X chromosome inactivation in human parthenogenetic embryonic stem cells following prolonged passaging

The present study aimed to investigate the X chrochromosome inactivation (XCI) status in long-term cultured human parthenogenetic embryonic stem cells. One human embryonic stem (hES) cell line and 2 human parthenogenetic embryonic stem (hPES) cell lines were subjected to long-term culture in vitro (>50 passages). Karyotyping, array-based comparative genomic hybridization (aCGH), X-inactive specific transcript (XIST) RNA, immunofluorescence staining and real-time PCR were used to assess the chromosome karyotypes of these cells and the XCI status. X chromosome microdeletion was observed in the hPES-2 cells following culture for 50 passages. As early as 20 passages, XIST RNA expression was detected in the hPES-2 cells and was followed by low X-linked gene expression. The XIST RNA expression level was higher in the differentiated hPES-2 cells. The hPES-2′ cells that were subclones of hPES-2 retained the XCI status, and had low XIST and X-linked gene expression. XIST RNA expression remained at a low level in the differentiated hPES-2′ cells. The human biparental embryonic stem (hBES)-1 and hPES-1 cells did not exhibit XCI, and the differentiated hPES-1 cells had high expression levels of XIST RNA. In conclusion, the chromosome karyotypes of some hPES cell lines revealed instabilities. Similar to the hES cells, the hPES cells exhibited 3 XCI statuses. The unstable XCI status of the hPES-2 line may have been related to chromosome instability. These unstable chromosomes renedered these cells susceptible to environmental conditions and freezing processes, which may be the result of environmental adaptations.