Identifying Molecular Ageing signatures of Neural Stem Cells to uncover novel Geroneuroprotectors

现在有一个需要和机会，以提高我们对NSC老化的动力学和分子机制的理解，以利用和控制成年人内源性NSC可塑性，以维持大脑功能在aging.Proposed工作计划：目标1：揭示NSC老化的分子特征我们已经产生了来自海马祖细胞暴露于年轻与老年血清的转录组数据。学生将使用生物信息学工具来生成NSC老化的分子特征。这些途径将被完善的认知相关的额外的转录组学数据，目前正在生成的老血清从低认知性能与血清从认知高性能在成人海马neurogenesis-dependent tasks.Objective 2：确定抗神经衰老化合物（geroneuroproectors）这一目标将启动临床可行的策略，以扭转目标1中确定的衰老诱导的变化。因此，药物再利用是绕过冗长的药物开发和安全性试验的有效方法。Thuret实验室最近成功地建立了第一个策略和方法，以确定药物重新定位为靶向NSC命运的抗抑郁药，并应用连接映射20。目标2将测试以下假设：目标1中确定的NSC衰老的全基因组转录组签名将映射由先前未确定其抗衰老潜在特性的已知药物诱导的相反mRNA变化。连接性作图是使用来自用生物活性小分子处理的培养的人类细胞的全基因组表达数据，通过共享基因表达变化的特征来发现药物、基因和疾病之间的功能关联21。学生将使用目标1中确定的衰老干细胞转录组学特征来探测NIH基于网络的细胞特征库（LINCS）22，以在20，143种已知药物中鉴定引起NSC中mRNA相反变化的化合物。目标3：验证已鉴定的老年神经保护剂将验证目标2中选定的抗衰老模拟药物，研究它们在挽救（i）细胞衰老表型方面的功效（增殖，干细胞，细胞死亡，DNA损伤，胶质细胞生成，神经发生和神经突生长）以及（ii）在我们的人海马祖细胞系的关键衰老分子途径。我们假设所选择的gerneuroproectors可以逆转细胞和分子衰老表型。目的4：产生一种新的体外加速NSC老化模型和交叉验证为了交叉验证目的3中验证的gerneuroproectors，将产生一种新的加速老化模型。我们将创建稳定的和四环素诱导的表达早老蛋白的NSC，以诱导年龄相关的表型，其特征在于增加的线粒体活性氧（mtROS）和增加的DNA双链断裂形成23。将交叉验证目标3中验证的老年神经保护剂减少或逆转增加的线粒体ROS产生和DNA损伤的潜力。

There is now a need and an opportunity to improve our understanding of the dynamics and molecular mechanisms of NSC ageing to exploit and control adult human endogenous NSC plasticity to maintain brain function during ageing.Proposed plan of work:Objective 1:Uncovering molecular signatures of NSC ageing We have already generated transcriptomic data from hippocampal progenitor cells exposed to young v. old serum. The student will use bioinformatic tools to generate molecular signatures of NSC ageing. Those pathways will be refined to be cognitively relevant with additional transcriptomic data currently being generated looking at old serum from low cognitive performers v. serum from cognitively high performers in adult hippocampal neurogenesis-dependant tasks.Objective 2:Identifying anti-neuroageing compounds (geroneuroproectors)This objective will initiate clinically feasible strategies to reverse ageing-induced changes identified in Objective 1. For that, drug repurposing is an efficient approach to by-pass lengthy drug development and safety trials. The Thuret lab recently successfully established the first strategy and method to identify drugs to reposition as antidepressants targeting NSC fate and applying connectivity mapping20.Objective 2 will test the hypothesisthat the genome-wide transcriptomic signature of NSC ageing identified in Objective 1 will map the opposite mRNA changes induced by known drugs not previously identified for their anti-ageing potential properties. Connectivity mapping is the use of genome-wide expression data from cultured human cells treated with bioactive small molecules to discover functional associations between drugs, genes and diseases through the feature of shared gene expression changes21. The student will use the ageing stem cell transcriptomic signature identified inObjective 1 to probe the NIH Library of Integrated Network-based Cellular Signatures (LINCS)22to identify compounds among the 20,143 known-drug that elicit opposite changes to mRNA in NSC.Objective 3:Validating the identifiedgeroneuroproectors Selected identified anti-ageing mimetic drugs in Objective 2 will be validated investigating their efficacy in rescuing (i) cellular ageing phenotypes (Proliferation, stemcellness, cell death, DNA damage, gliogenesis, neurogenesis and neurite outgrowth) as well as (ii) the key ageing molecular pathways in our human hippocampal progenitor cell line. We hypothesizethat the selected geroneuroproectorscan reverse cellular and molecular ageing phenotypes.Objective 4:Generating a novel in vitro accelerated model of NSC ageing & Cross-validation In order to cross-validate the validated geroneuroproectorsin Objective 3, a novel model of accelerated ageing will be generated. We will create stable and tetracycline inducible Progerin expressing NSCs to induce an age-related phenotype which is characterized by increased mitochondrial reactive oxygen species (mtROS) and increased formation of DNA double strand breaks 23. Validated geroneuroprotectors in Objective 3 will be cross-validated for their potential to reduce or reverse increased mtROS production and DNA Damage.