Impact of Lipofuscin in Retinal Pigment Epithelial Cells

描述（由申请人提供）：随着年龄的增长，眼睛的视网膜色素上皮细胞（RPE）会聚集类双维甲酸荧光团。这组色素，包括创始成员A2E，由于视黄醛的无意反应而在光感受器细胞中形成。它们在被吞噬的外节膜内沉积在RPE中，并以脂褐素的形式积累。这些化合物对RPE细胞的破坏作用与许多年龄相关和早发性黄斑疾病有关，包括隐性Stargardt病、elovl4相关疾病、最佳黄斑营养不良和年龄相关黄斑变性。在本应用中提出的研究的广泛目标是了解视网膜类双维甲酸促进威胁视力的疾病过程的机制。我们将讨论构成RPE脂褐素的类双维甲酸色素和这些化合物在光感受器细胞中的类双维甲酸前体。这项工作将阐明治疗途径。此外，由于类双维甲酸是眼底自身荧光的主要来源，这些研究将有助于对眼底自身荧光图像的临床解释。在Specific Aim 1中，我们将研究我们的初步观察，即类双维甲酸的光裂解释放小的二羰基（甲基乙二醛和乙二醛），它们负责蛋白质的晚期糖基化终产物（AGE）修饰。布鲁氏膜和细胞膜中的蛋白质容易发生age修饰。Bruch膜的老化变化也被认为是导致年龄相关性黄斑变性的原因。我们认为，与糖尿病的情况不同，age -加合物在布鲁氏膜上形成是由于覆盖的RPE释放了二羰基。在特异性目标2中，我们将探索体内RPE双维甲酸光氧化相关光降解的证据。在特异性目标3中，我们将探索位于感光细胞外段的类双维a荧光团，它们是RPE脂褐素的前体。我们将探讨在受损的感光细胞中增加双维甲酸形成的倾向；我们将测试光感受器类双维甲酸介导光感受器细胞脂质氧化和光损伤的能力；我们将证明在何种条件下，光感受器类双维甲酸可以促进眼底自体荧光图像中的超自体荧光。这些目标将通过使用动物和体外模型以及采用生化和组织学方法来实现。这项研究的完成将促进我们对囊肿形成的理解，并将阐明RPE脂褐素与Bruch膜老化变化之间的联系，这可能是AMD的前哨。这些研究将有助于了解光和氧化机制是如何在单基因视网膜疾病的光感受器细胞死亡的因素。

DESCRIPTION (provided by applicant): Retinal pigment epithelial (RPE) cells of the eye amass bisretinoid fluorophores with age. This group of pigments, including the founding member A2E, forms in photoreceptor cells due to inadvertent reactions of retinaldehyde. They are deposited in RPE within phagocytosed outer segment membrane and they accumulate as lipofuscin. The damaging effects of these compounds on RPE cells are implicated in a number of age-associated and early-onset forms of macular disease including recessive Stargardt disease, ELOVL4-related disease, best macular dystrophy and age-related macular degeneration. The broad objectives of the studies proposed in this application are to understand mechanisms by which bisretinoids of retina contribute to disease processes that threaten vision. We will address both the bisretinoid pigments constituting RPE lipofuscin and the bisretinoid precursors of these compounds in photoreceptor cells. This work will elucidate therapeutic avenues. Additionally since bisretinoids are the major source of fundus autofluorescence, these studies will contribute to clinical interpretations of fundus autofluorescence images. In Specific Aim 1, we will investigate our preliminary observation that photo-cleavage of bisretinoid releases the small dicarbonyls (methylglyoxal and glyoxal) that are responsible for advanced glycation end-product (AGE) - modifications of proteins. Proteins in Bruch's membrane and drusen are prone to AGE-modification. Aging changes in Bruch's membrane are also considered to contribute to onset of age-related macular degeneration. We propose that, unlike the case in diabetes, AGE-adducts in Bruch's membrane form as a consequence of dicarbonyl release from overlying RPE. In Specific Aim 2, we will probe for evidence of photooxidation-associated photodegradation of RPE bisretinoid in vivo. In Specific Aim 3 we will explore the bisretinoid fluorophores located in photoreceptor cell outer segments that are precursors of RPE lipofuscin. We will probe the propensity for increased formation of bisretinoid in impaired photoreceptor cells; we will test the ability of photoreceptor bisretinoids to mediate oxidation of lipid in photoreceptor cells and photo-damage; and we will demonstrate conditions under which photoreceptor bisretinoids can contribute to hyperautofluorescence in fundus autofluorescence images. These aims will be achieved by the use of animal and in vitro models and by employing biochemical and histological approaches. Completion of this research will advance our comprehension of drusen formation and will elucidate links between RPE lipofuscin and the aging changes in Bruch's membrane that can be a prelude to AMD. These studies will contribute to an understanding of how light and oxidative mechanisms are factors in photoreceptor cell death in monogenic retinal disorders.