Elucidating the developmental and molecular mechanism of chorioretinal anastomoses in a model of type 3 neovascular age-related macular degeneration

项目总结 新生血管性老年性黄斑变性(AMD)是老年人视力丧失和失明的主要原因。 老年人。在新生血管性AMD的亚型中，3型新生血管(3型NV)与 保护视力预后，其独特的特征是视网膜和脉络膜中的血管增殖。 晚期3型NV的一个明显特征是发生脉络膜视网膜吻合口(CRA)，异常 视网膜和脉络膜循环之间的直接联系。不幸的是，CRA可能会成为 复发性渗出，对现有的治疗方法有弹性。 CRA的发生机制尚不清楚。我们建立了3型新城疫的小鼠模型， 其中携带IV型胶原突变(Col4a1)的小鼠会出现人类疾病的特征，包括 浆液性视网膜脱离，出血，萎缩，并明显的CRA。与患者一样，Col4a1突变中的CRA 小鼠在成年后形成视网膜和脉络膜循环之间的大型新生融合， 让人联想到高流量的动静脉分流术。最近，反病毒规范已经成为一种强有力的 血管构型调节因子和控制房室识别的关键基因失调导致房颤的形成 分流。值得注意的是，规格化的房室规范已被证明可以逆转房室分流，尽管其流量很大 在某些动物模型中的状态。阐明反病毒规范在CRA中的作用可能会为两者提供新的见解 疾病的发病机制和治疗策略。我们的中心假设是Col4a1突变小鼠的CRA 房室分流来自视网膜或脉络膜，具有保守的发育起源。我们最近做了 支持直接通过色素眼的巩膜进行多光子成像 多光子显微镜。因此，这项跨学科研究提案的目标是： 1)优化经巩膜多光子显微镜脉络膜视网膜血管动力学活体成像；2) 确定Col4a1突变小鼠CRA的来源、血流和动静脉特性；以及3)识别 用单细胞RNA测序研究血管内皮细胞CRA的候选通路。 这个职业发展研究项目的长期目标是为调查人员提供一个交叉的 具备先进的活体成像和单细胞转录学方面的专业技能，使他能够 探讨眼血管发育不全的基本机制，以期发现新的治疗方法 血管疾病。

PROJECT SUMMARY Neovascular age-related macular degeneration (AMD) is a leading cause of vision loss and blindness in the elderly. Among the subtype of neovascular AMD, type 3 neovascularization (type 3 NV) is associated with a guarded visual prognosis and is uniquely characterized by vascular proliferation in both the retina and choroid. A defining feature of advanced type 3 NV is the development of chorioretinal anastomoses (CRA), abnormal direct connections between the retinal and choroidal circulations. Unfortunately, CRA can act as ‘hot spots’ for recurrent exudation which are resilient to available therapies. The developmental mechanism of CRA remains unclear. We established a murine model for type 3 NV, wherein mice harboring a mutation of type IV collagen (Col4a1) develop features of human disease including serous retinal detachment, hemorrhage, atrophy, and pronounced CRA. As in patients, CRA in Col4a1 mutant mice form in adulthood and mature into large de novo fusions between retinal and choroidal circulations, reminiscent of high-flow arteriovenous (AV) shunts. Recently, AV specification has emerged as a potent regulator of vascular patterning, and dysregulation of key genes governing AV identity induces formation of AV shunts. Notably, normalizing AV specification has been shown to reverse AV shunts despite their high-flow state in certain animal models. Elucidating a role for AV specification in CRA may offer novel insights into both disease pathogenesis and therapeutic strategies. Our central hypothesis is that CRA in Col4a1 mutant mice are AV shunts with a conserved developmental origin from either the retina or choroid. We have recently enabled multiphoton imaging of the choroid directly through the sclera of pigmented eyes using optimized multiphoton microscopy. The objectives of this inter-disciplinary research proposal are therefore to: 1) optimize intravital imaging of chorioretinal vascular dynamics with transscleral multiphoton microscopy; 2) determine the origin, blood flow, and arteriovenous identity of CRA in Col4a1 mutant mice; and 3) identify candidate pathways driving CRA in the endothelium using single cell RNA sequencing. The long-term goal of this career development research project is to provide the investigator with a cross- disciplinary skillset in advanced intravital imaging and single-cell transcriptomics that will allow him to investigate fundamental mechanisms of oculovascular dysgenesis toward the discovery of new therapies in vascular disease.