title: Efficient derivation of human embryonic stem cell lines from discarded embryos through increases in the concentration of basic fibroblast growth factor.
abstract: We describe the derivation and characterization of three novel human embryonic stem (hES) cell lines (YT1, YT2, YT3). One hES line (YT1) was obtained from six discarded blastocysts in a culture medium supplemented with 12 ng/ml basic fibroblast growth factor (bFGF), and two lines (YT2,YT3)were obtained from three discarded blastocysts in the same medium but supplemented with 16 ng/ml bFGF. These cell lines were derived by partial or whole embryo culture followed by further expansion after manual dissection of the passaged cells. These cells were passaged continuously for more than 6 or 8 months and possessed all of the typical features of pluripotent hES cell lines, such as typical morphological characteristics and the expression of hES-specific markers (TRA-1-60, TRA-1-81, SSEA-4, SSEA-3, alkaline phosphatase, Oct4, Nanog) and pluripotency-related genes (Oct4, Nanog, TDGF1, Sox2, EBAF, Thy-1, FGF4, Rex1). The lines maintained normal karyotypes after long-term cultivation. The karyotype of YT1 and YT3 was 46,XX, and that of YT2 was 46, XY. Pluripotency was confirmed by in vitro and in vivo differentiation, and genetic identity was demonstrated by DNA fingerprinting.Our results indicate that higher concentrations of bFGF at the early culture stage support efficient the hES cell derivation.
authors.affiliation: Shandong Research Center of Stem Cell Engineering, Yantai Yuhuangding Hospital, Shandong Province, People's Republic of China.
authors.forename: Yanwei
authors.initials: Y
authors.lastname: Wang
authors.forename: Chenming
authors.initials: C
authors.lastname: Xu
authors.forename: Haiyan
authors.initials: H
authors.lastname: Wang
authors.forename: Juan
authors.initials: J
authors.lastname: Liu
authors.forename: Shi
authors.initials: S
authors.lastname: Hui
authors.forename: Ning
authors.initials: N
authors.lastname: Li
authors.forename: Fujun
authors.initials: F
authors.lastname: Liu
authors.forename: Jianyuan
authors.initials: J
authors.lastname: Li
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002460:Cell Line; D002469:Cell Separation; D046149:Embryo Culture Techniques; D053595:Embryonic Stem Cells / Q000737:chemistry* / Q000166:cytology; D016222:Fibroblast Growth Factor 2 / Q000032:analysis*; D006801:Humans; D013997:Time Factors; D009927:Tissue and Organ Procurement
chemical_list: D016222:Fibroblast Growth Factor 2