Visualization and Lineage Tracing of Hematopoietic Stem Cell Heterogeneity

描述（由申请人提供）：骨髓中的造血干细胞（HSC）在整个生命过程中不断产生所有类型的血细胞。老年人HSC的活性表现出异常，如淋巴发育减少、骨髓发育增强和易患骨髓增生性疾病。最近的证据表明，HSC包括两个不同的子集：“淋巴偏向”HSC有限的自我更新能力，和“骨髓偏向”HSC具有优越的上级长期重建能力。后一种亚型随着年龄的增长而占优势，限制淋巴细胞生成并易发生骨髓增生。虽然“HSC异质性”模型为理解HSC动力学和衰老提供了必要的框架，但它受到我们目前无法前瞻性地识别两个HSC亚群或在完整生物体中追踪其后代的限制。我们的初步研究的特点是转基因株表达绿色荧光蛋白或Cre重组酶的一部分HSC一致的“骨髓偏向”的HSC。我们建议，这些报告菌株可用于鉴定和谱系追踪的两个HSC子集。在目标1中，表达分析和过继转移将用于验证荧光和Cre报告基因对功能性HSC亚群的标记。在目的2中，这些报告基因将用于动员和免疫介导的应激后HSC的定量和谱系追踪。此外，造血干细胞亚群的发展髓性白血病的特点。这些实验将建立一个新的遗传系统来分析HSC异质性，并在没有过继转移的完整动物中表征HSC动力学。成功的结果将为白血病发生过程中HSC异质性的研究提供有用的工具和初步数据。了解HSC活性的细胞基础，应促进新的方法来“振兴”的年龄或压力受损的造血和造血系统恶性肿瘤的治疗。

DESCRIPTION (provided by applicant): Hematopoietic stem cells (HSC) in the bone marrow continuously give rise to all blood cell types throughout the entire life. The activity of HSC in aged individuals shows abnormalities such as reduced lymphoid development, enhanced myeloid development, and predisposition to myeloproliferative diseases. Recent evidence suggests that the HSC comprise two distinct subsets: the "lymphoid-biased" HSC with limited self-renewal capacity, and the "myeloid-biased" HSC with superior long-term reconstitution capacity. The latter subset becomes predominant with age, limiting lymphopoiesis and predisposing to myeloproliferation. While the "HSC heterogeneity" model provides essential framework for understanding HSC dynamics and aging, it is limited by our current inability to prospectively identify the two HSC subsets or to trace their progeny in intact organisms. Our preliminary studies characterized transgenic strains expressing green fluorescent protein or Cre recombinase in a fraction of HSC consistent with "myeloid-biased" HSC. We propose that these reporter strains can be used for the identification and lineage tracing of the two HSC subsets. In Aim 1, expression analysis and adoptive transfers will be used to verify the marking of functional HSC subsets by the fluorescent and Cre reporters. In Aim 2, these reporters will be used for the quantification and lineage tracing of the HSC after mobilization and immune-mediated stress. In addition, the contribution of HSC subsets to the development of myeloid leukemia will be characterized. These experiments would establish a novel genetic system to analyze HSC heterogeneity, and characterize HSC dynamics in intact animals without adoptive transfer. A successful outcome would provide useful tools and initial data for the study of HSC heterogeneity during leukemogenesis. Understanding the cellular basis of HSC activity should facilitate novel approaches to the "rejuvenation" of age- or stress-impaired hematopoiesis and to the treatment of hematopoietic malignancies.