Human embryonic stem cells maintained in the absence of mouse embryonic fibroblasts or conditioned media are capable of hematopoietic development.
To date, hematopoietic development of human embryonic stem cells (hESCs) has been limited to cell lines cultured in the presence of either mouse embryonic fibroblasts (MEFs) or MEF-conditioned media (MEF-CM). Anonymous xenogenic factors from MEFs or MEF-CM complicate studies of hESC self-renewal and also raise concerns for the potential clinical applications of generating primitive hematopoietic cells from hESC lines maintained under these ambiguous conditions. Here, we demonstrate that hESCs can be cultured over 30 passages in defined conditions in the absence of MEFs or MEF-CM using only serum replacement (SR) media and high concentrations of basic fibroblast growth factor (SR-bFGF). Similar to hESCs cultured in MEF-CM, hESCs cultured in SR-bFGF sustained characteristics of undifferentiated hESCs, proliferative potential, normal karyotype, in vitro and in vivo 3 germ-layer specification and gave rise to hemogenic-endothelial precursors required for subsequent primitive hematopoietic development. Our report demonstrates that anonymous factors produced by feeder cells are not necessary for hESC maintenance and subsequent hematopoietic specification, thereby providing a defined system for studies of hESC self-renewal and hESC-derived hematopoiesis.
Wang
Lisheng
L
Krembil Center for Stem Cell Biology and Regenerative Medicine, Robarts Research Institute, 100 Perth Dr, London, ON N6A 5K8, Canada.
Li
Li
L
Menendez
Pablo
P
Cerdan
Chantal
C
Bhatia
Mickie
M
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000494:pharmacology*; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D004622:Embryo, Mammalian / Q000166:cytology*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005434:Flow Cytometry; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D007621:Karyotyping; D017493:Leukocyte Common Antigens / Q000096:biosynthesis; D051379:Mice; D016513:Mice, SCID; D050814:Octamer Transcription Factor-3; D016133:Polymerase Chain Reaction; D013234:Stem Cells; D013724:Teratoma; D013997:Time Factors; D014157:Transcription Factors / Q000096:biosynthesis
D017077:Culture Media, Conditioned; D004268:DNA-Binding Proteins; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; C494174:Pou5f1 protein, mouse; D014157:Transcription Factors; D016222:Fibroblast Growth Factor 2; D000469:Alkaline Phosphatase; D017493:Leukocyte Common Antigens