CD43 but Not CD41 Marks the First Hematopoietic Stem Cells in the Human Embryo

Hematopoietic stem cells (HSCs) emerge during embryogenesis and maintain multilineage hematopoiesis throughout the entire lifespan of the organism. Due to limited access to human embryonic tissues, early human HSC development remains a largely unexplored area of research. A better understanding of this process may be instrumental in creating novel protocols for in vitro generation of human HSCs from pluripotent stem cells to overcome the shortage of donor cells and increase the safety of HSC transplantations in the clinic.

We have recently described the spatio-temporal distribution of HSCs in the early human embryo and established the immunophenotype of the first human HSCs (Ivanovs et al., J. Exp. Med., 2011; Ivanovs et al., Stem Cell Reports, 2014). The first human HSCs emerge in the ventral domain of the dorsal aorta, possess high regenerative potential, and reside within the CD34⁺VE-cadherin⁺CD45⁺c-Kit⁺Thy-1⁺Endoglin⁺Runx1⁺CD38^-/loCD45RA^- cell population, which harbors all HSCs and committed hematopoietic progenitors in the human aorta-gonad-mesonephros (AGM) region. Here, we aimed to find a surface marker which would further enrich for the human AGM region HSCs. We found that CD41, which is the earliest surface marker of hematopoietic commitment in the mouse embryo (Ferkowicz et al., Development, 2003; Mikkola et al., Blood, 2003), does not mark the first human HSCs (3 independent experiments). Instead, the first HSCs in the human AGM region express CD43 (4 independent experiments). Of note, it has been shown previously that during human embryonic stem cell differentiation in vitro, CD43 but not CD41 marks multipotent hematopoietic progenitors (Vodyanik et al., Blood, 2006).

Human embryonic tissues for our study were obtained after elective medical termination of pregnancy with informed written consent according to the Declaration of Helsinki. The study protocol was approved by the Lothian Research Ethics Committee. Employing magnetic cell separation and xenotransplantations, CD41- and CD43-positive and -negative cell populations were isolated from Carnegie stage 15–17 human AGM regions and individually transplanted into preconditioned adult NOD.Cg-Prkdc^scidIl2rg^tm1Wjl/Sz mice. Human hematopoietic contribution to the peripheral blood, bone marrow, spleen, and thymus of transplanted mice was assessed at 4 months after transplantation.

Our findings suggest that, although the process of HSC development is largely conserved between the mouse and the human, certain aspects vary substantially between the species, reflecting possible differences in the HSC specification program. We conclude that CD43 is a useful surface marker for the purification of the first human HSCs to enable further studies into molecular mechanisms underlying their development and high regenerative potential.