Hypoxic culture maintains self-renewal and enhances embryoid body formation of human embryonic stem cells.
Hypoxic environment is theoretically more physiological for the growth of human embryonic stem (hES) cells. It has been reported that hypoxic culture maintained better undifferentiation of hES cells, but the effects on differentiation are less well established. The hES cells were thus cultured and compared in hypoxia (2% oxygen [O2]) and normoxia (21% O2). The data showed that the undifferentiated state of hES cells was maintained more favorably in hypoxia during prolonged culture. Most tested genes belonging to FGF, TGF-beta/GMP, and Wnt signaling pathways were enriched in undifferentiated hES cells and downregulated upon differentiation, accompanied with differential expression of FGFR1, FGFR2, and FRAT2 between hypoxia and normoxia. Higher P-Smad2/3 level was identified in hypoxia, favoring the maintenance of hES cells in undifferentiation. Bisulfite sequencing showed similar imprinting status between different O2 tensions at H19 differentially methylated region (DMR) and KvDMR loci. Embryoid body formation was enhanced in hypoxia accompanied with suppressed Sox17, Desmin, Gata4, Brachyury, and Cdx2 expression. We concluded that hypoxia improved self-renewal of hES cells through modulation of major signaling pathways and was also more efficient for differentiation to embryoid bodies, though they might present with suppressed expression of some lineage-specific genes across all the three embryonic germ layers and trophectoderm.
Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, College of Medicine and the Hospital, National Taiwan University, Taipei, Taiwan.
Hsin-Fu
HF
Chen
Hung-Chih
HC
Kuo
Shau-Ping
SP
Lin
Chung-Liang
CL
Chien
Ming-Shan
MS
Chiang
Hong-Nerng
HN
Ho
D015153:Blotting, Western; D000071616:CDX2 Transcription Factor; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D015687:Cell Hypoxia / Q000502:physiology*; D002460:Cell Line; D003893:Desmin / Q000235:genetics / Q000378:metabolism; D058732:Embryoid Bodies / Q000166:cytology* / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D005326:Fetal Proteins / Q000235:genetics / Q000378:metabolism; D005346:Fibroblast Growth Factors / Q000235:genetics / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D050991:GATA4 Transcription Factor / Q000235:genetics / Q000378:metabolism; D018398:Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D051496:Receptor, Fibroblast Growth Factor, Type 1 / Q000235:genetics / Q000378:metabolism; D051497:Receptor, Fibroblast Growth Factor, Type 2 / Q000235:genetics / Q000378:metabolism; D055760:SOXF Transcription Factors / Q000235:genetics / Q000378:metabolism; D020825:T-Box Domain Proteins / Q000235:genetics / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000235:genetics / Q000378:metabolism; D000098908:Brachyury Protein
D000071616:CDX2 Transcription Factor; D002352:Carrier Proteins; D003893:Desmin; D005326:Fetal Proteins; D005346:Fibroblast Growth Factors; D050991:GATA4 Transcription Factor; D018398:Homeodomain Proteins; D009363:Neoplasm Proteins; D051496:Receptor, Fibroblast Growth Factor, Type 1; D051497:Receptor, Fibroblast Growth Factor, Type 2; D055760:SOXF Transcription Factors; D020825:T-Box Domain Proteins; D016212:Transforming Growth Factor beta; D000098908:Brachyury Protein; C491270:CDX2 protein, human; C491817:FRAT2 protein, human; C484170:GATA4 protein, human; C109188:SOX17 protein, human