Transfer: Molecular Mechanisms of Glaucoma

摘要 青光眼是一组以缓慢进行性视网膜神经节细胞丢失为特征的视神经病变。 视网膜神经节细胞(RGCs)，视神经变性，并因此导致视力丧失。据估计，有70多个 目前有100万人受到青光眼的影响，其中约10%是双眼失明，使其成为 导致世界上不可逆转的失明的主要原因。有几种类型的青光眼，但在美国， 大多数病例是原发性开角型青光眼(POAG)，这是一种在非洲特别流行的变种 美国人。POAG被认为是一种由多种遗传和环境因素引起的复杂疾病 互动。眼压升高和年龄这两个主要危险因素与眼压升高的程度和年龄有关 研资局损失率。基因组学的最新进展使研究人员能够描述基因关联 青光眼的风险和特定的基因组基因座之间的关系。然而，尽管经过多年的研究，分子 青光眼的发病基础尚不清楚，导致其发展的因素也不完全清楚。 特色化的。在我们最近的工作中，我们使用了小鼠模型来研究Six6风险的分子影响。 青光眼的发展和RGC死亡的变异。我们观察到，当眼压升高时， Six6增加并直接调节p16INK4a的表达，导致视网膜节细胞衰老加速 最有可能直接导致RGC死亡。编码p16INK4a的基因CDKN2a位于肿瘤内。 人类染色体9p21上的抑制子基因。这个基因座已经被几个小组独立鉴定出来 在不同人群样本中与POAG的关联度最高。9p21中的基因调控 基因座已经在许多实验室得到了广泛的研究；然而，分子分析从未被 专门针对青光眼患者进行的。鼠标六号位于141号位置，因此非常适合 研究该变异在青光眼中的分子作用。然而，由于在小鼠中缺乏非风险变异体 菌株，不可能研究这两个变异体对RGC发育和退化的贡献。 老鼠模型。在这里，我们建议使用CRISPR/Cas9技术来改造携带人类非风险的小鼠 研究Six6基因各变异在青光眼发病机制中的作用。我们将使用最先进的 分子和细胞技术研究视网膜发育和视网膜节细胞变性 眼压是Six6的特定变种的函数。此外，我们还将研究分子 利用转录和表观基因组学研究p16INK4a在疾病病因学中的上调机制 方法，我们将提出方法，以下调其在眼睛的表达。建议数 各种方法的结合将推动对青光眼病因的总体理解，并提供 发展新颖的、个性化的、提高生活质量的治疗策略的分子基础 适用于青光眼患者。

SUMMARY Glaucoma is a group of optic neuropathies characterized by slow, progressive loss of retinal ganglion cells (RGCs), optic nerve degeneration and as a consequence, vision loss. It has been estimated that more than 70 million people are currently affected by glaucoma with approximately 10% being bilaterally blind, making it the leading cause of irreversible blindness in the world. Several glaucoma categories exist, but in United States, most of the cases are primary open-angle glaucoma (POAG), a variant particularly prevalent amongst African Americans. POAG is recognized as a complex disease in which multiple genetic and environmental factors interact. The two leading risk factors, increase intraocular pressure (IOP) and age are related to the extent and rate of RGC loss. Recent advances in genomics have allowed researchers to describe genetic association between the risk of glaucoma and specific genomic loci. Nevertheless, despite years of research, the molecular basis of glaucoma is poorly understood and the factors contributing to its progression have not been fully characterized. In our recent work, we have used a mouse model to study the molecular impact of Six6 risk variant in development of glaucoma and in RGC death. We observed that upon increased IOP, expression of Six6 increases and directly regulates the expression of p16Ink4a, leading to enhanced senescence in RGCs and most likely directly causing RGC death. The gene encoding p16INK4a, CDKN2A, lies within the tumor suppressor locus on human chromosome 9p21. This locus has been independently identified by several groups to have the highest association with POAG in different population samples. Gene regulation within the 9p21 locus has been extensively studied in many laboratories; however, a molecular analysis has never been performed specifically in relation to glaucoma. Mouse Six6 harbors His at position 141 and therefore is ideal to investigate the molecular role of this variant in glaucoma. However, due to the lack of non-risk variant in mouse strains, it is not possible to study the contribution of both variants on RGC development and degeneration in a mouse model. Here, we propose to use CRISPR/Cas9 technology to engineer mice harboring human non-risk variant of SIX6 to study the impact of each variant in pathogenesis of glaucoma. We will use state-of-the art molecular and cellular technologies to study retinal development and RGC degeneration upon elevated intraocular pressure as a function of the particular variant of SIX6. In addition, we will investigate the molecular mechanisms of p16Ink4a upregulation in the etiology of the disease using transcriptomic and epigenomic approaches, and we will propose the methodology to downregulate its expression in the eye. The proposed combination of approaches will move forward the general understanding of the etiology of glaucoma and provide the molecular basis for development of novel, personalized, therapeutic strategies to improve the quality of life for glaucoma patients.