title: Accumulation of spontaneous γH2AX foci in long-term cultured mesenchymal stromal cells.
abstract: Expansion of mesenchymal stromal/stem cells (MSCs) used in clinical practices may be associated with accumulation of genetic instability. Understanding temporal and mechanistic aspects of this process is important for improving stem cell therapy protocols. We used γH2AX foci as a marker of a genetic instability event and quantified it in MSCs that undergone various numbers of passage (3-22). We found that γH2AX foci numbers increased in cells of late passages, with a sharp increase at passage 16-18. By measuring in parallel foci of ATM phosphorylated at Ser-1981 and their co-localization with γaH2AX foci, along with differentiating cells into proliferating and resting by using a Ki67 marker, we conclude that the sharp increase in γH2AX foci numbers was ATM-independent and happened predominantly in proliferating cells. At the same time, gradual and moderate increase in γH2AX foci with passage number seen in both resting and proliferating cells may represent a slow, DNA double-strand break related component of the accumulation of genetic instability in MSCs. Our results provide important information on selecting appropriate passage numbers exceeding which would be associated with substantial risks to a patient-recipient, both with respect to therapeutic efficiency and side-effects related to potential neoplastic transformations due to genetic instability acquired by MSCs during expansion.
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Margarita
authors.initials: M
authors.lastname: Pustovalova
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Anna
authors.initials: A
authors.lastname: Grekhova
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Тatiana
authors.initials: Т
authors.lastname: Astrelina
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Viktoria
authors.initials: V
authors.lastname: Nikitina
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Dobrovolskaya
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Yulia
authors.initials: Y
authors.lastname: Suchkova
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Irina
authors.initials: I
authors.lastname: Kobzeva
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Darya
authors.initials: D
authors.lastname: Usupzhanova
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Natalia
authors.initials: N
authors.lastname: Vorobyeva
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Aleksandr
authors.initials: A
authors.lastname: Samoylov
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.
authors.forename: Andrey
authors.initials: A
authors.lastname: Bushmanov
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.|Insilico Medicine, Inc., Emerging Technology Centers, Johns Hopkins University at Eastern, Baltimore, MD 21218, USA.
authors.forename: Ivan V
authors.initials: IV
authors.lastname: Ozerov
authors.affiliation: Insilico Medicine, Inc., Emerging Technology Centers, Johns Hopkins University at Eastern, Baltimore, MD 21218, USA.|Life Sciences Center, Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region 141700, Russia.
authors.forename: Alex
authors.initials: A
authors.lastname: Zhavoronkov
authors.affiliation: Life Sciences Center, Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region 141700, Russia.
authors.forename: Sergey
authors.initials: S
authors.lastname: Leonov
authors.affiliation: Canadian Nuclear Laboratories, Chalk River, ON K0J1P0, Canada.
authors.forename: Dmitry
authors.initials: D
authors.lastname: Klokov
authors.affiliation: State Research Center-Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency (SRC-FMBC), Moscow 123098, Russia.|Insilico Medicine, Inc., Emerging Technology Centers, Johns Hopkins University at Eastern, Baltimore, MD 21218, USA.|Life Sciences Center, Moscow Institute of Physics and Technology, Dolgoprudny, Moscow Region 141700, Russia.
authors.forename: Andreyan N
authors.initials: AN
authors.lastname: Osipov
mesh_terms: D000328:Adult; D002454:Cell Differentiation; D049109:Cell Proliferation / Q000502:physiology*; D002478:Cells, Cultured; D042822:Genomic Instability; D006657:Histones / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism*; D010766:Phosphorylation
chemical_list: C468783:H2AX protein, human; D006657:Histones
keywords: DNA double-strand breaks; cellular senescence; genome instability; long-term cultivation; mesenchymal stromal cells; replicative senescence; γH2AX foci