title: DNA damage response in neonatal and adult stromal cells compared with induced pluripotent stem cells.
abstract: Comprehensive analyses comparing individual DNA damage response (DDR) of induced pluripotent stem cells (iPSCs) with neonatal stromal cells with respect to their developmental age are limited. The imperative necessity of providing developmental age-matched cell sources for meaningful toxicological drug safety assessments in replacement of animal-based testing strategies is evident. Here, DDR after radiation or treatment with N-methyl-N-nitrosurea (MNU) was determined in iPSCs compared with neonatal and bone marrow stromal cells. Neonatal and adult stromal cells showed no significant morphologically detectable cytotoxicity following treatment with 1 Gy or 1 mM MNU, whereas iPSCs revealed a much higher sensitivity. Foci analyses revealed an effective DNA repair in stromal cell types and iPSCs, as reflected by a rapid formation and disappearance of phosphorylated ATM and γH2AX foci. Furthermore, quantitative polymerase chain reaction analyses revealed the highest basic expression level of DDR and repair-associated genes in iPSCs, followed by neonatal stromal cells and adult stromal cells with the lowest expression levels. In addition, the influence of genotoxic stress prior to and during osteogenic differentiation of neonatal and adult stromal cells was analyzed applying common differentiation procedures. Experiments presented here suggest a developmental age-dependent basic expression level of genes involved in the processing of DNA damage. In addition a differentiation-dependent downregulation of repair genes was observed during osteogenesis. These results strongly support the requirement to provide adequate cell sources for toxicological in vitro drug testing strategies that match to the developmental age and differentiation status of the presumptive target cell of interest.
  
  
  CONCLUSIONS
  The results obtained in this study advance the understanding of DNA damage processing in human neonatal stromal cells as compared with adult stromal cells and induced pluripotent stem cells (iPSCs). The data suggest developmental age-dependent differences in DNA damage repair capacity. In iPSCs (closest to embryonic stem cells), the highest expression level of DNA damage response and repair genes was found, followed by neonatal stromal cells and adult stromal cells with the lowest overall expression. In addition, a differentiation-dependent downregulation of repair capacity was observed during osteogenic differentiation in neonatal stromal cells. Notably, the impact of genotoxic stress on osteogenic differentiation depended on the time the genotoxic insult took place and, moreover, was agent-specific. These results strongly support the necessity of offering and establishing adequate cell sources for informative toxicological testing matching to the developmental age and differentiation status of the respective cell of interest.
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Stefanie
authors.initials: S
authors.lastname: Liedtke
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Sophie
authors.initials: S
authors.lastname: Biebernick
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Teja Falk
authors.initials: TF
authors.lastname: Radke
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Daniela
authors.initials: D
authors.lastname: Stapelkamp
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Carolin
authors.initials: C
authors.lastname: Coenen
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Holm
authors.initials: H
authors.lastname: Zaehres
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Fritz
authors.affiliation: Institute for Transplantation Diagnostics and Cell Therapeutics and Institute of Toxicology, Heinrich-Heine-University Medical Center, Düsseldorf, Germany; Department Cell and Developmental Biology, Max Planck Institute for Molecular Biomedicine, Münster, Germany.
authors.forename: Gesine
authors.initials: G
authors.lastname: Kogler
mesh_terms: D000328:Adult; D000477:Alkylating Agents / Q000494:pharmacology*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D004249:DNA Damage; D004353:Drug Evaluation, Preclinical / Q000379:methods; D005260:Female; D006657:Histones / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D007231:Infant, Newborn; D008297:Male; D008770:Methylnitrosourea / Q000494:pharmacology*; D010012:Osteogenesis / Q000187:drug effects*; D010766:Phosphorylation / Q000187:drug effects; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism; D014965:X-Rays
chemical_list: D000477:Alkylating Agents; C468783:H2AX protein, human; D006657:Histones; D008770:Methylnitrosourea; C576324:ATM protein, human; D064007:Ataxia Telangiectasia Mutated Proteins
keywords: DNA Repair; DNA damage response; Ionizing radiation; MNU; MSC; Neonatal and adult stromal cells; Osteogenesis; iPSC