Comparative study of mouse and human feeder cells for human embryonic stem cells.
Various types of feeder cells have been adopted for the culture of human embryonic stem cells (hESCs) to improve their attachment and provide them with stemness-supporting factors. However, feeder cells differ in their capacity to support the growth of undifferentiated hESCs. Here, we compared the expression and secretion of four well-established regulators of hESC pluripotency and/or differentiation among five lines of human foreskin fibroblasts and primary mouse embryonic fibroblasts throughout a standard hESC culture procedure. We found that human and mouse feeder cells secreted comparable levels of TGF beta 1. However, mouse feeder cells secreted larger quantities of activin A than human feeder cells. Conversely, FGF-2, which was produced by human feeder cells, could not be detected in culture media from mouse feeder cells. The quantity of BMP-4 was at about the level of detectability in media from all feeder cell types, although BMP-4 dimers were present in all feeder cells. Production of TGF beta 1, activin A, and FGF-2 varied considerably among the human-derived feeder cell lines. Low- and high-producing human feeder cells as well as mouse feeder cells were evaluated for their ability to support the undifferentiated growth of hESCs. We found that a significantly lower proportion of hESCs maintained on human feeder cell types expressed SSEA3, an undifferentiated cell marker. Moreover, SSEA3 expression and thus the pluripotent hESC compartment could be partially rescued by addition of activin A. Cumulatively, these results suggest that the ability of a feeder layer to promote the undifferentiated growth of hESCs is attributable to its characteristic growth factor production.
Eiselleova
Livia
L
Department of Biology, Faculty of Medicine, Masaryk University, Brno, Czech Republic.
Peterkova
Iveta
I
Neradil
Jakub
J
Slaninova
Iva
I
Hampl
Ales
A
Dvorak
Petr
P
D028341:Activins / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D015295:Antigens, Tumor-Associated, Carbohydrate / Q000235:genetics; D001483:Base Sequence; D015415:Biomarkers / Q000378:metabolism; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000032:analysis / Q000235:genetics; D002454:Cell Differentiation; D049109:Cell Proliferation; D018920:Coculture Techniques / Q000379:methods*; D017077:Culture Media, Conditioned / Q000032:analysis; D017931:DNA Primers / Q000235:genetics; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D006028:Glycosphingolipids / Q000235:genetics; D006801:Humans; D051379:Mice; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D055746:Stage-Specific Embryonic Antigens; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis / Q000235:genetics
D015295:Antigens, Tumor-Associated, Carbohydrate; C525809:BMP4 protein, human; D015415:Biomarkers; C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D017077:Culture Media, Conditioned; D017931:DNA Primers; D006028:Glycosphingolipids; D012333:RNA, Messenger; D055746:Stage-Specific Embryonic Antigens; D053773:Transforming Growth Factor beta1; C422405:activin A; C052924:stage-specific embryonic antigen-3; D016222:Fibroblast Growth Factor 2; D028341:Activins