title: Analysis of Lymphocytic DNA Damage in Early Multiple Sclerosis by Automated Gamma-H2AX and 53BP1 Foci Detection: A Case Control Study.
abstract: BACKGROUND
  In response to DNA double-strand breaks, the histone protein H2AX becomes phosphorylated at its C-terminal serine 139 residue, referred to as γ-H2AX. Formation of γ-H2AX foci is associated with recruitment of p53-binding protein 1 (53BP1), a regulator of the cellular response to DNA double-strand breaks. γ-H2AX expression in peripheral blood mononuclear cells (PBMCs) was recently proposed as a diagnostic and disease activity marker for multiple sclerosis (MS).
  
  
  OBJECTIVE
  To evaluate the significance of γ-H2AX and 53BP1 foci in PBMCs as diagnostic and disease activity markers in patients with clinically isolated syndrome (CIS) and early relapsing-remitting MS (RRMS) using automated γ-H2AX and 53BP1 foci detection.
  
  
  METHODS
  Immunocytochemistry was performed on freshly isolated PBMCs of patients with CIS/early RRMS (n = 25) and healthy controls (n = 27) with γ-H2AX and 53BP1 specific antibodies. Nuclear γ-H2AX and 53BP1 foci were determined using a fully automated reading system, assessing the numbers of γ-H2AX and 53BP1 foci per total number of cells and the percentage of cells with foci. Patients underwent contrast enhanced 3 Tesla magnetic resonance imaging (MRI) and clinical examination including expanded disability status scale (EDSS) score. γ-H2AX and 53BP1 were also compared in previously frozen PBMCs of each 10 CIS/early RRMS patients with and without contrast enhancing lesions (CEL) and 10 healthy controls.
  
  
  RESULTS
  The median (range) number of γ-H2AX (0.04 [0-0.5]) and 53BP1 (0.005 [0-0.2]) foci per cell in freshly isolated PBMCs across all study participants was low and similar to previously reported values of healthy individuals. For both, γ-H2AX and 53BP1, the cellular focus number as well as the percentage of positive cells did not differ between patients with CIS/RRMS and healthy controls. γ-H2AX and 53BP1 levels neither correlated with number nor volume of T2-weighted lesions on MRI, nor with the EDSS. Although γ-H2AX, but not 53BP1, levels were higher in previously frozen PBMCs of patients with than without CEL, γ-H2AX values of both groups overlapped and γ-H2AX did not correlate with the number or volume of CEL.
  
  
  CONCLUSIONS
  γ-H2AX and 53BP1 foci do not seem to be promising diagnostic or disease activity biomarkers in patients with early MS. Lymphocytic DNA double-strand breaks are unlikely to play a major role in the pathophysiology of MS.
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Ludwig
authors.initials: L
authors.lastname: Rasche
authors.affiliation: Medipan GmbH, Berlin-Dahlewitz, Germany.
authors.forename: Lisa
authors.initials: L
authors.lastname: Heiserich
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Janina Ruth
authors.initials: JR
authors.lastname: Behrens
authors.affiliation: Department of Medical Biometrics and Clinical Epidemiology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Klaus
authors.initials: K
authors.lastname: Lenz
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Catherina
authors.initials: C
authors.lastname: Pfuhl
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Katharina
authors.initials: K
authors.lastname: Wakonig
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: René Markus
authors.initials: RM
authors.lastname: Gieß
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Erik
authors.initials: E
authors.lastname: Freitag
authors.affiliation: Medipan GmbH, Berlin-Dahlewitz, Germany.
authors.forename: Caroline
authors.initials: C
authors.lastname: Eberle
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|MIAC AG, Basel, Switzerland.|Experimental and Clinical Research Center, Charité - Universitätsmedizin Berlin and Max Delbrück Center for Molecular Medicine, Berlin-Buch, Germany.|Institute of Neuroradiology, Universitätsmedizin Göttingen, Göttingen, Germany.
authors.forename: Jens
authors.initials: J
authors.lastname: Wuerfel
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Dörr
authors.affiliation: Medipan GmbH, Berlin-Dahlewitz, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Bauer
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Experimental and Clinical Research Center, Charité - Universitätsmedizin Berlin and Max Delbrück Center for Molecular Medicine, Berlin-Buch, Germany.
authors.forename: Judith
authors.initials: J
authors.lastname: Bellmann-Strobl
authors.affiliation: NeuroCure Clinical Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Friedemann
authors.initials: F
authors.lastname: Paul
authors.affiliation: Medipan GmbH, Berlin-Dahlewitz, Germany.|Faculty of Science, Brandenburg University of Technology Cottbus - Senftenberg, Senftenberg, Germany.
authors.forename: Dirk
authors.initials: D
authors.lastname: Roggenbuck
authors.affiliation: Clinical and Experimental Multiple Sclerosis Research Center, Charité - Universitätsmedizin Berlin, Berlin, Germany.|Department of Neurology, Charité - Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Klemens
authors.initials: K
authors.lastname: Ruprecht
mesh_terms: D000328:Adult; D015415:Biomarkers / Q000378:metabolism; D016022:Case-Control Studies; D004249:DNA Damage; D005260:Female; D006657:Histones / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D008214:Lymphocytes / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D009103:Multiple Sclerosis / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D000071857:Tumor Suppressor p53-Binding Protein 1; D055815:Young Adult
chemical_list: D015415:Biomarkers; C468783:H2AX protein, human; D006657:Histones; D047908:Intracellular Signaling Peptides and Proteins; C498301:TP53BP1 protein, human; D000071857:Tumor Suppressor p53-Binding Protein 1
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