Epithelial-mesenchymal transition in colonies of rhesus monkey embryonic stem cells: a model for processes involved in gastrulation.
Rhesus monkey embryonic stem (rhES) cells were grown on mouse embryonic fibroblast (MEF) feeder layers for up to 10 days to form multilayered colonies. Within this period, stem cell colonies differentiated transiently into complex structures with a disc-like morphology. These complex colonies were characterized by morphology, immunohistochemistry, and marker mRNA expression to identify processes of epithelialization as well as epithelial-mesenchymal transition (EMT) and pattern formation. Typically, differentiated colonies were comprised of an upper and a lower ES cell layer, the former growing on top of the layer of MEF cells whereas the lower ES cell layer spread out underneath the MEF cells. Interestingly, in the central part of the colonies, a roundish pit developed. Here the feeder layer disappeared, and upper layer cells seemed to ingress and migrate through the pit downward to form the lower layer while undergoing a transition from the epithelial to the mesenchymal phenotype, which was indicated by the loss of the marker proteins E-cadherin and ZO-1 in the lower layer. In support of this, we found a concomitant 10-fold upregulation of the gene Snail2, which is a key regulator of the EMT process. Conversion of epiblast to mesoderm was also indicated by the regulated expression of the mesoderm marker Brachyury. An EMT is a characteristic process of vertebrate gastrulation. Thus, these rhES cell colonies may be an interesting model for studies on some basic processes involved in early primate embryogenesis and may open new ways to study the regulation of EMT in vitro.
Behr
Rüdiger
R
Institut für Anatomie, Lehrstuhl für Anatomie und Entwicklungsbiologie, Univer-sitätsklinikum Essen, Hufelandstr. 55, D-45122 Essen, Germany.
Heneweer
Carola
C
Viebahn
Christoph
C
Denker
Hans-Werner
HW
Thie
Michael
M
D000199:Actins / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D015820:Cadherins / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D002460:Cell Line; D002465:Cell Movement; D002478:Cells, Cultured; D018031:Connexin 43 / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology*; D047108:Embryonic Development; D004848:Epithelium / Q000473:pathology*; D017629:Gap Junctions; D005775:Gastrula / Q000166:cytology / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008253:Macaca mulatta; D008565:Membrane Proteins / Q000378:metabolism; D008648:Mesoderm / Q000378:metabolism / Q000473:pathology*; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D023421:Models, Animal; D010641:Phenotype; D010750:Phosphoproteins / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D018075:RNA, Complementary / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013234:Stem Cells / Q000166:cytology*; D013997:Time Factors; D015854:Up-Regulation; D062826:Zonula Occludens-1 Protein
D000199:Actins; D015820:Cadherins; D018031:Connexin 43; D008565:Membrane Proteins; D010750:Phosphoproteins; D018075:RNA, Complementary; D012333:RNA, Messenger; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; D012313:RNA; D000469:Alkaline Phosphatase