Establishment of pluripotent cell lines from porcine preimplantation embryos

Embryonic stem (ES) cells are pluripotent cells isolated from in vitro cult ure of preimplantation embryos. Experiments were undertaken to identify pre implantation embryonic stages and culture conditions under which pluripoten t, porcine embryo-derived cell lines could be isolated. Cell lines were est ablished from in vitro culture of intact, porcine early hatched blastocysts and isolated inner cell masses (ICM) from intermediate and late hatched bl astocysts on feeder layers prepared from permanent mouse embryonic fibrobla sts (STO). The cells of these porcine embryo-derived cell lines had a morph ology similar to that of murine ES cells, but colony morphology was more ep itheliallike. The cell lines retained a normal diploid karyotype, consisten tly expressed alkaline phosphatase activity, and survived cryopreservation. When subjected to in vitro differentiation, either spontaneous or induced, the embryo-derived cell lines differentiated extensively into a wide range of cell types representing the 3 embryonic germ layers. In vivo pluripoten cy of the cells was demonstrated by birth of a chimeric piglet, documented by pigmentation and DNA markers, and the ability to direct the development of nuclear-transfer embryos to the blastocyst stage. Such pluripotent embry o-derived cells provide a potential route for porcine genetic manipulation. (C) 1999 by Elsevier Science Inc.