A highly efficient reporter system for identifying and characterizing in vitro expanded hematopoietic stem cells
A highly efficient reporter system for identifying and characterizing in vitro expanded hematopoietic stem cells
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DOI：
10.1101/2021.06.18.448972
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发表时间：
2021-06
期刊：
影响因子：
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通讯作者：
J. Che;D. Bode;I. Kucinski;A. Cull;Fiona Bain;Melania Barile;Grace Boyd;M. Belmonte;M. Jassinskaj
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文献类型：
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作者：
J. Che;D. Bode;I. Kucinski;A. Cull;Fiona Bain;Melania Barile;Grace Boyd;M. Belmonte;M. Jassinskaj
体外培养的造血干细胞（HSC）是广泛的细胞和基因疗法的基本组成部分。最近的努力已经实现了功能性HSC的超过200倍的扩增，但是由于绝大多数细胞是非HSC并且单细胞起始的培养物显示出显著的克隆间变异性，因此它们的分子表征是不可能的。使用Fgd5报告小鼠与EPCR表面标记物组合，我们报告了扩增培养物中非HSC的HSC的唯一鉴定。将单克隆功能移植数据与单克隆基因表达谱联系起来，我们发现扩增的HSC的分子谱与活跃循环的胎肝HSC相似，并且与所有来源的功能性HSC（包括Prdm 16、Fstl1和Palld）共享基因表达特征。这种新工具现在可以应用于广泛的功能筛选和分子实验，以前由于有限的HSC数量而不可能。
Hematopoietic stem cells (HSCs) cultured outside the body are the fundamental component of a wide range of cellular and gene therapies. Recent efforts have achieved more than 200-fold expansion of functional HSCs, but their molecular characterization has not been possible due to the substantial majority of cells being non-HSCs and single cell-initiated cultures displaying substantial clone-to-clone variability. Using the Fgd5 reporter mouse in combination with the EPCR surface marker, we report exclusive identification of HSCs from non-HSCs in expansion cultures. Linking single clone functional transplantation data with single clone gene expression profiling, we show that the molecular profile of expanded HSCs is similar to actively cycling fetal liver HSCs and shares a gene expression signature with functional HSCs from all sources, including Prdm16, Fstl1 and Palld. This new tool can now be applied to a wide-range of functional screening and molecular experiments previously not possible due to limited HSC numbers.