HES-1 preserves purified hematopoietic stem cells ex vivo and accumulates side population cells in vivo

Mouse long-term hematopoietic reconstituting cells exist in the c-Kit+Sca-1+Lin− (KSL) cell population; among them, CD34low/− cells represent the most highly purified population of hematopoietic stem cells in the adult bone marrow. Here, we demonstrate that retrovirus-mediated transduction of CD34low/−c-Kit+Sca-1+Lin−(34−KSL) cells with the HES-1 gene, which encodes a basic helix-loop-helix transcription factor functioning downstream of the Notch receptor, and is a key molecule for the growth phase of neural stem cells in the embryo, preserves the long-term reconstituting activity of these cells in vitro. We also show that cells derived from the HES-1–transduced 34−KSL population produce progenies characterized by negative Hoechst dye staining, which defines the side population, and by CD34low/− profile in the bone marrow KSL population in each recipient mouse at ratios 3.5- and 7.8-fold those produced by nontransduced 34−KSL-derived competitor cells. We conclude that HES-1 preserves the long-term reconstituting hematopoietic activity of 34−KSL stem cells ex vivo. Up-regulation of HES-1 protein in the 34−KSL population before unnecessary cell division, that is, without retrovirus transduction, may represent a potent approach to absolute expansion of hematopoietic stem cells.