Abstract
Signals via fibroblast growth factor receptors (FGFRs) are involved in mesoderm induction events and may be also critical for early hematopoietic specification and proliferation of the hemangioblast. In vitro differentiated embryonic stem cells represent excellent system for the study of early hematopoietic commitment, particularly for understanding signals regulating the onset of hematopoietic differentiation. We have used human embryonic stem cells (hESCs) to study the expression of FGFR1, 2, 3, and 4 in undifferentiated cells and their differentiated progeny. Culturing hESCs i/ in high densities (protocol 1), ii/ without feeder layer of mouse embryonal fibroblasts and basic fibroblast growth factor (protocol 2), and iii/ in three-dimensional aggregates called embryoid bodies (protocol 3), was used to induce the differentiation. To achieve more directed and homogenous differentiation feeder-free hESCs were first subjected to the aggregation step (formation of embryoid bodies) that resembles the gastrulation process. This was followed by differentiation in monolayer in the presence of basic fibroblast growth factor (protocol 4). Such two-step differentiation protocol (5 + 10 days) was shown to activate ectodermal and mesodermal genes and form ectodermal and mesodermal cells (
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