Human embryonic stem (hES) cells are typically derived and serially propagated on inactivated murine embryonic fibroblast (MEF) feeders. The use of MEFs and other components of animal origin in the culture media for hES cell support substantially elevates the risk of contaminating these cell lines with infectious agents of animal origin thereby severely limiting their potential for clinical application. We have previously shown that it is possible to derive and establish new hES cell lines in a xeno-free culture system using human fetal muscle fibroblast feeders. In this report, we have comparatively evaluated a panel of 11 different human adult, fetal, and neonatal feeders for hES cell support and have ranked them as supportive and non-supportive. We report that two adult skin fibroblast cell lines established in-house from abdominal skin biopsies supported prolonged undifferentiated hES cell growth for over 30 weekly passages in culture. Furthermore, hES cell lines cultured on adult skin fibroblast feeders retain hES cell morphology and remain pluripotent. Also, differences in feeder support exist between human cell types and sources. The use of human adult skin feeders is convenient for hES cell support given the ease of obtaining skin biopsies.
All current 78 National Institutes of Health-listed human embryonic stem cell (hESC) lines approved for U.S. government federal research funding have been derived and propagated on mouse embryonic fibroblasts (MEFs) and in the presence of culture medium containing animal-based ingredients. The use of a feeder layer of animal origin and animal components in the culture media substantially elevates the risk of the cross-transfer of viruses and other pathogens to the embryonic stem (ES) cells. Hence, safer current good manufacturing practice (CGMP) and good tissue culture practice (GTCP)-compliant hESC lines and differentiated hESC progenitors need to be derived for clinical application.Several attempts at improving hESC culture conditions have been reported. These advances include the use of conditioned media together with Matrigel TM as an attachment sub-
Human embryonic stem cell (hESC) biology is expected to revolutionize the future of medicine by the provision of cell-based therapies for the treatment of a variety of deliberatig diseases. The tremendous versatility of hESCs has reinforced this hope. To understand the biology of these mysterious cells and attempt to differentiate them into desirable tissues, bona fide hESCs that maintain their stability with time are required for research and clinical application. This review discusses the various protocols to derive and propagate hESCs from high quality embryos. The nature and properties of hESCs are also described together with unanswered questions that need to be addressed if this science is to be taken to the bedside.
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