HSC Diversity: Regulation by Clonal Selection vs Epigenetic Induction

描述(申请人提供)：造血干细胞(HSC)被定义为既能自我更新又能产生所有血细胞系的克隆性细胞。在过去的赠款期间，我们展示了随着小鼠和人类年龄的增长，血细胞谱系的表达发生了倾斜，从年轻成人的淋巴细胞生成和骨髓单核细胞生成之间的平衡，到老年人的骨髓单核细胞生成占主导地位。这种与衰老相关的造血分化结果的倾斜可能解释了晚年对髓系恶性肿瘤和贫血的易感性增加，以及对感染性疾病的适应性免疫能力降低。在这个建议中，我们考虑了两种可能的机制来解释这种与衰老相关的现象：(1)年轻生命中的所有HSC克隆型都产生了平衡的血细胞谱系，但随着时间的推移获得了一系列表观遗传学变化，这些变化沉默了一些基因并诱导了其他基因，导致了偏向于粒单核细胞生成的老年HSC；(2)在年轻人中同时存在“平衡的”和“偏向髓系的”HSC克隆型，随着时间的推移，环境、反馈和前馈调节外部刺激优先选择偏向髓系的HSC克隆型。虽然我们的初步数据支持后一种解释，但目前的建议将通过对不同HSC克隆类型的多样性和成熟度进行全面分析来最终评估这些替代方案。我们将初步确定不同类型的HSC克隆的数量，使用单细胞生物标记分析不同的基因表达模式，并评估从胚胎、胎儿、年轻成年和老年小鼠获得的HSC。我们将用敏感的克隆标记技术标记不同类型的单个HSC，然后研究它们在小鼠体内的起源、成熟、迁移和调节。我们将从成年人开始，追溯他们到胎儿或胚胎发育的后期阶段。我们将把它们追溯到衰老和白血病的形成。我们将测试当缺乏或输注过多时，终末血细胞类型[红细胞、血小板、粒细胞、单核细胞和巨噬细胞，各种淋巴细胞]是否将内源性HSC克隆带入或退出细胞周期，以扩大或减少某些HSC克隆型的数量。我们将评估与衰老相关的HSC分化结果的倾斜是否在人类中发生。这项关于HSC克隆多样性是否由出生预先决定，一生中的造血平衡是通过对不同生理事件的克隆选择来塑造的研究，是推动这项拨款基础研究的主题。

DESCRIPTION (provided by applicant): Hematopoietic stem cells (HSC) are defined as clonogenic cells that can both self-renew and give rise to all blood cell lineages. In the past grant period we demonstrated a skewing in blood cell lineage representation as mice and humans age, from a balance between lymphopoiesis and myelomonocytopoiesis in young adult life, to a predominance of myelomonocytopoiesis in the elderly. This aging-related skewing of hematopoietic differentiation outcome potentially explains the increased susceptibility late in life to myeloid malignancies and anemia, as well as diminished adaptive immunity to infectious disease. In this proposal, we consider two possible mechanisms for this aging-related phenomenon (1) that all HSC clonotypes in young adult life produce a balanced repertoire of blood cell lineages, but acquire a series of epigenetic changes over time which silence some genes and induce others, resulting in elderly HSC which are biased to myelomonocytopoiesis; or (2) that "balanced" and "myeloid-biased" HSC clonotypes both exist in young adults, with myeloid-biased HSC clonotypes being preferentially clonally selected over time by environmental, feed- back, and feed-forward regulatory external stimuli. While our preliminary data favor the latter explanation, the current proposal will definitively evaluate these alternatives by undertaking a comprehensive analysis of the diversity and maturation of different HSC clonotypes. We will initially determine the number of different kinds of HSC clones, using the single cell Biomark assay for diverse gene expression patterns, and evaluating HSCs obtained from embryonic, fetal, young adult, and aged mice. We will label individual HSCs of different types with sensitive clone marking techniques, then study their origin, maturation, migration, and regulation in mice. We will start in adults, tracing them back to the fetal or late embryonic stages of development. We will trace them forward to aging and to leukemogenesis. We will test whether the end blood cell types [RBC, platelets, granulocytes, monocytes and macrophages, the various lymphocytes] when lacking or transfused in overabundance bring endogenous HSC clones into or out of cell cycle to expand or diminish the number of certain HSC clonotypes. We will evaluate whether the same aging-related skewing of HSC differentiation outcomes occurs in humans. This examination of whether HSC clonal diversity is pre-determined by birth, with hematopoietic balance throughout life being shaped by clonal selection in response to diverse physiological events, is the theme that will drive the basic research of this grant.