Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells

Human embryonic stem (ES) cells are pluripotent cells derived from the inne r cell mass of in vitro fertilized human blastocysts. We examined the poten tial of eight growth factors [basic fibroblast growth factor (bFGF). transf orming growth factor beta 1 (TGF-beta 1). activin-A, bone morphogenic prote in 4 (BMP-4), hepatocyte growth factor (HGF), epidermal growth factor (EGF) . beta nerve growth factor (beta NGF). and retinoic acid] to direct the dif ferentiation of human ES-derived cells in vitro. We show that human ES cell s that have initiated development as aggregates (embryoid bodies) express a receptor for each of these factors, and that their effects are evident by differentiation into cells with different epithelial or mesenchymal morphol ogies. Differentiation of the cells was assayed by expression of 24 cell-sp ecific molecular markers that cover all embryonic germ layers and 11 differ ent tissues. Each growth factor has a unique effect that may result from di rected differentiation and/or cell selection, and we can divide the overall effects of the factors into three categories: growth factors (Activin-A an d TGF beta 1) that mainly induce mesodermal cells; factors (retinoic acid, EGF. BMP-4, and bFGF) that activate ectodermal and mesodermal markers; and factors (NGF and HGF) that allow differentiation into the three embryonic g erm layers, including endoderm. None of the growth factors directs differen tiation exclusively to one cell type. This analysis sets the stage for dire cting differentiation of human ES cells in culture and indicates that multi ple human cell types may be enriched in vitro by specific factors.