Ferroxidases in RPE Iron Transport

铁是视网膜氧化磷酸化、膜生物发生和视黄醇异构化所必需的。 但如果调控不当，也会产生氧化应激，导致细胞死亡。这可能有助于 视网膜疾病如下：1)铁中毒导致直接进入视网膜的铁变性。 眼内异物携带的眼睛。2)人类AMD视网膜比年龄匹配的对照组含有更多的铁， 提示铁超载可能在AMD的发病机制中起一定作用。3)铁氧合酶的遗传缺陷 铜蓝蛋白(CP)可导致视网膜铁蓄积和早发性黄斑变性。4)小鼠 CP及其同源基因Hephestn(Heph)的突变具有年龄相关性的视网膜铁超载和 变性，具有许多类似于AMD的特征，包括视网膜下新生血管。后两者 这些分数表明CP和HEPH对视网膜健康很重要。其他器官的证据表明，CP 或者，Heph可以与质膜铁转运蛋白(FPN)合作，从细胞中输出铁。 这项建议的目标是增加对CP、HEPH和FPN在视网膜铁中的作用的理解 体内平衡及其分泌激素海普西丁(HepC)的调节丙型肝炎病毒在视网膜(AS)中产生 以及肝脏)，并触发FPN的内化和降解。丙型肝炎病毒可以作为来自 视网膜细胞感知铁过量(如光感受器)以降解FPN并限制RPE和RPE的铁转移 穆勒细胞。 我们现有的CP/Heph双突变体和HepC-/-小鼠表明，这三种蛋白对视网膜至关重要 铁的动态平衡和健康，但很少提供有关体内蛋白质的特定功能的信息 视网膜。有条件的鼠标淘汰技术(lox/cre)提供了机会来确定这些 蛋白质在特定的视网膜细胞类型中发挥作用，以及细胞间铁转移是如何执行和调节的。 在Aim1中，Heeph的光感受器特异性功能，可能是阻止PR铁的“铁释放阀” 将在CP-/-背景下使用HEPH条件基因敲除来调查超载。在《目标2》中，铁杆 FPN的转运功能将用RPE和光感受器特异的条件性基因敲除小鼠来研究。 在目标3中，将在基因敲除和条件性基因敲除小鼠中研究HepC的视网膜功能。这些 研究之所以重要，是因为：1)它们将提供有关细胞类型特定功能的新信息 HEPH、FPN和HEPC以及控制视网膜铁稳态的细胞间铁转移途径。2) 条件性基因敲除小鼠可能为AMD的几个特征提供模型，包括视网膜下 新生血管的同时避免了我们现有的CP/HEPH双重病例中限制寿命的脑铁超载 突变的小鼠。

Iron is necessary in the retina for oxidative phosphorylation, membrane biogenesis and retinol isomerization, but can also produce oxidative stress if improperly regulated, leading to cell death. This can contribute to retinal disease as follows: 1) Iron toxicity causes rapid retinal degeneration following direct entry of iron into the eye carried by an intraocular foreign body. 2) Human AMD retinas have more iron than age-matched controls, suggesting that iron overload may play a role in AMD pathogenesis. 3) Inherited defects in the ferroxidase ceruloplasmin (Cp) result in retinal iron accumulation and early onset macular degeneration. 4) Mice with mutation in Cp and its homolog hephaestin (Heph) have an age-dependent retinal iron overload and degeneration with a number of features similar to AMD, including subretinal neovascularization. The latter two points indicate that Cp and Heph are important for retinal health. Evidence from other organs suggests that Cp or Heph can cooperate with the plasma membrane iron transporter ferroportin (Fpn) to export iron from cells. The goal of this proposal is to increase understanding of the roles of Cp, Heph and Fpn in retinal iron homeostasis and their regulation by the secreted hormone hepcidin (Hepc). Hepc is produced in the retina (as well as the liver) and triggers internalization and degradation of Fpn. Hepc may serve as a message from retinal cells sensing iron excess (such as photoreceptors) to degrade Fpn and limit iron transfer from RPE and Muller cells. Our existing Cp/Heph double mutant and Hepc-/- mice indicate that these three proteins are critical for retinal iron homeostasis and health, but provide little information about the specific functions of the proteins within the retina. Conditional mouse knockout technology (lox/cre) affords the opportunity to determine how these proteins function within specific retinal cell types and how intercellular iron transfer is executed and regulated. In Aim1, the photoreceptor-specific functions of Heph, a possible "iron release valve" to prevent PR iron overload will be investigated using a Heph conditional knockout on a Cp-/- background. In Aim 2, the iron transport function of Fpn will be investigated using RPE and photoreceptor-specific conditional knockout mice. In Aim 3, the retinal function of Hepc will be investigated in knockout and conditional knockout mice. These studies are important because: 1) They will provide new information about the cell-type specific functions of Heph, Fpn and Hepc and the routes of intercellular iron transfer that control retinal iron homeostasis. 2) The conditional knockout mice are likely to provide models for several features of AMD, including subretinal neovascularization while avoiding the lifespan-limiting brain iron overload in our existing Cp/Heph double mutant mice.