title: X-ray and ultraviolet C irradiation-induced γ-H2AX and p53 formation in normal human periosteal cells in vitro: markers for quality control in cell therapy.
abstract: OBJECTIVE
  For successful cell transplantation therapy, the quality of cells must be strictly controlled. Unfortunately, to exclude inappropriate cells that possess structurally abnormal chromosomes, currently only karyotyping functions as an assessment. Unfortunately, this methodology is time-consuming and only effective for metaphasic cells. To develop a more efficient, inclusive and sensitive methodology, we examined the phosphorylation of histone H2AX and the p53 levels in normal human periosteal cells exposed to x-rays or other oxidative stressors.
  
  
  METHODS
  Periosteal cells were obtained from human alveolar bone before being exposed to x-rays, ultraviolet C or hydrogen peroxide. The cell cycle, electric nuclear volume and CD44 expression were evaluated using flow cytometry, and the phosphorylated H2AX (γ-H2AX), p53, p21 and proliferating cell nuclear antigen (PCNA) levels were evaluated by Western blot analyses.
  
  
  RESULTS
  Each oxidative stress dose-dependently arrested cell growth and partially induced premature cellular senescence. In parallel, each oxidative stress rapidly phosphorylated H2AX and stabilized p53, and intense stress sustained these high levels for at least 8 days.
  
  
  CONCLUSIONS
  Intensive oxidative stress induces sustained high levels of γ-H2AX and p53, which force cells toward senescence or non-apoptotic cell death. Lower doses of oxidative stress induced more modest and transient increases in γ-H2AX and p53, and these cells eventually survive. However, because DNA is repaired without a template in the majority of these cells, G1 mutations accumulate. Therefore, we recommend that any cell population expressing elevated γ-H2AX and p53 levels be excluded from cell transplantation therapy.
authors.affiliation: Division of Oral Bioengineering, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan; Advanced Research Center, The Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan. Electronic address: kawase@dent.niigatau.ac.jp.
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Kawase
authors.affiliation: Division of Oral Bioengineering, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan; Division of Periodontology, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.
authors.forename: Mana
authors.initials: M
authors.lastname: Kamiya
authors.affiliation: Department of Oral and Maxillofacial Radiology, The Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan.
authors.forename: Kazuhide
authors.initials: K
authors.lastname: Hayama
authors.affiliation: Division of Oral and Maxillofacial Surgery, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Nagata
authors.affiliation: Division of Periodontology, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Okuda
authors.affiliation: Division of Periodontology, Institute of Medicine and Dentistry, Niigata University, Niigata, Japan.
authors.forename: Hiromasa
authors.initials: H
authors.lastname: Yoshie
authors.affiliation: Midwest Biomedical Research Foundation, Research Service, Department of Veterans Affairs Medical Center, Kansas City, Missouri, USA.
authors.forename: Douglas M
authors.initials: DM
authors.lastname: Burns
authors.affiliation: Department of Oral and Maxillofacial Radiology, The Nippon Dental University School of Life Dentistry at Niigata, Niigata, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Tsuchimochi
authors.affiliation: Bioscience Medical Research Center, Niigata University Medical and Dental Hospital, Niigata, Japan.
authors.forename: Koh
authors.initials: K
authors.lastname: Nakata
mesh_terms: D002453:Cell Cycle / Q000502:physiology / Q000528:radiation effects; D064987:Cell- and Tissue-Based Therapy / Q000379:methods; D016922:Cellular Senescence / Q000502:physiology / Q000528:radiation effects; D006657:Histones / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D018384:Oxidative Stress / Q000528:radiation effects; D011786:Quality Control; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D014466:Ultraviolet Rays; D014965:X-Rays
chemical_list: C468783:H2AX protein, human; D006657:Histones; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53
keywords: cell cycle; oxidative stress; p53; periosteal cells; quality control; γ-H2AX