The Neurodevelopmental Consequences of Genomic Stress

摘要： 神经元中DNA损伤的异常积累是神经元中DNA损伤的共同病理特征。 神经退行性疾病（阿尔茨海默病，帕金森病），它也与正常 衰老转基因小鼠模型进一步证明了在哺乳动物中对DNA修复的特定要求。 神经发生还已知基因组应激，由正在进行的转录和代谢副产物引起， 在衰老中起作用，因为它有助于DNA损伤的积累。以前，研究衰老在 人类的大脑一直是令人难以置信的挑战;死后组织很难获得， 到纵向研究。此外，对衰老的研究通常着眼于生命后期的模式生物， 这意味着很少有人评估压力暴露如何影响大脑的发育，以及如何影响大脑的发育。 最终影响衰老过程。 随着人类诱导多能干细胞（hiPSC）的发展，我们现在有一个体外模型， 它为研究正常和病理性神经发育模型提供了工具。 这项研究的目的是确定基因组应激和随后的DNA损伤如何改变多样性 在hiPSC衍生的神经祖细胞（NPC）及其后代中。目标一是确定NPC 对复制应激具有抗性，以及与同基因成纤维细胞相比，这种抗性是否是NPC独有的， 星形胶质细胞和神经元。多光谱成像流式细胞术、免疫印迹和细胞周期分析都将在 用于通过研究DNA损伤修复蛋白来阐明抗性机制。该目的是 建立在初步数据表明NPC对复制应激具有抵抗力并且特别易感的基础上 转录相关的基因组应激;显示DNA损伤，细胞周期停滞和DNA 损伤信号激活。目的二试图确定基因组应激是否促进神经炎症 通过单细胞RNA-seq和人Luminex测定来确定表型。基于hiPSC的神经发生提供了 人类模型系统，以了解衰老和神经退行性变中基因组应激相关机制。这 一项研究将确定神经发育压力如何使神经元亚群更容易发生神经变性 通过神经炎症表型，并最终导致随后的研究集中在保护NPC 以及它们的后代免受基因组应激的影响，以防止加速衰老和神经退行性疾病。

ABSTRACT: The abnormal accumulation of DNA damage in neurons is a shared pathological feature among neurodegenerative diseases (Alzheimer's disease, Parkinson's disease) and it is also correlated with normal aging. Transgenic mouse models further demonstrate a specific requirement for DNA repair during neurogenesis. It is also known that genomic stress, arising from ongoing transcription and metabolic byproducts, plays a role in aging because it contributes to the accumulation of DNA damage. Previously, studying aging in the human brain has been incredibly challenging; post-mortem tissue is difficult to acquire and is not amenable to longitudinal studies. Furthermore, studies of aging have typically looked at model organisms later in life, meaning that few have assessed how stress exposure impacts the development of the brain and how that ultimately affects the aging process. With the development of human-induced pluripotent stem cells (hiPSCs), we now have an in vitro model of human neurodevelopment that provides a tool to study models of normal and pathological neurodevelopment. The goal of this study is to determine how genomic stress and subsequent DNA damage alters the diversity among hiPSC-derived neural progenitor cells (NPCs) and their progeny. Aim one seeks to determine how NPCs are resistant to replicative stress and whether this resistance is unique to NPCs compared to isogenic fibroblasts, astrocytes, and neurons. Multispectral imaging flow cytometry, immunoblotting, and cell cycle analysis will all be used to elucidate the mechanisms of resistance by investigating DNA damage repair proteins. This aim is founded on preliminary data indicating that NPCs are resistant to replication stress and particularly susceptible to transcription associated genomic stress; showing an increase in DNA damage, cell cycle arrest, and DNA damage signaling activation. Aim two seeks to determine if genomic stress promotes neuroinflammatory phenotypes through single cell RNA-seq and the human Luminex assay. hiPSC-based neurogenesis provides a human model system to understand genomic stress related mechanisms in aging and neurodegeneration. This study will define how neurodevelopmental stress may predispose subsets of neurons for later neurodegeneration through neuroinflammatory phenotypes, and ultimately lead to consequent studies focused on protecting NPCs and their progeny from genomic stress to prevent accelerated aging and neurodegenerative disease.