Epithelial-mesenchymal transition in rhesus monkey embryonic stem cell colonies: the role of culturing conditions.

Colonies of rhesus monkey embryonic stem cells (rhESC; cell line R366.4) have been described before to show a spatially ordered process of epithelial-mesenchymal transition in vitro. In the present investigations, we have studied variables of culturing conditions which influence the reproducibility of the formation of crater-like ingression centers in the colonies. Critical parameters are found to be age and density of mouse embryonic fibroblast (MEF) feeder cell layers, the mode of mitotic inactivation of the MEFs (mitomycin C, or irradiation), and the mode of rhESC isolation during subculturing (enzymatic/mechanical cell cluster isolation; type of enzyme). The described culturing system appears to offer a reproducible in vitro model potentially useful for studies on cellular processes involved in gastrulation in the primate.