Evaluation of X-Inactivation Status and Cytogenetic Stability of Human Dermal Fibroblasts after Long-Term Culture

• Published in: International Journal of Cell Biology 2010, Vol.2010, p.73-77
• Main Authors: Xue, Zhi-Gang, Shi, Zhan-Ping, Dong, Juan, Liao, Ting-Ting, Wang, Yan-Peng, Sun, Xue-Ping, Yan, Zheng-Jie, Qian, Xiao-Qiao, Cui, Yu-Gui, Chen, Juan, Liu, Jia-Yin, Fan, Guoping
• Format: Article
• Language: English
• Subjects: Chromosomes; DNA methylation; Reproductive health; Stem cells
• ISSN: 1687-8876; 1687-8884
• DOI: 10.1155/2010/289653

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).