Ferroxidases in RPE iron transport

RPE 铁转运中的铁氧化酶

• 批准号: 6826940
• 负责人: JOSHUA L DUNAIEF
• 金额: $ 31.7万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6826940
• 关键词: Adenoviridae; angiogenesis; biological models; confocal scanning microscopy; ferroxidase; gene induction /repression; gene targeting; genetically modified animals; immunoelectron microscopy; in situ hybridization; ion transport; iron; laboratory mouse; model design /development; protein localization; retina degeneration; retinal pigment epithelium; transfection /expression vector; vascular endothelial growth factors

DESCRIPTION (provided by applicant): Iron is necessary in the retina for oxidative phosphorylation, membrane biogenesis and many other metabolic processes, but can also produce oxidative stress if improperly regulated, leading to cell death. Iron plays a significant role in retinal degeneration, as follows: 1) Iron toxicity causes retinal degeneration following direct entry of iron into the eye carried by an intraocular foreign body 2) Iron has been implicated in the RCS rats' retinal degeneration, in which the mechanism of iron overload is indirect. 3) Human AMD retinas have more iron than age-matched controls, suggesting that iron overload may play a role in AMD pathogenesis. 4) Inherited defects in iron metabolism result in retinal degeneration in Friedreich's ataxia, pantothenate kinase associated neuropathy, and aceruloplasminemia. The focus of the current proposal is to understand the functions in ocular iron homeostasis of the ferroxidases ceruloplasmin (Cp) and its homologue hephaestin (Heph). In Aim 1, the retinal localization of Cp and Heph will be assessed. In Aim 2, an RPE-specific Heph knockout will be generated. This will determine whether RPE Heph expression is necessary for RPE iron homeostasis. It should also provide a mouse model of retinal degeneration that like our Cp-/-Heph-/- mice, has subretinal neovascularization, but unlike Cp-/-Heph-/- mice, has a normal lifespan. In addition it will provide reagents for generating RPE-specific knockouts for the eye research community. In Aim 3, the retinal degeneration in Cp-/-Heph-/- and Cp-/-RPE-specific Heph-/- mice will be characterized. Both oxidative stress and mechanisms of neovascularization will be studied. In Aim 4 an RPE in vitro system will be used to determine whether Cp and Heph mediate RPE iron import or export, and assess the polarity of transport. These studies are important because: 1) The mechanism of retinal degeneration in aceruloplasminemia is not known. 2) The retinal functions of Cp and Heph are not known. 3) Our preliminary studies show that Cp-/-Heph-/- mice will provide a model for several features of AMD, including subretinal neovascularization. 4) These mice will provide a model of neurodegeneration caused by age-dependent increases in iron levels, which recent reports suggest may contribute to the pathogenesis of Alzheimer's and Parkinson's diseases and AMD. 5) These mice will provide the opportunity to test iron chelators and antioxidants as potential treatments/preventions for those age-related diseases.

描述(申请人提供)：铁是视网膜氧化磷酸化、膜生物发生和许多其他代谢过程所必需的，但如果调控不当也会产生氧化应激，导致细胞死亡。铁在视网膜变性中起着重要的作用，主要表现在：1)铁中毒导致眼内异物携带铁直接进入眼内，引起视网膜变性；2)铁与RCS大鼠的视网膜变性有关，其机制是间接的。3)人类AMD视网膜中铁含量高于年龄匹配的对照组，提示铁超载可能在AMD的发病机制中起一定作用。4)遗传性铁代谢缺陷可导致Friedreich‘s共济失调视网膜变性、泛酸激酶相关性神经病和无纤维蛋白原血症。本研究的重点是了解铜蓝蛋白铁氧合酶(CP)及其同系物肝素(Heph)在眼部铁稳态中的作用。在目标1中，将评估CP和HEPH的视网膜定位。在AIM 2中，将产生RPE特异性的HEPH基因敲除。这将决定RPE Heph的表达是否对RPE铁稳态是必需的。它还应该提供一种视网膜变性的小鼠模型，与我们的CP-/-HEPH-/-小鼠一样，具有视网膜下新生血管，但与CP-/-HEPH-/-小鼠不同，具有正常寿命。此外，它还将为眼科研究界提供产生RPE特异性基因敲除的试剂。在目标3中，将描述CP-/-HEPH-/-和CP-/-RPE特异性HEPH-/-小鼠的视网膜变性。氧化应激和新生血管的机制都将被研究。在目标4中，将使用RPE体外系统来确定CP和HEPH是否介导RPE铁的输入或输出，并评估运输的极性。这些研究很重要，因为：1)无纤维蛋白溶血症性视网膜变性的机制尚不清楚。2)CP和HEPH的视网膜功能未知。3)我们的初步研究表明，CP-/-HEPH-/-小鼠将为AMD的几个特征提供一个模型，包括视网膜下新生血管。4)这些小鼠将提供一种由铁水平随年龄增加而引起的神经退行性变的模型，最近的报告表明，这可能与阿尔茨海默氏症、帕金森氏病和AMD的发病机制有关。5)这些小鼠将提供测试铁络合剂和抗氧化剂作为这些年龄相关疾病的潜在治疗/预防的机会。