Defining the role of Wnt/beta-catenin signaling in the survival, proliferation, and self-renewal of human embryonic stem cells.
We used a panel of human and mouse fibroblasts with various abilities for supporting the prolonged growth of human embryonic stem cells (hESCs) to elucidate growth factors required for hESC survival, proliferation, and maintenance of the undifferentiated and pluripotent state (self-renewal). We found that supportive feeder cells secrete growth factors required for both hESC survival/proliferation and blocking hESC spontaneous differentiation to achieve self-renewal. The antidifferentiation soluble factor is neither leukemia inhibitory factor nor Wnt, based on blocking experiments using their antagonists. Because Wnt/beta-catenin signaling has been implicated in cell-fate determination and stem cell expansion, we further examined the effects of blocking or adding recombinant Wnt proteins on undifferentiated hESCs. In the absence of feeder cell-derived factors, hESCs cultured under a feeder-free condition survived/proliferated poorly and gradually differentiated. Adding recombinant Wnt3a stimulated hESC proliferation but also differentiation. After 4-5 days of Wnt3a treatment, hESCs that survived maintained the undifferentiated phenotype but few could form undifferentiated hESC colonies subsequently. Using a functional reporter assay, we found that the beta-catenin-mediated transcriptional activation in the canonical Wnt pathway was minimal in undifferentiated hESCs, but greatly upregulated during differentiation induced by the Wnt treatment and several other methods. Thus, Wnt/beta-catenin activation does not suffice to maintain the undifferentiated and pluripotent state of hESCs. We propose a new model for the role of Wnt/beta-catenin signaling in undifferentiated hESCs.
Dravid
Gautam
G
The Institute for Cell Engineering, Department of Gynecology & Obstetrics, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Ye
Zhaohui
Z
Hammond
Holly
H
Chen
Guibin
G
Pyle
April
A
Donovan
Peter
P
Yu
Xiaobing
X
Cheng
Linzhao
L
D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D049109:Cell Proliferation; D033041:Embryo Research; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008954:Models, Biological; D015398:Signal Transduction; D013234:Stem Cells / Q000166:cytology*; D015854:Up-Regulation; D051153:Wnt Proteins / Q000235:genetics / Q000494:pharmacology / Q000502:physiology*; D051176:beta Catenin / Q000235:genetics / Q000502:physiology*
D051153:Wnt Proteins; D051176:beta Catenin; D005346:Fibroblast Growth Factors