From birth to adulthood in a nutshell : hematopoietic stem cell development in vitro & HOXB4

Abstract

Ectopic expression of the homeodomain transcription factor HOXB4 has been shown to enhance the in vivo repopulation ability of in vitro differentiated mouse ES‐cells. How it promotes the conversion of ES‐cells to HSCs is not yet known. Here we demonstrate that its expression enforces the development of the developmentally earliest known hematopoietic cell during ES‐cell differentiation and promotes its expansion in stroma‐cell free suspension cultures. As these cultures lead to long‐term engraftment after transplantation into Rag2(‐/‐)γ(‐/‐) mice, the ability to engraft may correlate with the presence of these earliest, CD41hi cells. To test whether the CD41+ subpopulation depends on expression levels of HOXB4, we designed new tetracycline regulatable cassettes in which all necessary components were embedded in a single retroviral vector.The presence of a CD41hi / c‐Kit+/− / CD45− subpopulation correlated with HOXB4 expression levels in clonal ES‐HC cultures. This subpopulation was capable of reconstituting the entire heterogeneity of the primary suspension culture and differentiated towards all lineages in colony assays.In summary, we show that HOXB4 expression enhances the in vitro generation and expansion of the earliest known HSCs from mouse ES‐cells. Its tight and fine‐tuned regulation may also support the generation and safe expansion of patient‐specific pluripotent stem cell derived HSCs.