Abstract
BackgroundFor therapeutic usage of induced Pluripotent Stem (iPS) cells, to accomplish xeno-free culture is critical. Previous reports have shown that human embryonic stem (ES) cells can be maintained in feeder-free condition. However, absence of feeder cells can be a hostile environment for pluripotent cells and often results in karyotype abnormalities. Instead of animal feeders, human fibroblasts can be used as feeder cells of human ES cells. However, one still has to be concerned about the existence of unidentified pathogens, such as viruses and prions in these non-autologous feeders.Methodology/Principal FindingsThis report demonstrates that human induced Pluripotent Stem (iPS) cells can be established and maintained on isogenic parental feeder cells. We tested four independent human skin fibroblasts for the potential to maintain self-renewal of iPS cells. All the fibroblasts tested, as well as their conditioned medium, were capable of maintaining the undifferentiated state and normal karyotypes of iPS cells. Furthermore, human iPS cells can be generated on isogenic parental fibroblasts as feeders. These iPS cells carried on proliferation over 19 passages with undifferentiated morphologies. They expressed undifferentiated pluripotent cell markers, and could differentiate into all three germ layers via embryoid body and teratoma formation.Conclusions/SignificanceThese results suggest that autologous fibroblasts can be not only a source for iPS cells but also be feeder layers. Our results provide a possibility to solve the dilemma by using isogenic fibroblasts as feeder layers of iPS cells. This is an important step toward the establishment of clinical grade iPS cells.
Highlights
Both embryonic stem (ES) cells and induced Pluripotent Stem cells, are generally maintained on mouse embryonic fibroblasts (MEF), which are mitotically inactivated by treatment with mitomycin C or c-ray irradiation [1,2,3]
All the five cell lines of feeder cells were supportive for undifferentiated growth of induced Pluripotent Stem (iPS) cells at least 19 additional passages (Fig. 1A)
The percentage of TRA-1-60 positive colonies was similar among different human fibroblasts and SNL cells (Fig. 1B)
Summary
Both embryonic stem (ES) cells and induced Pluripotent Stem (iPS) cells, are generally maintained on mouse embryonic fibroblasts (MEF), which are mitotically inactivated by treatment with mitomycin C or c-ray irradiation [1,2,3]. To examine whether human fibroblasts support self-renewal of human iPS cells, we treated four independent human fibroblast lines (1388, 1392, 1503 and NHDF; see Table S1) and SNL cells [12] with mitomycin C, and seeded them on culture plates (Fig. S1). All the five cell lines of feeder cells were supportive for undifferentiated growth of iPS cells at least 19 additional passages (Fig. 1A).
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