A Long-term Follow-up Study on the Engraftment of Human Hematopoietic Stem Cells in Sheep

Abstract

Xenograft models of human hematopoiesis are essential to the study of the engraftment and proliferative potential of human hematopoietic stem cells (HSCs) in vivo. Immunodeficient mice and fetal sheep are often used as xenogeneic recipients because they are immunologically naive. In this study, we transplanted human HSCs into fetal sheep and assessed the long-term engraftment of transplanted human HSCs after birth. Fourteen sheep were used in this study. In 4 fetal sheep, HSCs were transduced with homeo-box B4 (HOXB4) gene before transplantation, which promoted the expansion of HSCs. Another 4 fetal sheep were subjected to non-myeloablative conditioning with busulfan. Seven of these 8 sheep showed successful engraftment of human HSCs (1-3% of colonyforming units) as assessed after the birth of fetal sheep (5 months post-transplantation), although HOXB4-transduced HSCs showed sustained engraftment for up to 40 months. Intact HSCs were transplanted into six non-conditioned fetal sheep, and human colony-forming units were not detected in the sheep after birth. These results suggest that, as compared with mouse models, where the short lifespan of mice limits long-term follow-up of HSC engraftment, the fetal sheep model provides a unique perspective for evaluating long-term engraftment and proliferation of human HSCs.

Fetal sheep support the development of hematopoietic cells in vivo from human induced pluripotent stem cells

“…In fact, we showed that human cord blood-derived HSPCs engrafted in sheep for long-term after transplantation into the fetal liver [21][22][23][24]. Here, we demonstrate that a sheep fetal liver provides a microenvironment for turning human iPSCderived HEs to hematopoietic cells with the capability of long-term engraftment and multilineage differentiation (Figure 1).…”

“…Although the engraftment of human iPSC-derived hematopoietic cells was successful, human cells were only detected in the bone marrow of sheep but not in the peripheral blood by sensitive PCR (Figure 4B). It was also the case when human cord blood CD34+ cells were transplanted into sheep fetuses [21][22][23][24]. Actually, it was observed in immunodeficient NSG mice transplanted with human HSPCs [44,45].…”

“…The in utero transplantation procedures were performed as described previously [19][20][21][22][23][24]50]. Briefly, at 6 days before transplantation, the pregnant ewes wer e injected intravenously with busulfan (Wako Pure Chemical Industries, Osa ka, Japan) at 3 mg/kg body weight to treat the fetuses [22].…”

“…To evaluate the engraftment of human hematopoietic cells in sheep, the presence of human cells and human colony-forming units (CFUs) in the peripheral blood or bone marrow were determined by nested-PCR as previously described [19,[21][22][23].…”