Identification of a new role of membrane‐type 1 matrix metalloproteinases in corneal neovascularization

膜 1 型基质金属蛋白酶在角膜新生血管形成中的新作用的鉴定

• 批准号: 10585794
• 负责人: Kyuyeon Han
• 金额: $ 39.98万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-01-01 至 2027-11-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10585794
• 关键词: Angiogenesis Inhibitors; Angiogenic Factor; Angiostatins; Animal Model; Antibodies; Binding; Biological Assay; Biophysics; Blindness; Blood Vessels; Cells; Cellular Infiltration; Clioquinol; Cornea; Corneal Diseases; Corneal Injury; Corneal Neovascularization; Effectiveness; Endostatins; Endothelial Cells; Enzymes; Equilibrium; Event; Extracellular Matrix; Extracellular Matrix Degradation; FDA approved; FGF2 gene; FGFR2 gene; Fibroblast Growth Factor; Fibroblasts; Folic Acid; Gelatinase A; Gelatinase B; Goals; Impairment; Implant; In Vitro; Infection; Infiltration; Inflammation; Inflammatory; Injury; KDR gene; Knock-out; Knockout Mice; Lead; Libraries; MMP14 gene; Macrophage; Matrix Metalloproteinase Inhibitor; Matrix Metalloproteinases; Measures; Mediating; Molecular Biology Techniques; Mus; Neutrophil Collagenase; Pharmaceutical Preparations; Proteins; Proteolysis; Regulation; Reporting; Role; Signal Transduction; Specificity; System; TNF-alpha converting enzyme; Testing; Therapeutic; Therapeutic Agents; Tissues; VEGFA gene; Validation; Vascular Endothelial Cell; Vascular Endothelial Growth Factor Receptor-1; Vascular Endothelial Growth Factors; Vision; Wild Type Mouse; analog; cell type; conditional knockout; cytokine; cytotoxicity; enzyme activity; experimental study; immune cell infiltrate; in vivo; inhibitor; injured; innovation; membrane activity; mutant; novel; novel therapeutics; pigment epithelium-derived factor; proMMP-2; protein expression; quinoline; quinoline analog; receptor; scaffold; side effect; small molecule; small molecule inhibitor

Project Summary/Abstract Corneal neovascularization (NV) can be caused by severe corneal injury or infection and is a leading cause of blindness. Balance of pro-angiogenic factors and anti-angiogenic factors are important to maintain avascular corneal tissue. Pro-angiogenic factors such as VEGFA and FGF2 are highly induced in inflamed corneas and lead to activation of its receptor proteins such as VEGFR2 and FGFR2. Membrane-type 1 matrix metalloproteinase (MMP-14) is involved in remodeling of extracellular matrix (ECM) through its proteolytic activity. Recent studies have revealed that MMP-14 is a regulator of VEGFA/VEGFR2-mediated corneal NV via unique and selective cleavage of VEGFR1 which is a decoy receptor for VEGFR2. FGF2-induced corneal NV is delayed in MMP-14 knockout (KO) mice, indicating there is some correlation between FGF2 and MMP-14. However, how MMP-14 interacts with FGF2 is still largely unknown. The goal of this application is to characterize mechanism of MMP-14 on regulation of FGFR2 levels via ADAM-9 enzyme. We demonstrated that FGFR2 level was low in MMP-14 KO corneal fibroblast cells. On the other hand, ADAM-9, which is substrate of MMP-14, was higher in MMP-14 KO fibroblast than WT cells. We have also discovered that expression of MMP-8, MMP-9, and ADAM-17, all of which are underlying FGF2/FGFR2-system, were highly induced in WT than MMP-14 KO cells upon stimulation of FGF2. Thus, inhibition of MMP-14 can reduce FGF2/FGFR2-mediated corneal NV and inflammation. Furthermore, our results show that FDA-approved small molecule drugs, clioquinol, chloroxine, and folic acid, all of which contain a quinoline scaffold, inhibit MMP-14 enzyme activity. We propose three specific aims: (1) Mechanism of two enzyme cascades, MMP-14 and ADAM-9, to regulate FGFR2 level and expression of FGF2/FGFR2-mediated MMPs; (2) investigate the effect of MMP-14 in FGF2/FGFR2-mediated corneal inflammation; (3) Characterize quinoline analogs as selective MMP-14 inhibitors. We will complete these aims using innovative techniques from molecular biology and biophysics in vitro and in vivo.

项目摘要/摘要 角膜新生血管(NV)可由严重的角膜损伤或感染引起，是主要原因 失明的恐惧。促血管生成因子和抗血管生成因子的平衡对维持无血管状态很重要 角膜组织。促血管生成因子如VEGFA和FGF2在炎症的角膜和 导致其受体蛋白的激活，如VEGFR2和FGFR2。膜型1基质 基质金属蛋白酶-14通过其蛋白水解酶参与细胞外基质的重塑 活动。最近的研究表明，基质金属蛋白酶-14是VEGFA/VEGFR2介导的角膜新生血管的调节因子。 VEGFR2诱骗受体VEGFR1的独特和选择性切割。成纤维细胞生长因子2诱导的角膜新生血管 在MMP14基因敲除(KO)小鼠中延迟表达，表明FGF2和MMP14之间存在一定的相关性。 然而，基质金属蛋白酶-14如何与成纤维细胞生长因子2相互作用在很大程度上仍不清楚。本应用程序的目标是将 基质金属蛋白酶-14通过ADAM-9酶调节FGFR2水平的机制我们证明了FGFR2水平 低表达基质金属蛋白酶-14KO的角膜成纤维细胞。另一方面，作为基质金属蛋白酶-14底物的ADAM-9被 MMP14KO成纤维细胞高于WT细胞。我们还发现，基质金属蛋白酶-8、基质金属蛋白酶-9和基质金属蛋白酶-9的表达 ADAM-17均为FGF2/FGFR2-系统，在WT细胞中的诱导率高于MMP-14 KO细胞 在FGF2的刺激下。因此，抑制基质金属蛋白酶-14可以减少FGF2/FGFR2介导的角膜新生血管，并 发炎。此外，我们的结果表明，FDA批准的小分子药物，CLOQUOL，氯氧辛， 叶酸，所有这些都含有喹啉支架，抑制基质金属蛋白酶-14的活性。我们提出了三个具体的 目的：(1)MMP14和ADAM-9两种酶调节FGFR2水平和表达的机制 (2)探讨基质金属蛋白酶-14在FGF2/FGFR2介导的角膜中的作用。 炎症；(3)将喹啉类似物表征为选择性的基质金属蛋白酶-14抑制剂。我们将完成这些目标 利用来自分子生物学和生物物理学的创新技术，在体外和体内。