BLRD Research Career Scientist Award Application

项目概要/摘要 弗里曼博士的研究的首要目标是了解基因的改变是如何改变的。 基因组/表观基因组对衰老的贡献，以及治疗如何可能针对这些变化来维持 “年轻”的表观基因组，延缓衰老，延迟/预防与年龄有关的疾病，并改善健康和 老年人的生活质量。表观遗传失调是衰老的标志，但这究竟是如何导致 细胞功能障碍和疾病是未知的。为了阐明这一贡献，弗里曼博士的VA研究已经 集中于随着年龄的增长，星形胶质细胞、小胶质细胞和海马神经元的表观基因调控。使用 新的细胞分离和先进的测序方法，细胞类型特异性碱基分辨率图， 在CG和非CG背景下，产生了甲基化（mC）和羟甲基化（hmC）。与配对 RNA表达数据、生物信息学技术正被用来了解基因表达的调控 随着年龄的增长。这种类型的碱基特异性、全基因组和细胞类型特异性研究尚未在 老化研究，并可能提供最深入的看法与老化的表观基因组变化的日期。 此外，为了证明预防年龄相关的表观基因组变化，正在使用热量限制。在 总的来说，这些研究不仅将增加对衰老的神经表观基因组学的理解，而且还将提供 未来使用表观基因组编辑进行干预研究的目标。重要的是，这些研究检查了 和女性提供性别知情的见解，因为他以前的研究表明，大多数年龄- 相关的表观基因组变化是性别差异的。 博士弗里曼还是三个NIH项目和一个基金会资助项目的PI或共同PI。这些研究扩展了 他通过使用新型转基因小鼠模型和大脑抗衰老干预对衰老进行了VA研究， 视网膜和GI系统。他的实验室开发的这些模型允许时间控制，细胞类型特异性标记 以分离用于表观基因组分析的DNA和用于表达分析的RNA， 需要细胞分选。在脑和视网膜中，异时血浆转移，其中来自年轻小鼠的血浆是 给老年小鼠进行返老还童，正在被用来确定是否可以与年龄相关的表观基因组变化。 发生后，就被逆转了。同时，使用蛋白质组学方法，DNMT和泰特相互作用的蛋白质 将它们靶向特定的基因组区域进行表观基因组改变。 更广泛地说，Freeman博士还与其他OKC VAMC研究人员进行了广泛的合作。在 在这些VA和NIH资助的研究中，弗里曼博士正在研究肌肉衰老的细胞类型特异性机制， 大脑卵巢关节和结肠除了为这些研究带来分子和生物化学专业知识外， 衰老的表观遗传学的共同线索允许在器官系统之间进行比较和对比。 这些合作与他作为最近更新的基因组科学核心主任的工作相结合。 P30 NIA中心赠款：俄克拉荷马州内森休克衰老中心。该核心提供了表观基因组和 转录组学分析以及线粒体基因组分析，为全国各地的老龄化研究人员。 此外，弗里曼博士领导了许多跨机构（OMRF，OUHSC）的举措，重点是学员和教师 发展，包括培训学生和博士后（F部队）和初级教师（NeuroGrant Seekers）， 成功准备和提交赠款。他还与OKC VAMC的领导合作， 招募新的，非常成功的调查员到VA系统，其中一个已经获得了MERIT奖， 目前正在审查另外三名调查员的MERIT申请。

Project Summary/Abstract The overarching goal of Dr. Freeman’s research has been to understand how alterations of the genome/epigenome contribute to aging and how therapies could potentially target these changes to maintain a ‘youthful’ epigenome that retards aging, delays/prevents age-related diseases, and improves the health and quality of life of the elderly. Epigenetic dysregulation is a hallmark of aging, but exactly how this contributes to cellular dysfunction and disease is not known. To elucidate this contribution, Dr. Freeman’s VA research has focused on epigenomic regulation in astrocytes, microglia, and neurons in the hippocampus with aging. Using novel cell isolation and advanced sequencing approaches, cell-type specific base resolution maps of both methylation (mC) and hydroxymethylation (hmC) in CG and non-CG contexts are being generated. With paired RNA expression data, bioinformatic techniques are being used to understand the regulation of gene expression with aging. This type of base-specific, genome-wide, and cell-type specific studies have not been performed in aging research and will potentially offer the most in-depth view of epigenomic changes with aging to date. Further, to demonstrate the prevention of age-related epigenomic changes, caloric restriction is being used. In total, these studies will not only increase the understanding of the neuroepigenomics of aging but also provide targets for future interventional studies using epigenome editing. Importantly these studies examine both males and females to provide sex-informed insight as his prior studies have demonstrated that the majority of age- related epigenomic changes are sexually divergent. Dr. Freeman is also PI or co-PI on three NIH projects and one foundation funded project. These studies extend his VA research in aging through use of novel transgenic mouse models and anti-aging interventions in the brain, retina, and GI system. These models developed by his lab allow temporally controlled, cell-type specific tagging of nuclei and ribosomes to isolate DNA for epigenomic analysis and RNA for expression analysis without the need for cell sorting. In the brain and retina, heterochronic plasma transfer, in which plasma from young mice is administered to old mice for rejuvenation, is being used to determine if age-related epigenomic changes can be reversed after they have occurred. In parallel, using proteomic approaches, DNMT and TET interacting proteins are being identified that target them to specific genomic regions for epigenomic alteration. More broadly, Dr. Freeman also has an extensive collaborative portfolio with other OKC VAMC investigators. In these VA and NIH funded studies, Dr. Freeman is examining cell-type specific mechanisms of aging in muscle, brain, ovary, joints, and colon. In addition to bringing molecular and biochemical expertise to these studies, a common thread of epigenetics of aging allows comparisons and contrasts to be made across the organ systems. These collaborations are paired with his work as Director of the Genomics Sciences Core of a recently renewed P30 NIA Center grant: the Oklahoma Nathan Shock Aging Center. This core provides epigenomic and transcriptomic analyses as well as mitochondrial genomic assays for aging researchers across the country. Additionally, Dr. Freeman leads many cross-institution (OMRF, OUHSC) initiatives focused on trainee and faculty development, including training students and post-docs (F-Troop) and junior faculty (NeuroGrant Seekers) in successful preparation and submission of grants. He has also worked with leaders at the OKC VAMC to help recruit new, highly successful investigators to the VA system, one of which has received a MERIT award with three more investigators having MERIT applications currently under review.