Enhanced plating efficiency of trypsin-adapted human embryonic stem cells is reversible and independent of trisomy 12/17.
Human embryonic stem cells (hESCs) can be cultured abundantly and indefinitely, but are subject to accumulations of chromosomal aberrations. To preserve their genetic integrity, hESCs are commonly maintained as cell aggregates or clumps during passaging. However, clump passaging hinders large-scale culture and complicates the isolation of single cell clones. To facilitate the isolation of genetically modified clones of hESCs while preserving their genetic integrity, we employed trypsin single-cell passaging for brief periods before returning to clump passaging for long-term maintenance. We observed that accommodation to trypsin passage as single cells is an adaptive process where over three to four passages considerably increases the plating efficiency. However, trypsin passage was associated with abnormalities of chromosomes 12 and 17. Nevertheless, the high plating efficiency of trypsin passaged hESCs is a reversible phenotype, regardless of chromosomal abnormalities, suggesting that epigenetic events are responsible for the switch in phenotype.
Chan
Elayne M
EM
Division of Hematology/Oncology, Children's Hospital, Boston, Massachusetts 02115, USA.
Yates
Frank
F
Boyer
Leah F
LF
Schlaeger
Thorsten M
TM
Daley
George Q
GQ
D000222:Adaptation, Physiological / Q000187:drug effects* / Q000235:genetics; D000465:Algorithms; D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D002881:Chromosomes, Human, Pair 12; D002886:Chromosomes, Human, Pair 17; D002999:Clone Cells; D004526:Efficiency; D053595:Embryonic Stem Cells / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D005434:Flow Cytometry; D006801:Humans; D051379:Mice; D014314:Trisomy; D014357:Trypsin / Q000494:pharmacology*
D014357:Trypsin