title: Histone H2AX phosphorylation as a measure of DNA double-strand breaks and a marker of environmental stress and disease activity in lupus.
abstract: OBJECTIVE
  Defective or inefficient DNA double-strand break (DSB) repair results in failure to preserve genomic integrity leading to apoptotic cell death, a hallmark of systemic lupus erythematosus (SLE). Compelling evidence linked environmental factors that increase oxidative stress with SLE risk and the formation of DSBs. In this study, we sought to further explore genotoxic stress sensitivity in SLE by investigating DSB accumulation as a marker linking the effect of environmental stressors and the chromatin microenvironment.
  
  
  METHODS
  DSBs were quantified in peripheral blood mononuclear cell subsets from patients with SLE, healthy controls, and patients with rheumatoid arthritis (RA) by measuring phosphorylated H2AX (phospho-H2AX) levels with flow cytometry. Phospho-H2AX levels were assessed in G0/G1, S and G2 cell-cycle phases using propidium iodide staining, and after oxidative stress using 0.5 µM hydrogen peroxide exposure for 0, 2, 5, 10, 30 and 60 min.
  
  
  RESULTS
  DSB levels were significantly increased in CD4+ T cells, CD8+ T cells and monocytes in SLE compared with healthy controls (p=2.16×10(-4), 1.68×10(-3) and 4.74×10(-3), respectively) and RA (p=1.05×10(-3), 1.78×10(-3) and 2.43×10(-2), respectively). This increase in DSBs in SLE was independent of the cell-cycle phase, and correlated with disease activity. In CD4+ T cells, CD8+ T cells and monocytes, oxidative stress exposure induced significantly higher DSB accumulation in SLE compared with healthy controls (60 min; p=1.64×10(-6), 8.11×10(-7) and 2.04×10(-3), respectively).
  
  
  CONCLUSIONS
  Our data indicate that SLE T cells and monocytes have increased baseline DSB levels and an increased sensitivity to acquiring DSBs in response to oxidative stress. Although the mechanism underlying DSB sensitivity in SLE requires further investigation, accumulation of DSB may serve a biomarker for disease activity in SLE and help explain increased apoptotic cell accumulation in this disease.
authors.affiliation: Division of Rheumatology, Department of Internal Medicine , University of Michigan , Ann Arbor, Michigan , USA.
authors.forename: Rajaie
authors.initials: R
authors.lastname: Namas
authors.affiliation: Division of Rheumatology, Department of Internal Medicine , University of Michigan , Ann Arbor, Michigan , USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Renauer
authors.affiliation: Division of Rheumatology, Department of Internal Medicine , University of Michigan , Ann Arbor, Michigan , USA.
authors.forename: Mikhail
authors.initials: M
authors.lastname: Ognenovski
authors.affiliation: Division of Rheumatology, Department of Internal Medicine , University of Michigan , Ann Arbor, Michigan , USA.
authors.forename: Pei-Suen
authors.initials: PS
authors.lastname: Tsou
authors.affiliation: Division of Rheumatology, Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan, USA; Center for Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, Michigan, USA.
authors.forename: Amr H
authors.initials: AH
authors.lastname: Sawalha
keywords: Double strand breaks; H2AX phosphorylation; Oxidative stress