Ex Vivo Expansion Potential of Murine Hematopoietic Stem Cells: A Rare Property Only Partially Predicted by Phenotype

The scarcity of hematopoietic stem cells (HSCs) restricts their use in both clinical settings and experimental research. Here, we examined a recently developed method for expanding rigorously purified murine HSCs ex vivo. Input HSCs displayed varying potential for ex vivo self-renewal, with alternative outcomes revealed by single cell multimodal RNA- and ATAC-seq profiling. While most HSC progeny offered only transient in vivo reconstitution, these cells efficiently rescued mice from lethal myeloablation. The amplification of functional HSC activity allowed for long-term multilineage engraftment in unconditioned hosts that somewhat surprisingly associated with a return of HSCs to quiescence. Thereby, our findings identify several key considerations for ex vivo HSC expansion, with major implications also for assessment of normal HSC activity. Overall design: To gain a deeper understanding of the cells generated in ex vivo HSC cultures, we collected cells from the entire culture or EPCR+ cells and isolated nuclei for single cell multiome sequencing