Differentiation of Rhesus Monkey Embryonic Stem Cells in Three-dimensional Collagen Matrix.
From: NASA/NIH Center for Three Dimensional Tissue Culture, Laboratory of Cellular and Molecualr Biophysics, Bethesda, MD, USA.
Methods in molecular biology (Clifton, N.J.)
- Publish Date: 2006
- ISSN: 1064-3745
- Volume: 330
- Issue:
- Pages: 431-43
- Medium: Print
- Language: English
- Citation (JAMA): Chen Silvia Sihui, Revoltella Roberto P, Zimmerberg Joshua, et al. Differentiation of Rhesus Monkey Embryonic Stem Cells in Three-dimensional Collagen Matrix.. Methods Mol. Biol. 2006;330:431-43
Abstract
During normal embryogenesis, embryonic stem cells (ESCs) reside in the context of complex three-dimensional tissue structures, in particular of extracellular matrices (ECMs), which determine cell migration, proliferation, and differentiation. Therefore, to study ESC differentiation in an in vivo-like microenvironment, three-dimensional culture systems are necessary. Here, we developed protocols for ESC cultures in three-dimensional systems consisting of collagen matrices (collagen gels and porous collagen sponges) to investigate the mechanisms of ESC differentiation as well as the formation of tissue-like structures. In collagen matrices, ESCs differentiate into neural, epithelial, and endothelial lineages. In this system, ESCs form various tissue-like structures. The abilities of ESCs to form such structures in two chemically similar but topologically different matrices are different. In particular, in collagen gels ESCs form gland-like circular structures, whereas in collagen sponges ESCs are scattered through the matrix and form aggregates. To mimic the in vivo situation further, we developed a protocol for co-cultures of ESCs with human dermal fibroblasts or keratinocytes in collagen matrixes. Co-culture with fibroblasts in collagen gel facilitates ESC differentiation into cells of a neural lineage expressing nestin, neural cell adhesion molecule (NCAM), and class III beta-tubulin. In collagen sponges, keratinocytes facilitated ESC differentiation into cells of an endothelial lineage expressing factor VIII. Thus, the developed protocols promote ESC differentiation into a particular lineage, accompanied by the formation of tissue-like structures. Three-dimensional culture systems are a valuable tool for directing ESC differentiation and the formation of organs and tissues.
Mesh Headings (Keywords): Animals, Biological Markers, Cell Culture Techniques, Cell Differentiation, Cells, Cultured, Collagen, Embryo, Mammalian, Extracellular Matrix, Humans, Immunohistochemistry, Macaca mulatta, Stem Cells
Check for Full Text / PubMed Unique Identifier (PMID): 16846041
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