Long-term ex vivo haematopoietic-stem-cell expansion allows nonconditioned transplantation
Long-term ex vivo haematopoietic-stem-cell expansion allows nonconditioned transplantation
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DOI：
10.1038/s41586-019-1244-x
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发表时间：
2019-07-04
期刊：
影响因子：
64.8
通讯作者：
Yamazaki, Satoshi
中科院分区：
文献类型：
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作者：
Wilkinson, Adam C.;Ishida, Reiko;Yamazaki, Satoshi
多能自我更新造血干细胞 (HSC) 移植后可再生成人血液系统 (1)，这是包括免疫缺陷和白血病 (2) 在内的多种疾病的治疗方法。尽管已付出大量努力通过体内骨髓 HSC 微环境或生态位的表征来识别 HSC 维持因子 (3-5)，但以前无法实现稳定的离体 HSC 扩增 (6,7)。在这里，我们描述了一种明确的、无白蛋白的培养系统的开发，该系统支持功能性小鼠 HSC 的长期离体扩增。我们采用系统优化方法，发现高水平的血小板生成素与低水平的干细胞因子和纤连蛋白协同作用，维持 HSC 的自我更新。血清白蛋白长期以来被认为是 HSC 培养物中生物污染物的主要来源 (8)；我们认为聚乙烯醇是功能优越的血清白蛋白替代品，符合良好的生产规范。这些条件使功能性 HSC 在 1 个月内扩增 236 至 899 倍，尽管对克隆衍生培养物的分析表明 HSC 离体自我更新能力存在相当大的异质性。使用该系统，仅源自 50 个细胞的 HSC 培养物可以牢固地植入受体小鼠体内，无需正常的毒性预处理（例如辐射），这可能与人类 HSC 移植相关。因此，这些发现对 HSC 基础研究和临床血液学具有重要意义。
Multipotent self-renewing haematopoietic stem cells (HSCs) regenerate the adult blood system after transplantation(1), which is a curative therapy for numerous diseases including immunodeficiencies and leukaemias(2). Although substantial effort has been applied to identifying HSC maintenance factors through the characterization of the in vivo bone-marrow HSC microenvironment or niche(3-5), stable ex vivo HSC expansion has previously been unattainable(6,7). Here we describe the development of a defined, albumin-free culture system that supports the long-term ex vivo expansion of functional mouse HSCs. We used a systematic optimization approach, and found that high levels of thrombopoietin synergize with low levels of stem-cell factor and fibronectin to sustain HSC self-renewal. Serum albumin has long been recognized as a major source of biological contaminants in HSC cultures(8); we identify polyvinyl alcohol as a functionally superior replacement for serum albumin that is compatible with good manufacturing practice. These conditions afford between 236- and 899-fold expansions of functional HSCs over 1 month, although analysis of clonally derived cultures suggests that there is considerable heterogeneity in the self-renewal capacity of HSCs ex vivo. Using this system, HSC cultures that are derived from only 50 cells robustly engraft in recipient mice without the normal requirement for toxic pre-conditioning (for example, radiation), which may be relevant for HSC transplantation in humans. These findings therefore have important implications for both basic HSC research and clinical haematology.