NOTCH-mediated ex vivo expansion of human hematopoietic stem and progenitor cells by culture under hypoxia

Summary

Activation of NOTCH signaling in human hematopoietic stem/progenitor cells (HSPCs) by treatment with an engineered Delta-like ligand (DELTA1 ^ext-IgG [DXI]) has enabled ex vivo expansion of short-term HSPCs, but the effect on long-term repopulating hematopoietic stem cells (LTR-HSCs) remains uncertain. Here, we demonstrate that ex vivo culture of human adult HSPCs with DXI under low oxygen tension limits ER stress in LTR-HSCs and lineage-committed progenitors compared with normoxic cultures. A distinct HSC gene signature was upregulated in cells cultured with DXI in hypoxia and, after 21 days of culture, the frequency of LTR-HSCs increased 4.9-fold relative to uncultured cells and 4.2-fold compared with the normoxia + DXI group. NOTCH and hypoxia pathways intersected to maintain undifferentiated phenotypes in cultured HSPCs. Our work underscores the importance of mitigating ER stress perturbations to preserve functional LTR-HSCs in extended cultures and offers a clinically feasible platform for the expansion of human HSPCs.

Graphical abstract

Highlights

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  Superior NOTCH-mediated expansion of human HSPCs in hypoxic cultures

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  Culture under hypoxia mitigates ER stress in human HSPCs

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  Hypoxia potentiates NOTCH intracellular signaling in cultured human HSPCs

Abstract

Larochelle and colleagues concurrently activate NOTCH and hypoxic pathways in ex vivo cultures of human adult HSPCs to enable a clinically relevant expansion of cells with long-term repopulating potential after transplantation. They characterize the molecular intersection between the two signaling pathways and demonstrate the important role of low oxygen tension in mitigating ER stress during NOTCH-mediated expansion of human HSPCs.