Lysosomal Enzymes in Outflow Pathway Physiology and Pathophysiology

流出途径生理学和病理生理学中的溶酶体酶

• 批准号: 9284304
• 负责人: Paloma Liton
• 金额: $ 45.75万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-06-01 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9284304
• 关键词: Aging; Agreement; Alpha Cell; Applications Grants; Aqueous Humor; Biochemical; Blindness; Caspase; Cathepsins; Cathepsins B; Caveolae; Cell Surface Receptors; Cell surface; Cells; Clinical; Clinical Trials; Collagen; Collagen Fiber; Connective Tissue; Cues; Data; Degradation Pathway; Deposition; Disease; Drainage procedure; Drug Targeting; Elastic Fiber; Endothelial Cells; Environment; Enzymes; Etiology; Extracellular Matrix; Eye; Functional disorder; Future; Glaucoma; Ground Substance; Human; Image; In Vitro; Laboratories; Matrix Metalloproteinases; Mechanics; Mediating; Modeling; Molecular; Mus; Nature; Ocular Hypertension; Pathogenesis; Pathway interactions; Patients; Peptide Hydrolases; Pharmacology; Physiologic Intraocular Pressure; Physiology; Play; Primary Open Angle Glaucoma; Proteomics; Resistance; Retinal Ganglion Cells; Risk; Role; Route; Serine Protease; Testing; Time; Tissues; Trabecular meshwork structure; Transforming Growth Factor Beta 2; Transforming Growth Factor beta; Work; base; carboxypeptidase C; extracellular; in vivo; novel; novel therapeutics; prevent; protein B; uptake

SUMMARY Defective aqueous humor (AH) drainage through the trabecular meshwork (TM) / Schlemm's cannal (SC) conventional outflow pathway is usually associated to elevated intraocular pressure (IOP), and hence, increased risk for developing glaucoma, a blinding disease second leading cause of permanent blindness in the US. The nature of such resistance to AH outflow is far from being elucidated. The TM consists of sheets of connective tissue beams lined by TM endothelial cells. Each beam is composed of a central elastic core surrounded by collagen fibers embedded in a ground substance. The glaucomatous outflow pathway is characterized by thickening of the trabecular lamellae and accumulation of long-spacing collagen bundles and elastic fiber sheaths, which is presumed to stiffen the TM and prevent the tissue to respond to mechanical cues. The exact causes underlying the deposit of extracellular material (ECM) and thickening of the beams remain unknown; but it is likely a consequence of excessive synthesis of ECM components, decreased proteolytic degradation, or both. Matrix metalloproteinases (MMPs) have been historically believed to be the major proteases involved in ECM degradation; however, emerging evidence indicates that while MMPs play a critical role in initiating ECM degradation in the extracellular environment, other proteases or the coordinated action of several types of proteases (i.e. cysteine and serine proteases) are responsible for the bulk matrix degradation, occurring pericellularly and intracellularly in the lysosomal compartment, associated to lysosomal cathepsins. In agreement, studies conducted in our laboratory have clearly demonstrated the constitutive cell surface expression and secretion of cathepsin B (CTSB) in TM cells and its participation in the endocytic uptake and intralysosomal degradation of native and denature collagens. Here we propose to investigate for the first time the contribution of a CTSB-mediated pericellular and intracellular ECM degradative pathway in TM cells and the effect on outflow physiology. We hypothesize that CTSB plays a critical role in ECM remodeling and outflow physiology by initiating a proteolytic cascade leading to the pericellular and intracellular degradation of ECM components in TM cells. We further hypothesize that modulation of CTSB activity may represent a novel therapeutic strategy in glaucoma. To test this hypothesis, we will (1) demonstrate that CTSB associates with caveolae and initiates a proteolytic cascade resulting in the partial degradation of ECM components; (2) determine the cell surface receptors participating in of ECM components in TM cells; and (3) evaluate the effects of modulating CTSB activity in outflow pathway function of murine glaucoma models.

概括 通过小梁网（TM） / Schlemm的Cannal（SC）的缺陷水幽默（AH）排水 常规的流出途径通常与眼内压（IOP）升高有关，因此，因此 增加青光眼的风险增加，这是盲目疾病的第二主要原因，导致永久性失明 美国。这种对AH流出的抵抗力的性质远非被阐明。 TM由TM内皮细胞衬里的结缔组织束组成。每个光束都组成 被嵌入地面物质中的胶原纤维包围的中央弹性芯。青光眼 流出途径的特征是小梁薄片的增厚和长间距的积累 胶原蛋白束和弹性纤维鞘，假定可以使TM变硬并防止组织 响应机械提示。确切的原因是细胞外材料（ECM）沉积的根本原因和 梁的增厚仍然未知；但这可能是由于过度合成ECM的结果 成分，蛋白水解降解或两者兼而有之。基质金属蛋白酶（MMP）已经 从历史上看，据信是与ECM降解有关的主要蛋白酶；但是，新兴的证据 表明MMP在在细胞外环境中启动ECM降解方面起关键作用，但 其他蛋白酶或几种类型的蛋白酶（即半胱氨酸和丝氨酸蛋白酶）的协调作用是 负责散装基质降解，细胞周围发生在溶酶体中 车厢，与溶酶体组织蛋白酶有关。同意，在我们的实验室进行的研究 清楚地证明了TM细胞中组织蛋白酶B（CTSB）的组成型细胞表面表达和分泌 以及它参与天然和成熟胶原蛋白的内吞摄取和内溶性降解。 在这里，我们建议第一次调查CTSB介导的周围和 TM细胞中细胞内ECM降解途径以及对流出生理学的影响。我们假设这一点 CTSB通过启动蛋白水解在ECM重塑和流出生理中起关键作用 导致TM细胞中ECM成分的细胞周围和细胞内降解。我们 进一步假设CTSB活性的调节可能代表了一种新颖的治疗策略 青光眼。为了检验这一假设，我们将（1）证明CTSB与Caveolae相关并启动A 蛋白水解级联反应导致ECM成分的部分降解； （2）确定细胞表面 参与TM细胞中ECM成分的受体； （3）评估调节CTSB的效果 鼠青光眼模型的流出途径函数中的活性。