title: Evaluation of x-inactivation status and cytogenetic stability of human dermal fibroblasts after long-term culture.
abstract: Human primary fibroblasts are a popular type of somatic cells for the production of induced pluripotent stem (iPS) cells. Here we characterized biological properties of primary fibroblasts in terms of cell-growth rate, cytogenetic stability, and the number of inactive X chromosomes during long-term passaging. We produced eight lines of female human dermal fibroblasts (HDFs) and found normal karyotype and expected pattern of X chromosome inactivation (XCI) at low passages (Passage P1-5). However, four out of the eight HDF lines at high passage numbers (≥ P10) exhibited duplicated hallmarks of inactive X chromosome including two punctuate signals of histone H3 lysine 27 trimethylation (H3K27me3) and X inactive-specific transcript (XIST) RNA signals in approximately 8.5-18.5% of the cells. Our data suggest that the copy number of inactive X chromosomes in a subset of female HDF is increased by a two-fold. Consistently, DNA fluorescent in situ hybridization (FISH) identified 3-4 copies of X chromosomes in one nucleus in this subset of cells with two inactive Xs. We conclude that female HDF cultures exhibit a higher risk of genetic anomalies such as carrying an increased number of X chromosomes including both active and inactive X chromosomes at a high passage (≥ P10).
authors.affiliation: Jiangsu Province Key Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing 210029, China.
authors.forename: Juan
authors.initials: J
authors.lastname: Chen
authors.forename: Zhan-Ping
authors.initials: ZP
authors.lastname: Shi
authors.forename: Juan
authors.initials: J
authors.lastname: Dong
authors.forename: Ting-Ting
authors.initials: TT
authors.lastname: Liao
authors.forename: Yan-Peng
authors.initials: YP
authors.lastname: Wang
authors.forename: Xue-Ping
authors.initials: XP
authors.lastname: Sun
authors.forename: Zheng-Jie
authors.initials: ZJ
authors.lastname: Yan
authors.forename: Xiao-Qiao
authors.initials: XQ
authors.lastname: Qian
authors.forename: Yu-Gui
authors.initials: YG
authors.lastname: Cui
authors.forename: Zhi-Gang
authors.initials: ZG
authors.lastname: Xue
authors.forename: Guoping
authors.initials: G
authors.lastname: Fan
authors.forename: Jia-Yin
authors.initials: JY
authors.lastname: Liu