Wellcome MRC Cambridge Stem Cell Institute, University of Cambridge, Cambridge, UK.
Department of Haematology, University of Cambridge, Cambridge, UK.
EMBO Rep. 2022 Oct 6;23(10):e55502. doi: 10.15252/embr.202255502. Epub 2022 Aug 16.
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 > 200-fold expansion of functional HSCs, but their molecular characterization has not been possible since the majority of cells are non-HSCs and single cell-initiated cultures have 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. By directly linking single-clone functional transplantation data with single-clone gene expression profiling, we show that the molecular profile of expanded HSCs is similar to proliferating fetal HSCs and reveals a gene expression signature, including Esam, Prdm16, Fstl1, and Palld, that can identify functional HSCs from multiple cellular states. This "repopulation signature" (RepopSig) also enriches for HSCs in human datasets. Together, these findings demonstrate the power of integrating functional and molecular datasets to better derive meaningful gene signatures and opens the opportunity for a wide range of functional screening and molecular experiments previously not possible due to limited HSC numbers.
造血干细胞（HSCs）在体外培养是广泛的细胞和基因治疗的基本组成部分。最近的研究已经实现了功能 HSCs 的>200 倍扩增，但由于大多数细胞是非 HSCs，并且单细胞起始培养具有显著的克隆间变异性，因此其分子特征尚未可知。我们使用 Fgd5 报告小鼠与 EPCR 表面标记物相结合，从扩增培养物中的非 HSCs 中报告了 HSCs 的独特鉴定。通过将单细胞功能移植数据与单细胞基因表达谱直接关联，我们表明扩增的 HSCs 的分子特征与增殖的胎儿 HSCs 相似，并揭示了一个基因表达特征，包括 Esam、Prdm16、Fstl1 和 Palld，该特征可从多种细胞状态中识别功能 HSCs。这个“再群体签名”（RepopSig）也可在人类数据集富集 HSCs。总之，这些发现证明了整合功能和分子数据集以更好地得出有意义的基因特征的强大功能，并为以前由于 HSCs 数量有限而无法进行的广泛功能筛选和分子实验开辟了机会。
