Autophagic Lysosomal Pathway and Glaucoma

自噬溶酶体途径与青光眼

• 批准号: 8461206
• 负责人: Paloma Liton
• 金额: $ 32.01万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-01 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8461206
• 关键词: Affect; Age; Aging; Aging-Related Process; Alzheimer' s Disease; Anterior; Applications Grants; Aqueous Humor; Atherosclerosis; Autophagocytosis; Autophagolysosome; Autophagosome; Biological; Cadaver; Cathepsins; Cell Culture Techniques; Cell physiology; Cells; Chronic; Data; Degradation Pathway; Deposition; Disease; Endocytosis; Enzymes; Excision; Extracellular Matrix; Extracellular Space; Eye; Functional disorder; Galactosidase; Garbage; Genes; Genetic; Glaucoma; Human; In Vitro; Laboratories; Life; Lysosomes; Mediating; Mitochondria; Modeling; Molecular; Monitor; Mus; Organelles; Oxidative Stress; Parkinson Disease; Pathogenesis; Pathology; Pathway interactions; Phenotype; Physiologic Intraocular Pressure; Pigments; Predisposition; Primary Open Angle Glaucoma; Proteins; Proteolysis; Rattus; Reactive Oxygen Species; Risk; Stress; Structure of sinus venosus of sclera; Testing; Tissues; Trabecular meshwork structure; Transgenic Organisms; Vacuole; Vesicle; age related; base; in vivo; mouse model; mutant; myocilin; normal aging; response; senescence; stressor; therapeutic target; tool; wasting

ABSTRACT The fundamental abnormality occurring in the conventional outflow pathway associated with elevated intraocular pressure and therefore, increased risk of developing glaucoma, an age-related disease affecting more than 70 million people world wide still remains obscure. However, data from several laboratories, including ours, support a key role of reactive oxygen species, both present in the aqueous humor as well as generated during the normal aging process within the outflow pathway, in the pathogenesis of glaucoma. Autophagy, a lysosomal pathway responsible for the degradation of long-lived proteins and organelles, has emerged as an important cellular homeostatic mechanism that is part of the early protective cellular response against oxidative stress. A general decline in autophagic activity has been observed in several tissues with aging and in age-related disorders. A corollary question is whether autophagy function declines with age in the outflow pathway tissue, and if so, whether this could contribute to the susceptibility to disease. Our preliminary data show that exposure of primary cultures of trabecular meshwork (TM) cells to chronic oxidative stress causes profound changes in the lysosomal degradative pathway, including: (1) Increased lysosomal mass and lysosomal enzymes protein content, (2) increased autophagic vacuoles content, (3) upregulated levels of LC3-II, (4) accumulation of intralysosomal oxidized material and damaged mitochondria, and (5) decreased cathepsin activities. In addition, stressed cultures showed elevated senescence-associated- ¿-galactosidase (SA-¿-gal), a marker found to be also upregulated in the TM from glaucoma donors. We hypothesize that aging of the outflow pathway is accompanied by a decline in the autophagic degradative capacity, thus leading to the inefficient removal of oxidized components and to the intracellular accumulation of nonfunctional aberrant cellular components, which reduce the ability of TM cells to respond against additional stressors of the autophagic pathway (i.e. mutant myocilin, pigment) further compromises the autophagic cellular function, thus promoting the secretion of autophagolysosomes into the extracellular space, which can contribute to the abnormal deposition observed in glaucoma. To test this hypothesis, we will investigate (1) whether aging of TM cells is associated with a decrease in autophagic flux in vitro and in vivo; (2), whether the experimentally-induced decreased in autophagic capacity in TM cells results in the accumulation of damaged proteins and organelles, as well as extracellular matrix vesicles; and (3), whether autophagy dysfunction is associated with the acquisition of a glaucoma phenotype, including the presence of extracellular matrix vesicles, in human eyes and in established mice glaucoma models.

抽象的 与升高相关的常规出口通路中发生的基本异常 眼内压，因此增加了青光眼发展的风险，这种疾病会影响 全球超过7000万人仍然晦涩。但是，来自几个实验室的数据， 包括我们的包括，支持活性氧的关键作用，既存在于水性幽默以及 在出口通路内的正常老化过程中，在青光眼的发病机理中产生。 自噬是一种导致长寿命蛋白质和细胞器降解的溶酶体途径，具有 成为重要的细胞稳态机制，是早期受保护的细胞反应的一部分 反对氧化应激。在几次中观察到自噬活动的总体下降 衰老和与年龄有关的疾病。推论问题是自噬功能是否随着年龄的年龄而下降 出口途径组织，如果是这样，这是否会导致对疾病的易感性。 我们的初步数据表明，小梁网（TM）细胞的主要培养物暴露于慢性 氧化应激会导致溶酶体降解途径的深刻变化，包括：（1）增加 溶酶体质量和溶酶体酶蛋白含量，（2）自噬液泡含量增加，（3） LC3-II的上调水平，（4）杂物体内氧化材料和线粒体受损的积累 （5）开发了组织蛋白酶的活动。此外，显示的压力培养物升高了感应相关 - - 半乳糖苷酶（Sa-®-Gal），发现在青光眼供体的TM中也更新了标记。 我们假设出口通路的衰老是通过自噬的下降来实现的 降解能力，从而导致氧化成分的效率低下，并导致细胞内 非功能异常细胞成分的积累，这降低了TM细胞反应的能力 反对自噬途径的其他应力源（即突变肌蛋白，色素）进一步损害了 自噬性细胞功能，从而促进自噬物质体向细胞外空间的分泌， 这可以有助于在青光眼中观察到的异常沉积物。为了检验这一假设，我们将 （1）TM细胞的衰老是否与体外和体内自噬通量的降低有关； （2），实验诱导的TM细胞自噬能力的降低是否导致 受损的蛋白质和细胞器以及细胞外基质蔬菜的积累； （3），是否 自噬功能障碍与获得青光眼表型的获取有关，包括存在 细胞外基质蔬菜，在人眼中和已建立的小鼠青光眼模型中。