Establishment of a human, age-specific model for axon growth and regeneration studies

建立用于轴突生长和再生研究的人类特定年龄模型

基本信息

批准号：
9978418
负责人：
Darcie Leann Moore
金额：
$ 44.1万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-05-01 至 2022-10-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9978418
关键词：
ASCL1 gene Adult Affect Age Animal Model Animals Axon Behavior Biological Birth Bypass Cell Aging Cell Culture System Cell Maintenance Cells Clinical Trials Communities DNA Damage Development Doxycycline Embryo Encyclopedia of DNA Elements Environment Epigenetic Process Expression Profiling Fibroblasts Gene Expression Genes Genetic Transcription Glutamates Growth Homologous Gene Human In Vitro Injury Knock-out Lead Length Life Maintenance Measures Medical Methods Mitochondria Modeling Motor Neurons Mus Natural regeneration Neonatal Neuraxis Neurons PTEN gene Paralysed Pathway interactions Phenotype Play Population Protocols documentation Research Research Personnel Resources Rodent Role Somatic Cell Spinal cord injury System Testing Treatment Efficacy Validation Work aged axon growth axon regeneration cell age disability excitatory neuron human model human tissue in vitro Model induced pluripotent stem cell juvenile animal knock-down novel overexpression pluripotency postnatal psychologic regenerative response screening social stem cells targeted treatment telomere therapeutic target tool transcription factor transdifferentiation young adult

项目摘要

Human spinal cord injury leads to life-long disability, with limited treatment options despite decades of research in rodents. Recent evidence indicates that there are fundamental differences in gene expression between human and mouse, suggesting a critical need for identification of novel or human-specific candidates as targets to increase axon regeneration following spinal cord injury. Additionally, in vitro studies of axon growth are typically performed in neurons from embryonic or young postnatal rodents, ages with increased intrinsic growth ability, and a very different transcriptional landscape from adult neurons. Thus, we propose to create an age-relevant, human model for axon growth and regeneration. Recent technological advances now allow for human adult somatic cells to be reprogrammed into stem cells, and then differentiated into neurons. However, reprogramming these cells through pluripotency has been shown to erase the “age” of the original cell, resulting in a more embryonic identity of the resulting neuron. Using a direct reprogramming protocol, we will transdifferentiate neonatal, ~35 year old, and ~70 year old human somatic cells directly into neurons through overexpression of specific neuronal factors, allowing for maintenance of the cell’s biological age identity. We will determine the maintenance of age in these cells at the level of the epigenetic age signature, gene expression, and markers of cellular aging. As the first study of axon growth in age-maintained human neurons, we will characterize the basal axon growth behavior on permissive and inhibitory substrates to identify if age plays a role in human axon growth ability. To test this new resource, we will target 20 different genes and pathways previously identified in other species to regulate axon growth for their conservation in function in human neurons. In rodents it has recently been shown that certain axon growth modifiers such as PTEN knockout, do not function similarly in aged animals as young animals (Geoffroy et al, 2016). Thus, we will utilize our age-maintained human iNs to determine if age drives changes in axon growth phenotypes. Completion of this research will 1) establish a new human, age-relevant cell culture system for axon growth studies, 2) identify genes and their regulators that have conserved functions in human neurons, and 3) determine the role age plays in axon growth ability. Identification of translational targets specifically in human neurons ultimately may lead to the development of effective therapeutic targets for human spinal cord injury.

人脊髓损伤导致终身残疾，而治疗方案有限啮齿动物的研究。最近的证据表明，基因表达存在根本差异在人与小鼠之间，表明对新颖或人类特异性候选者的迫切需要作为增加脊髓损伤后轴突再生的目标。另外，轴突的体外研究生长通常是在胚胎或年轻产后啮齿动物的神经元中进行的，年龄增加内在的生长能力以及与成人神经元的转录景观非常不同。那我们建议创建一个与年龄相关的人类模型，用于轴突生长和再生。现在的技术进步现在允许将人类的成年体细胞重编程为茎细胞，然后分化为神经元。但是，通过多能性重新编程这些细胞已经显示以消除原始细胞的“年龄”，从而产生了所得神经元的胚胎身份。使用直接重新编程的协议，我们将转变为新生儿，约35岁，约70岁通过特定神经元因子的过表达，人类体细胞细胞直接进入神经元，从而使维持细胞的生物年龄认同。我们将确定在这些细胞中的年龄维持表观遗传年龄特征，基因表达和细胞衰老标记的水平。作为轴突的首次研究维持年龄的人类神经元的增长，我们将表征允许性的基本轴突生长行为并抑制底物确定年龄是否在人轴突生长能力中起作用。为了测试这个新资源，我们将针对以前在其他物种中鉴定的20种不同基因和途径，以调节轴突生长以进行它们在人类神经元功能中的保护。在啮齿动物中，最近已经显示出某些轴突生长诸如PTEN敲除之类的修饰符在老年动物中不像年轻动物一样起作用（Geoffroy等人， 2016）。这是，我们将利用我们年龄增长的人类来确定年龄是否驱动轴突生长的变化表型。这项研究的完成将1）建立一个新的人类，年龄与年龄相关的轴突的细胞培养系统增长研究，2）鉴定构成人类神经元功能的基因及其调节因子，3）确定年龄在轴突生长能力中的作用。专门在人类中识别翻译目标神经元最终可能导致为人脊髓损伤的有效治疗靶标发展。