Human Adipose-Derived Stem Cells Expanded Under Ambient Oxygen Concentration Accumulate Oxidative DNA Lesions and Experience Procarcinogenic DNA Replication Stress.
Adipose-derived stem cells (ADSCs) have led to growing interest in cell-based therapy because they can be easily harvested from an abundant tissue. ADSCs must be expanded in vitro before transplantation. This essential step causes concerns about the safety of adult stem cells in terms of potential transformation. Tumorigenesis is driven in its earliest step by DNA replication stress, which is characterized by the accumulation of stalled DNA replication forks and activation of the DNA damage response. Thus, to evaluate the safety of ADSCs during ex vivo expansion, we monitored DNA replication under atmospheric (21%) or physiologic (1%) oxygen concentration. Here, by combining immunofluorescence and DNA combing, we show that ADSCs cultured under 21% oxygen accumulate endogenous oxidative DNA lesions, which interfere with DNA replication by increasing fork stalling events, thereby leading to incomplete DNA replication and fork collapse. Moreover, we found by RNA sequencing (RNA-seq) that culture of ADSCs under atmospheric oxygen concentration leads to misexpression of cell cycle and DNA replication genes, which could contribute to DNA replication stress. Finally, analysis of acquired small nucleotide polymorphism shows that expansion of ADSCs under 21% oxygen induces a mutational bias toward deleterious transversions. Overall, our results suggest that expanding ADSCs at a low oxygen concentration could reduce the risk for DNA replication stress-associated transformation, as occurs in neoplastic tissues. Stem Cells Translational Medicine 2017;6:68-76.
Equipe Labellisée La Ligue Contre Le Cancer, Paris, France.|Laboratoire d'Excellence Toulouse Cancer Labex Toucan, Cancer Research Center of Toulouse, INSERM U1037, CNRS ERL5294, Toulouse, France.|University Paul Sabatier, Toulouse, France.
Rémy
R
Bétous
University Paul Sabatier, Toulouse, France.|Etablissement Français du Sang Pyrénées Méditerranée, Toulouse, France.|INSERM U1031, UMR5273, Toulouse, France.
Marie-Laure
ML
Renoud
Institut National de la Recherche Agronomique (INRA), UR 875, Unité de Mathématique et Informatique Appliquées, PF Bioinfo Genotoul, Castanet Tolosan, France.
Claire
C
Hoede
Institut National de la Recherche Agronomique (INRA), UR 875, Unité de Mathématique et Informatique Appliquées, PF Bioinfo Genotoul, Castanet Tolosan, France.
Ignacio
I
Gonzalez
INSERM U916 Vinco, Université de Bordeaux, Institut Bergonié, Bordeaux, France.
Natalie
N
Jones
INSERM U916 Vinco, Université de Bordeaux, Institut Bergonié, Bordeaux, France.
Michel
M
Longy
University Paul Sabatier, Toulouse, France.|Etablissement Français du Sang Pyrénées Méditerranée, Toulouse, France.|INSERM U1031, UMR5273, Toulouse, France.
Luc
L
Sensebé
Equipe Labellisée La Ligue Contre Le Cancer, Paris, France.|Laboratoire d'Excellence Toulouse Cancer Labex Toucan, Cancer Research Center of Toulouse, INSERM U1037, CNRS ERL5294, Toulouse, France.|University Paul Sabatier, Toulouse, France.
Christophe
C
Cazaux
Equipe Labellisée La Ligue Contre Le Cancer, Paris, France.|Laboratoire d'Excellence Toulouse Cancer Labex Toucan, Cancer Research Center of Toulouse, INSERM U1037, CNRS ERL5294, Toulouse, France.|University Paul Sabatier, Toulouse, France.
Jean-Sébastien
JS
Hoffmann
D000273:Adipose Tissue / Q000166:cytology*; D063646:Carcinogenesis / Q000473:pathology*; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002877:Chromosomes, Human / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication / Q000187:drug effects*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D009154:Mutation / Q000235:genetics; D018384:Oxidative Stress / Q000187:drug effects*; D010100:Oxygen / Q000494:pharmacology*; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D013312:Stress, Physiological
D004247:DNA; D010100:Oxygen
DNA damage; DNA replication stress; Ex vivo expansion; Human adipose-derived stem cells; Oxygen concentration