Novel strategies for induction of aging in human iPSC-derived lineages towards improved models of late-onset diseases

项目摘要 大多数神经退行性疾病和许多其他人类疾病(如癌症)的主要风险因素 就是老了。研究迟发性疾病的一个主要挑战是对衰老的准确描述 该疾病的体外和体内模型的研究背景。这在一定程度上是由于对 衰老过程的细胞和分子特征。我们之前定义了一组细胞变化 与来自老年捐赠者的细胞老化有关，并证明这些与年龄相关的特征被恢复 通过重新编程为诱导的多能干细胞(IPSC)并在 在重新分化为IPSC来源的细胞时，处于“恢复活力”的状态。这一现象，虽然从一个 科学观点，也是使用IPSC研究年龄相关性的具体障碍 精神错乱。我们的研究小组证明，这些老化特征可以重新引入IPSC来源的细胞 通过简单表达孕激素，一种突变型的层蛋白A(LMNA)蛋白，负责 早衰症Hutchinson-Gilford Progeria。然而，使用一种致病物质诱导细胞老化 因子可能不是生理老化的真实代表，并可能导致 对衰老相关疾病进行建模。因此，有必要开发一种准确的细胞模型 重现生理老化的背景。我们的假设得到了有希望的初步结果的支持，我们的假设是 细胞衰老是通过特定的遗传和表观遗传变化来促进的，这些变化可以被用来触发衰老 迟发性疾病模型中的细胞状态。在这里，我们建议开发一种新的年龄建模方法 在IPSC衍生的神经元谱系中，有三个特定的步骤。首先，我们的目标是产生一个全面的 用原代成纤维细胞和脑组织表征衰老细胞的遗传和表观遗传学特征 来自年轻和老年捐赠者的，这将提供不同细胞系的基因组老化签名。到时候我们会的 在重新编程期间监控这些签名，并确定细胞年龄的候选决定因素。我们会 通过有针对性的实验以及在独立样本上验证这些签名。第二，使用 我们先前识别的衰老细胞特征与新发现的 遗传和表观遗传衰老标记，我们将设计最佳的策略来诱导细胞衰老。第三，我们将 使用这些策略来研究细胞年龄对帕金森病进展和病理的影响 (PD)使用来自遗传型PD患者的IPSC衍生的多巴胺神经元。这两个都将被使用 用于体外研究帕金森病的细胞表现以及移植到帕金森病后的体内表现 评估诱导衰老对体内细胞行为和功能的影响的小鼠模型。 这项研究的结果将提供对遗传和表观遗传学的更全面的理解 人类衰老的基础变化，更重要的是，提供了一种在IPSC中诱导衰老背景的方法 人类疾病的模型。

Project Summary The primary risk factor for most neurodegenerative diseases and many other human ailments such as cancer is old age. A major challenge in studying late-onset diseases is the accurate representation of the aging context in both in vitro and in vivo models of the disease. This is due in part to a lack of understanding of the cellular and molecular characteristics of the aging process. We previously defined a set of cellular changes associated with aging in cells from old donors and demonstrated that these age-related hallmarks are restored to a youthful state through reprogramming into induced pluripotent stem cells (iPSC) and maintained in such “rejuvenated” state upon re-differentiation into iPSC-derived cells. This phenomenon, while fascinating from a scientific perspective, also represents a concrete barrier for the use of iPSC for studying age-dependent disorders. Our group demonstrated that these aging characteristics can be reintroduced into iPSC-derived cells through simple expression of progerin, a mutant form of the lamin A (LMNA) protein that is responsible for the premature aging disorder Hutchinson-Gilford Progeria. However, inducing cellular age using a disease-causing factor may not be a faithful representation of physiological aging and potentially lead to pathological artifacts in modeling aging-related diseases. It is therefore necessary to develop a cellular model that will accurately reproduce the physiological aging context. Our hypothesis, supported by promising preliminary results, is that cellular aging is promoted by specific genetic and epigenetic changes that can be utilized to trigger an aged cellular state in models of late-onset disease. Here, we propose to develop a new approach for modeling age in iPSC-derived neuronal lineages in three specific steps. First, we aim to generate a comprehensive characterization of genetic and epigenetic features of aged cells using primary fibroblasts and brain tissues from young and old donors, which will provide genomic aging signatures of different cell lineages. We will then monitor those signatures during reprogramming and identify candidate determinants of cellular age. We will validate these signatures by targeted experiments as well as on independent samples. Second, using a combination of our previously identified cellular hallmarks of aging in conjunction with the newly identified genetic and epigenetic aging markers we will design optimal strategies to induce cellular aging. Third, we will use these strategies to study the impact of cellular age on the progression and pathology of Parkinson disease (PD) using iPSC-derived dopamine neurons from patients with genetic forms of PD. These will both be used for in vitro studies characterizing cellular PD manifestations as well as in vivo upon transplantation into PD mouse models to assess the impact of induced aging on cellular behavior and function in vivo. The results from this study will provide a more comprehensive understanding of the genetic and epigenetic changes that underlie human aging and importantly, provide a methodology to induce aging context in iPSC models of human diseases.