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

Abstract

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.

Non‐coding RNAs: emerging regulatory factors in the derivation and differentiation of mammalian parthenogenetic embryonic stem cells

“…As early as 1990s, imprinted lncRNA genes X-inactive specific transcript (Xist) and H19 have aroused researchers' interest (Uh et al, 2014;Qi et al, 2015). Qi et al (2015) studied XCI status in long-term cultured hPESCs. Xchromosome microdeletion was observed in the hPESCs cells after culture for 50 passages.…”

“…Both X-chromosomes in PESCs originate from a single X in the MII oocyte, so X-chromosome inactivation (XCI) and parental imprinting are very important for PESCs (Feil et al, 1994). As early as 1990s, imprinted lncRNA genes X-inactive specific transcript (Xist) and H19 have aroused researchers' interest (Uh et al, 2014;Qi et al, 2015). Qi et al (2015) studied XCI status in long-term cultured hPESCs.…”