Efficient Hematopoietic Differentiation of Human Embryonic Stem Cells on Stromal Cells Derived from Hematopoietic Niches

Abstract

Hematopoietic stem cells derived from human embryonic stem cells (hESCs) could provide a therapeutic alternative to bone marrow transplants, but the efficiency of currently available derivation protocols is low. In this study, we investigated whether coculture with monolayers of cells derived from mouse AGM and fetal liver, or with stromal cell lines derived from these tissues, can enhance hESC hematopoietic differentiation. We found that under such conditions hESC-derived differentiating cells formed early hematopoietic progenitors, with a peak at day 18-21 of differentiation that corresponded to the highest CD34 expression. These hESC-derived hematopoietic cells were capable of primary and secondary hematopoietic engraftment into immunocompromised mice at substantially higher levels than described previously. Transcriptional and functional analysis identified TGF-beta1 and TGF-beta3 as positive enhancers of hESC hematopoietic differentiation that can further stimulate this process when added to the culture. Overall, our findings represent significant progress toward the goal of deriving functional hematopoietic stem cells from hESCs.

Generation of T Cells from Human Embryonic Stem Cell-Derived Hematopoietic Zones

“…During the course of these experiments we noted the formation of endothelium-lined cell clumps, reminiscent of blood islands during embryonic development, which we termed hematopoietic zones (HZs). These structures have since been described by two other groups and were shown to contain HPC with self renewing capacity and HPC capable of generating massive amounts of megakaryocytes (5,11). In this study, we report that these HZs generate large numbers of T cells when transferred on OP9-DL1 cells in the presence of growth factors.…”

“…However, as discussed by the authors, expression of adult globin in red lineage cells was absent, and definitive proof of lymphoid engraftment was lacking (5). Therefore, to the best of our knowledge, formal proof of the existence of hESC-derived HSC is still lacking.…”

“…The three-dimensional embryoid body system and coculture systems on stromal cell lines such as murine OP9 cells or aorta-gonad-mesonephros-derived stromal cells are often used (2,3). In both systems, multipotent HPC with self-renewing activity were demonstrated (4,5). Vodyanik et al (6) performed an extensive analysis of precursors generated from hESC in OP9 coculture and described a CD34 ϩ CD43 ϩ population, with precursor activity for B, NK, and myeloid cells.…”

A short history of hemogenic endothelium

“…This is reflected in the difficulty to generate bona fide HSCs de novo from ES/iPS cells [50,85,[91][92][93], or through direct reprogramming [94,95]. This indicates that critical components of EHT, responsible for regulating the hematopoietic nature of these cells are missing in these culture systems and indicate the importance of understanding the cellular, molecular and niche requirements for EHT in vivo.…”

Migration of dorsal aorta mesenchymal stem cells induced by mouse embryonic circulation

“…The emergence and maintenance of HSCs in the DA depend upon their interaction with the local microenvironment, including the mesenchymal and endothelial cells (Ohneda et al, 1998;Xu et al, 1998;Oostendorp et al, 2002a,b;Durand et al, 2007;Ledran et al, 2008). Regarding stromal cells from the AGM region, the hematopoietic supportive potential other than the mesenchymal capacity has been intensely investigated.…”