fibrotic phenotype and corneal neovascularization. Taken together, these extracellular enzymes may regulate various aspects of the repair process, both positively and negatively. In this application, we propose to investigate the roles of the extracellular enzymes MMP-9, MMP-12, and Sulf-1 in the epithelial, stromal, and inflammatory corneal response to injury. Specific Aim 1 will determine the effects of MMP-9 or Sulf-1 activity on the epithelial cell proliferation response to injury. Specific Aim 2 will use MMP-12 knockout mice to examine the role of MMP- 12 in fibrosis and angiogenesis during the corneal stromal response to injury. Specific Aim 3 will examine the ability of these extracellular enzymes to recruit inflammatory cells to the cornea and will use a novel imaging technique based on spinning disk confocal microscopy to study inflammatory cell dynamics in the cornea in vivo and in real-time. Understanding the roles of extracellular enzymes in wounded corneas should further our understanding of the mechanisms important to the maintenance of corneal clarity and provide basic insights that will, in the long term, allow for the development of better treatment of corneal opacification.