Expanding and defining human hematopoietic stem and progenitor cells ex vivo using small molecules
Human hematopoietic stem cells (HSCs) are defined by their capacity to self-renew and to differentiate into all blood lineages during an adult lifetime. Based on these unique properties, HSCs are used in transplantation procedures to treat various hematological diseases. However, the low number of HSCs in a graft limits the use of this treatment. To overcome this restrain, different approaches were established to expand HSCs ex vivo; yet, the absence of a reliable surface maker that correlates with HSC activity in culture made the assessment labor-intensive and time-consuming. Using a library of small molecules, we were able to identify pyrimidoindole derivative named UM171 as an agonist for HSC self-renewal. UM171 promotes ex vivo expansion of hematopoietic and stem cell progenitors (HSPC) independently of AhR suppression- a pathway reported by Boitano et al. to have the greatest effect in HSC expansion. Unlike AhR suppression that targets a hematopoietic population with limited proliferative potential, UM171 targets the long-term HSCs. Transcriptome analysis showed that UM171 reduces the levels of transcripts associated with lineage differentiation and induces the expression of genes encoding for membrane proteins, one of the best differentially expressed being the endothelial protein c receptor (EPCR). Cell sorting and transplantation experiments of EPCR expressing cells showed a high correlation with HSC activity. We demonstrated EPCR as a first reliable marker to enrich for HSC in culture and that it is required for HSPC function in vivo. These findings provide a valuable tool for clinical and research applications to optimize further HSPC expansion protocols and understand the molecular machinery that governs the HSC self-renewal.
Cord Blood Hematopoietic Stem Cell Self-renewal Small Molecule Surface Marker