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Propagation of human embryonic and induced pluripotent stem cells in an indirect co-culture system |
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| Authors: | Sheena Abraham Steven D Sheridan Louise C Laurent Kelsey Albert Igor Ulitsky Jeanne F Loring Raj R Rao |
| Institution: | a Chemical and Life Science Engineering, Virginia Commonwealth University, Richmond, VA, USA b Center for Human Genetic Research, Harvard Medical School, Massachusetts General Hospital, Boston, MA, USA c Department of Brain and Cognitive Science, Picower Institute for Learning and Memory, Massachusetts Institute of Technology, Cambridge, MA 02139, USA d Center for Regenerative Medicine, Scripps Research Institute, La Jolla, CA, USA e Reproductive Medicine, University of California, San Diego, CA, USA f Burnham Institute for Medical Research, La Jolla, CA, USA g School of Computer Science, Tel Aviv University, Israel h Department of Pathology, University of Virginia, Charlottesville, VA, USA |
| Abstract: | We have developed and validated a microporous poly(ethylene terephthalate) membrane-based indirect co-culture system for human pluripotent stem cell (hPSC) propagation, which allows real-time conditioning of the culture medium with human fibroblasts while maintaining the complete separation of the two cell types. The propagation and pluripotent characteristics of a human embryonic stem cell (hESC) line and a human induced pluripotent stem cell (hiPSC) line were studied in prolonged culture in this system. We report that hPSCs cultured on membranes by indirect co-culture with fibroblasts were indistinguishable by multiple criteria from hPSCs cultured directly on a fibroblast feeder layer. Thus this co-culture system is a significant advance in hPSC culture methods, providing a facile stem cell expansion system with continuous medium conditioning while preventing mixing of hPSCs and feeder cells. This membrane culture method will enable testing of novel feeder cells and differentiation studies using co-culture with other cell types, and will simplify stepwise changes in culture conditions for staged differentiation protocols. |
| Keywords: | Human pluripotent stem cells Microporous membranes Indirect co-culture Cell separation |
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