© Andrei Tchernov
Human embryonic stem cells appear to accrue genomic changes that could make them unusable therapeutically when cultured at length. Anirban Maitra of Johns Hopkins University School of Medicine and colleagues in Sweden, Canada, Singapore, and the United States analyzed early- and late-passage cultures of nine human embryonic stem-cell lines approved for use by the US federal government.1 The late batches were cultured 22 to 175 passages more than early counterparts.
Four late-passage lines developed copy number aberrations. These ranged from large genomic amplifications or deletions, such as amplification of the entire 17q arm, to more discrete changes, such as a two-megabase amplification encompassing the MYC oncogene. The researchers found mitochondrial DNA sequence alterations in two late-passage lines, with five coding region changes: three resulting in missense mutations in NADH dehydrogenases and one causing a nonsense mutation in ATPase 6. Real-time quantitative methylation-specific PCR assays of 14 genes with known differential methylation patterns in cancer tissues revealed higher promoter methylation levels in eight late-passage lines.
"The very act of culturing and repeatedly passing cells introduces a selection process, and cells that accumulate any kind of DNA modification, genetic or epigenetic, that confers an advantage for doubling are ultimately going to take over," says James Battey, chair of the National Institutes of Health stem cell task force. "We need to be vigilant with these cells."