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undifferentiated hESCs in mixed cell cultures as well as when transplanted
into animal models. These marked cells can be analyzed not only in tissues
or organs ex vivo but also possibly in live animals by noninvasive whole-
body imaging. 83
In the following sections, we concentrate on the directed differentiation of
hESCs to the hematopoietic, cardiac, neural, and pancreatic lineages as these
are the areas in which most progress has been achieved. We also highlight
key findings following the transplantation of these hESC derivatives into
animal models.
1.4 Differentiation of hESCs to the Hematopoietic Lineage
The ability to direct the differentiation of hESCs in vitro to hematopoietic
lineages is aiding our understanding of blood cell development. Many stud-
ies have examined the development of hematopoietic and endothelial pro-
genitors in hESC cultures; however, the ability to develop transplantable
hematopoietic stem cells (HSCs) is still elusive. The role of particular cytok-
ines and growth factors on hematopoietic development from hESCs has
begun to be investigated, but many of these early studies were conducted
in the presence of fetal calf serum. 84 BMP4, a ventral mesoderm inducer, has
been shown to strongly promote hematopoietic differentiation from hESCs
being cultured as EBs, when combined with other hematopoietic cytok-
ines. 85 The addition of vascular endothelial growth factor A (VEGF-A) fur-
ther increased the efficiency of erythroid colony formation. 86 A modification
of the EB system allowing for the formation of uniform-size “spin EBs” in a
completely defined media free of animal product further defined the role of
BMP4 for the development of hematopoietic mesoderm but less frequently
supported the generation of hematopoietic progenitors. 51,52 However, the
combination of BMP4 with VEGF, stem cell factor (SCF), and FGF2 enhanced
the total yield of hematopoietic precursors. 52
Wnt signaling pathways also appear to play roles in hematopoiesis. Woll
et al. demonstrated that the inhibition of canonical Wnt signals in serum-
induced cultures resulted in a decrease in the proportion of cells with
hematopoietic potential. 87 This was further supported by the demonstra-
tion that a noncanonical Wnt (Wnt11) can direct hESCs toward mesoderm,
whereas canonical Wnt3a causes cells already committed to the hematopoi-
etic lineage to proliferate. 88
The kinetics of hematopoietic lineage development within hESC differentia-
tion parallels that observed with mESCs. Differentiating hESCs pass through
a primitive streak stage defined by the expression of either BRACHYURY or
MIXL1, markers of the primitive streak, to a KDR + or PDGFRα + mesodermal
progenitor, and subsequently to the yolk sac hematopoietic program. 51,53,54,89
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