Whole-Genome Sequencing Identifies Genetic Variances in Culture-Expanded Human Mesenchymal Stem Cells

•Whole-genome investigation reveals variances of cultured MSCs along serial passages•Human MSCs maintain a stable genomic composition in early stages of ex vivo culture•Human MSCs are subject to clonal growth upon extended expansion•The “new” SNCs present in the expanded MSCs preexist in the early cell population

SummaryCulture-expanded human mesenchymal stem cells (MSCs) are increasingly used in clinics, yet full characterization of the genomic compositions of these cells is lacking. We present a whole-genome investigation on the genetic dynamics of cultured MSCs under ex vivo establishment (passage 1 [p1]) and serial expansion (p8 and p13). We detected no significant changes in copy-number alterations (CNAs) and low levels of single-nucleotide changes (SNCs) until p8. Strikingly, a significant number (677) of SNCs were found in p13 MSCs. Using a sensitive Droplet Digital PCR assay, we tested the nonsynonymous SNCs detected by whole-genome sequencing and found that they were preexisting low-frequency mutations in uncultured mononuclear cells (∼0.01%) and early-passage MSCs (0.1%–1% at p1 and p8) but reached 17%–36% in p13. Our data demonstrate that human MSCs maintain a stable genomic composition in the early stages of ex vivo culture but are subject to clonal growth upon extended expansion.

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