title: Dynamic monitoring of oxidative DNA double-strand break and repair in cardiomyocytes.
abstract: DNA double-strand breaks (DSBs) are most dangerous lesions. To determine whether oxidative stress can induce DSBs and how they are repaired in cardiomyocytes (CMs), cultured neonatal rat CMs were treated with different doses of H2O2 and followed for up to 72 h for monitoring the spatiotemporal dynamics of DNA repair protein assembly/disassembly at DSB foci. The protein levels and foci numbers of histone H2AX phosphorylated at serine 139 (γ-H2AX) increased proportionally to 50, 100, and 200 μmol/L H2O2 after 30 min treatment. When H2O2 was at or above 400 μmol/L, γ-H2AX became predominantly pannuclear. After 30 min, 200 μmol/L of H2O2 treatment, γ-H2AX levels were highest within the first hour and then gradually declined during the recovery and returned to basal levels at 48 h. Among DNA damage transducer kinases, ataxia telangiectasia mutated (ATM) was significantly activated by H2O2 in contrast to mild activation of ATR (ATM and Rad3-related). A DSB binding protein, p53 binding protein 1, formed distinct nuclear foci that colocalized with γ-H2AX foci and phosphorylated ATM. Our findings indicate that DSBs can be induced by H2O2 and ATM is the main kinase to mediate DSB repair in CMs. Therefore, monitoring DSB repair can assess oxidative injury and response in CMs.
authors.affiliation: Department of Laboratory Medicine and Pathology, University of Minnesota, Room 293, Dwan Variety Club Cardiovascular Research Center, 425 East River Parkway, Minneapolis, MN 55455, USA; Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA.
authors.forename: Bo
authors.initials: B
authors.lastname: Ye
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA.
authors.forename: Ning
authors.initials: N
authors.lastname: Hou
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA.
authors.forename: Lu
authors.initials: L
authors.lastname: Xiao
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA.
authors.forename: Yifan
authors.initials: Y
authors.lastname: Xu
authors.affiliation: Department of Pathology and Laboratory Medicine, UCLA Center for the Health Science, Room 13-145E, 10833 Le Conte Avenue, Los Angeles, CA 90095, USA.
authors.forename: Haodong
authors.initials: H
authors.lastname: Xu
authors.affiliation: Department of Laboratory Medicine and Pathology, University of Minnesota, Room 293, Dwan Variety Club Cardiovascular Research Center, 425 East River Parkway, Minneapolis, MN 55455, USA; Department of Pathology and Laboratory Medicine, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA. Electronic address: lif@umn.edu.
authors.forename: Faqian
authors.initials: F
authors.lastname: Li
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D053903:DNA Breaks, Double-Stranded; D004260:DNA Repair / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D006861:Hydrogen Peroxide / Q000633:toxicity; D018613:Microscopy, Confocal; D032383:Myocytes, Cardiac / Q000473:pathology*; D016877:Oxidants / Q000633:toxicity; D018384:Oxidative Stress / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D016877:Oxidants; D006861:Hydrogen Peroxide
keywords: Cultured neonatal rat cardiomyocytes; DNA damage; DNA double-strand breaks; H(2)O(2); Heart; Oxidative stress