Genotoxic Effects of Culture Media on Human Pluripotent Stem Cells.
Culture conditions play an important role in regulating the genomic integrity of Human Pluripotent Stem Cells (HPSCs). We report that HPSCs cultured in Essential 8 (E8) and mTeSR, two widely used media for feeder-free culturing of HPSCs, had many fold higher levels of ROS and higher mitochondrial potential than cells cultured in Knockout Serum Replacement containing media (KSR). HPSCs also exhibited increased levels of 8-hydroxyguanosine, phospho-histone-H2a.X and p53, as well as increased sensitivity to γ-irradiation in these two media. HPSCs in E8 and mTeSR had increased incidence of changes in their DNA sequence, indicating genotoxic stress, in addition to changes in nucleolar morphology and number. Addition of antioxidants to E8 and mTeSR provided only partial rescue. Our results suggest that it is essential to determine cellular ROS levels in addition to currently used criteria i.e. pluripotency markers, differentiation into all three germ layers and normal karyotype through multiple passages, in designing culture media.
National Centre for Biological Sciences (TIFR), Bangalore - 560065, India.|Manipal University, Madhav Nagar, Manipal 576104, Karnataka, India.
Megha
M
Prakash Bangalore
Institute for Stem Cell Biology and Regenerative Medicine, Bangalore 560065, India.
Syama
S
Adhikarla
Institute for Stem Cell Biology and Regenerative Medicine, Bangalore 560065, India.
Odity
O
Mukherjee
National Centre for Biological Sciences (TIFR), Bangalore - 560065, India.
Mitradas M
MM
Panicker
D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology; D002466:Cell Nucleolus / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D003470:Culture Media / Q000633:toxicity*; D004249:DNA Damage; D017930:Genes, Reporter; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D009153:Mutagens / Q000633:toxicity*; D039904:Pluripotent Stem Cells / Q000187:drug effects* / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
D000975:Antioxidants; D003470:Culture Media; D009153:Mutagens; D017382:Reactive Oxygen Species; D005978:Glutathione; D001205:Ascorbic Acid