Understanding OPA1 mutation-driven dominant optic atrophy using human PSC-derived retinal ganglion cells

项目总结/摘要 显性视神经萎缩（DOA）是最普遍的遗传性视神经病变，影响大约1：12，000至1：12，000。 1：50，000人全世界。DOA患者表现出视网膜神经节细胞（RGC）变性，这导致 进行性双侧视力丧失。大多数DOA病例是由视神经萎缩基因突变引起的 1（OPA1），一种编码靶向线粒体内膜的蛋白质的核基因。有趣的是， 尽管OPA1在所有人体组织中普遍表达，但RGC似乎是受OPA1影响的唯一细胞类型。 OPA1突变。因此，研究人RGCs中的DOA以了解其病理学变化是必要的。 这些细胞中存在的机制使它们特别容易退化。然而，研究 由于原始视网膜组织的稀少和RGC的缺乏， 仅占视网膜细胞总数的2%。这项建议旨在解决以下重大未满足的需求： 开发DOA的人类RGC模型。干细胞技术的进步使我们的实验室能够 常规地从人多能干细胞（hPSC）衍生的3D视网膜类器官培养物产生人RGC。 此外，我已经通过使用基因编辑技术和通过重编程建立了OPA1突变hPSC系 DOA患者的外周血细胞。将这些OPA1突变hPSC系分化成视网膜类器官将有助于 提供了在真实的人RGC中建立DOA疾病模型的第一个机会。在拟议的研究中， 我将使用OPA1突变的hPSC衍生的人RGC群体来研究OPA1的病理机制。 突变介导的RGC变性。由于OPA 1在促进线粒体融合中起重要作用， 维持嵴的完整性，稳定呼吸链的超级复合体，RGC死亡 在DOA患者中观察到的这种情况可能是线粒体缺陷的结果，这可能导致能量供应不足， 增加的氧化应激和/或细胞色素C渗漏到细胞质中。我会研究线粒体 动态，嵴结构和代谢状态的OPA1突变的RGCs，以描绘是否在这些变化， 在DOA患者中观察到的RGC变性的基本过程。这项研究的结果将 提高我们对影响DOA患者RGCs的病理机制的理解， 开发可以保护或挽救DOA患者视力的治疗方法。此外，我们的发现可以 为其他神经退行性疾病提供了重要的见解，这些疾病与 doa.

PROJECT SUMMARY/ABSTRACT Dominant optic atrophy (DOA) is the most prevalent genetic optic neuropathy, affecting roughly 1:12,000 to 1:50,000 individuals worldwide. DOA patients exhibit retinal ganglion cell (RGC) degeneration, which leads to progressive bilateral vision loss. The majority of DOA cases are caused by mutations in the gene optic atrophy 1 (OPA1), a nuclear gene that encodes a protein targeted to the inner mitochondrial membrane. Interestingly, although OPA1 is ubiquitously expressed in all human tissues, RGCs appear to be the only cell type affected by OPA1 mutations. It is therefore essential to study DOA in human RGCs in order to understand the pathological mechanisms present in these cells that render them particularly prone to degeneration. However, studies of human RGCs have been historically difficult due to the rarity of primary retinal tissues and scarcity of RGCs, which only comprise ~2% of the total retinal cells. This proposal seeks to address the significant unmet need for developing human RGC models of DOA. Advances in stem cell technology have enabled our laboratory to routinely produce human RGCs from human pluripotent stem cell (hPSC)-derived 3D retinal organoid cultures. In addition, I have established OPA1 mutant hPSC lines by using gene editing technology and by reprogramming DOA patients’ peripheral blood cells. Differentiating these OPA1 mutant hPSC lines into retinal organoids will provide the first opportunity to establish DOA disease models in authentic, human RGCs. In the proposed study, I will use OPA1 mutant hPSC-derived human RGC populations to investigate pathological mechanisms of OPA1 mutation-mediated RGC degeneration. As OPA1 plays significant roles in promoting mitochondrial fusion, maintaining the integrity of the cristae, and stabilizing super complexes of the respiratory chain, RGC death observed in DOA patients is likely a result of mitochondrial defects that can lead to an insufficient energy supply, increased oxidative stress, and/or leakage of cytochrome c into the cytoplasm. I will investigate the mitochondrial dynamics, cristae structure, and metabolic state of OPA1-mutant RGCs to delineate whether changes in these fundamental processes underly the RGC degeneration observed in DOA patients. Findings from this study will advance our understanding of the pathological mechanisms affecting DOA patients’ RGCs and facilitate the development of therapies that can preserve or rescue vision in DOA patients. Additionally, our findings could provide important insights into other neurodegenerative diseases that share common metabolic deficiencies with DOA.