Genomic Instability-Induced Senescence in Brain Aging and Alzheimer's Disease

项目摘要/摘要 发展慢性疾病的最大风险因素，包括神经退行性疾病，如 阿尔茨海默病(AD)，即年龄。现在有大量证据表明，衰老过程可以由DNA驱动 损伤，这是普遍存在的，是许多不利的细胞命运的原因，如细胞凋亡和细胞衰老。 DNA损伤及其错误修复的一个主要病理后果是DNA碱基突变 替换为非常大的染色体改变。随着先进的单细胞技术的出现，它 最近发现，在人类衰老过程中，神经元中的突变会以更高的频率积累 在受神经退化影响的大脑中。这与使用细胞遗传学方法的早期发现是一致的。 表明与阿尔茨海默病相关的非整倍体增加。我们最近表明，这一数字增加了 非整倍体诱导细胞衰老，包括衰老相关分泌表型(SASP)。 衰老细胞的清除对阿尔茨海默病进展的各个方面都有有益的影响 在衰老相关生物学中，细胞衰老是一种新出现的重要细胞命运 神经退行性变。最近的研究表明，涉及非神经细胞(Neun-Negative)的细胞过程 细胞)在人类和小鼠模型中都对AD的病理有重要贡献。学习规范性 在小鼠的衰老过程中，我们发现从小鼠分离的NeuN阴性细胞中有显着的非整倍体积累。 大脑皮层，但不是来自老年小鼠的小脑。 在这个应用程序中，我们建议以我们的观察为基础来测试Neun-Negative的假设 大脑中的细胞特别容易受到与年龄相关的非整倍体和大规模基因组积累的影响 不稳定促进衰老。基因组不稳定诱导的衰老Neun阴性细胞可能无法 完成神经护理功能和/或获得神经毒性特性，并特别有害 对于AD的进展。为了验证我们的假设，在目标1中，我们将建立来自 正常衰老和AD时人的大脑皮质和海马区。使用新开发的高灵敏单片 基于细胞的分析包括多重置换扩增(SCMDA)和多色间期DNA-RNA- FISH(IDR-FISH)为了测量非整倍体和原位衰老，我们将比较Neun阴性和Neun- 从健康的老年供者到年龄匹配的AD患者和年轻人对照组的阳性细胞。这将确立 人类脑老化过程中非整倍体和/或其他形式的基因组不稳定性的综合分析 与AD和AD类型痴呆相关的区域和细胞类型。在目标2中，使用原电池以及 从晚发性AD患者和无疾病对照中诱导出人类多能干细胞(HiPSCs)，我们将 利用共培养模型研究非整倍体诱导的神经元衰老对细胞的非自主影响。 两个单独或共同促进非整倍体细胞死亡的小分子将被测试其潜力。 目的：杀死非整倍体诱导的衰老星形胶质细胞，并改善其对神经元的潜在有害影响。 在全球范围内，这些研究将提供关于老年人和老年人大规模基因组不稳定性的详细新知识 AD脑及其与衰老细胞的功能联系可能是AD的致病因素。这将开启新的 新干预措施的可能性，例如，使用新一代老年科学干预措施。

Project Summary/Abstract The largest risk factor for developing chronic disease, including neurodegenerative diseases, such as Alzheimer’s disease (AD), is age. There is now abundant evidence that aging processes can be driven by DNA damage, which is ubiquitous and a cause of many adverse cell fates, such as apoptosis and cellular senescence. A major pathologic consequence of DNA damage and its erroneous repair is DNA mutation, from base substitutions to very large chromosomal alterations. With the emergence of advanced single-cell technology it has recently been shown that mutations accumulate in neurons during human aging at a frequency that is higher in brains affected by neurodegeneration. This is in keeping with earlier findings using cytogenetic methods indicating increased aneuploidy associated with Alzheimer’s disease. We have recently shown that increased aneuploidy induces cellular senescence, including the senescence-associated secretory phenotype (SASP). Clearance of senescent cells has shown beneficial effects on various aspects of AD disease progression implicating cellular senescence as an emerging and important cell fate in the biology of age-related neurodegeneration. Recent work suggests that cellular processes involving non-neuronal cells (NeuN-negative cells) significantly contribute to the pathology of AD in both humans and mouse models. Studying normative aging in the mouse we identified a significant accumulation of aneuploidy in NeuN-negative cells isolated from the cerebral cortex, but not from the cerebellum of old mice. In this application we propose to build on our observations to test the hypothesis that NeuN-negative cells in the brain are particularly susceptible to age related accumulation of aneuploidy and large-scale genomic instability promoting senescence. Genomic instability-induced senescent NeuN-negative cells could fail to accomplish their neuronal nursing functions and/or acquire neurotoxic properties and be particularly detrimental for AD progression. To test our hypothesis, in Aim 1 we will establish the genomic landscape of cells from the human cortex and hippocampus during normal aging and AD. Using newly developed, highly sensitive single cell-based assays including multiple displacement amplification (SCMDA) and multicolor interphase DNA-RNA- FISH (iDR-FISH) to measure aneuploidy and senescence in situ we will compare NeuN-negative and NeuN- positive cells from healthy aged donors to age-matched AD patients and young adult controls. This will establish a comprehensive analysis of aneuploidy and/or other forms of genomic instability during human aging in brain regions and cell types associated with AD and AD-type dementias. In Aim 2 using primary cells as well as induced human pluripotent stem cells (hiPSCs) from late-onset AD patients and disease-free controls, we will study the cell non-autonomous effects of aneuploidy-induced senescence on neurons using co-culture models. Two small molecules that promote the death of aneuploid cells, alone or together, will be tested for the potential to kill aneuploidy-induced senescent astrocytes and ameliorate their potential detrimental effects on neurons. Globally, these studies will provide detailed new knowledge of large-scale genomic instability in aged and AD brain and its functional link to senescent cells as a possible causal factor in AD. This will open up new possibilities for novel interventions, e.g., using the new generation of geroscience interventions.