Multilineage differentiation of rhesus monkey embryonic stem cells in three-dimensional culture systems.
In the course of normal embryogenesis, embryonic stem (ES) cells differentiate along different lineages in the context of complex three-dimensional (3D) tissue structures. In order to study this phenomenon in vitro under controlled conditions, 3D culture systems are necessary. Here, we studied in vitro differentiation of rhesus monkey ES cells in 3D collagen matrixes (collagen gels and porous collagen sponges). Differentiation of ES cells in these 3D systems was different from that in monolayers. ES cells differentiated in collagen matrixes into neural, epithelial, and endothelial lineages. The abilities of ES cells to form various structures in two chemically similar but topologically different matrixes were different. In particular, in collagen gels ES cells formed gland-like circular structures, whereas in collagen sponges ES cells were scattered through the matrix or formed aggregates. Soluble factors produced by feeder cells or added to the culture medium facilitated ES cell differentiation into particular lineages. Coculture with fibroblasts in collagen gel facilitated ES cell differentiation into cells of a neural lineage expressing nestin, neural cell adhesion molecule, and class III beta-tubulin. In collagen sponges, keratinocytes facilitated ES cell differentiation into cells of an endothelial lineage expressing factor VIII. Exogenous granulocyte-macrophage colony-stimulating factor further enhanced endothelial differentiation. Thus, both soluble factors and the type of extracellular matrix seem to be critical in directing differentiation of ES cells and the formation of tissue-like structures. Three-dimensional culture systems are a valuable tool for studying the mechanisms of these phenomena.
NASA/NIH Center for Three Dimensional Tissue Culture, Laboratory of Cellular and Molecular Biophysics, National Institute of Child Health and Human Development (NICHD), NIH, Bethesda, Maryland 20892, USA.
Silvia S
SS
Chen
Roberto P
RP
Revoltella
Sandra
S
Papini
Monica
M
Michelini
Wendy
W
Fitzgerald
Joshua
J
Zimmerberg
Leonid
L
Margolis
D000818:Animals; D015415:Biomarkers; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000502:physiology*; D002455:Cell Division / Q000502:physiology; D019070:Cell Lineage / Q000502:physiology*; D002478:Cells, Cultured; D003094:Collagen; D017077:Culture Media, Conditioned / Q000494:pharmacology; D016207:Cytokines / Q000494:pharmacology; D015194:Diffusion Chambers, Culture; D005109:Extracellular Matrix / Q000502:physiology; D005781:Gelatin Sponge, Absorbable; D005782:Gels; D007150:Immunohistochemistry; D008253:Macaca mulatta; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000502:physiology*
D015415:Biomarkers; D017077:Culture Media, Conditioned; D016207:Cytokines; D005782:Gels; D003094:Collagen