Int. J. Devl Neuroscience 43 (2015) 8–15 Contents lists available at ScienceDirect International Journal of Developmental Neuroscience j ourna l ho me pa g e: www.elsevier.com/locate/ijdevneu Protective effect of antioxidants on DNA damage in leukocytes from X-linked adrenoleukodystrophy patients Desirèe P. Marchetti a,∗ , Bruna Donida b , Helen T. da Rosa c , Paula R. Manini c , Dinara J. Moura c , Jenifer Saffi c , Marion Deon b,d , Caroline P. Mescka a , Daniella M. Coelho d , Laura B. Jardim d,e , Carmen R. Vargas a,b,d,∗ a Programa de Pós-Graduac¸ ão em Ciências Biológicas, Bioquímica, UFRGS, Rua Ramiro Barcelos, 2600, CEP 90035-003 Porto Alegre, RS, Brazil b Programa de Pós-Graduac¸ ão em Ciências Farmacêuticas, UFRGS, Av. Ipiranga, 2752, CEP 90610-000 Porto Alegre, RS, Brazil c Universidade Federal de Ciências de Saúde de Porto Alegre, UFSPA, Rua Sarmento Leite, 245, CEP 90050170 Porto Alegre, RS, Brazil d Servic¸o de Genética Médica, HCPA, Rua Ramiro Barcelos, 2350, CEP 90035-003 Porto Alegre, RS, Brazil e Programa de Pós-Graduac¸ ão em Ciências Médicas, UFRGS, Rua Ramiro Barcelos, 2400, CEP 90035-003 Porto Alegre, RS, Brazil a r t i c l e i n f o Article history: Received 10 January 2015 Received in revised form 6 March 2015 Accepted 6 March 2015 Available online 10 March 2015 Keywords: X-linked adrenoleukodystrophy N-Acetyl-l-cysteine Trolox Rosuvastatin DNA damage a b s t r a c t Toxic metabolites accumulation and oxidative stress have been associated to the pathophysiology of X-linked adrenoleukodystrophy (X-ALD), an inborn error of peroxisome metabolism. Parameters of oxidative damage to proteins and lipids in X-ALD patients were already described in literature; however, DNA injuries were not studied yet. Considering that, the aims were to investigate DNA damage by comet assay in heterozygotes and symptomatic X-ALD patients, to look for associations between DNA damage and lipid peroxidation as measured by urinary 15-F2t-isoprostane; and to evaluate the in vitro effect of N-acetyl-l-cysteine (NAC), trolox (TRO) and rosuvastatin (RSV) on DNA damage in leukocytes from symptomatic patients. Symptomatic patients presented higher DNA damage levels than those found in heterozygotes and controls; heterozygotes and controls showed similar results. In order to investigate the in vitro antioxidant effect on DNA damage, whole blood cells from symptomatic patients were incubated with NAC (1 and 2.5 mM), TRO (25 and 75 M) and RSV (0.5, 2 and 5 M) before DNA damage analysis. NAC, TRO and RSV, at all tested concentrations, were all capable to reduce DNA damage in symptomatic X-ALD patients until control levels. Finally, DNA damage correlated with urinary isoprostanes and plas- matic levels of TBA-RS and DCFH-DA, allowing to hypothesize that DNA damage might be induced by lipid peroxidation in symptomatic patients. The present work yields experimental evidence that NAC, TRO and RSV reduce the in vitro DNA injury in symptomatic X-ALD patients, what may suggest that the administration of these antioxidants might be considered as an adjuvant therapy for X-ALD. © 2015 Elsevier Ltd. All rights reserved. Abbreviations: AMN, adrenomieloneuropathy; C22:0, docosanoic acid; C24:0, tetracosanoic acid; C26:0, hexacosanoic acid; CCER, childhood cerebral form; DCFH- DA, dihydrodichlorofluorescein diacetate; IEM, inborn error of metabolism; NAC, N-acetyl-l-cysteine; PBS, phosfate buffer saline; RSV, rosuvastatin; TAR, total antiox- idant reactive; TAS, total antioxidant status; TBA-RS, thiobarbituric acid-reactive substances; TRO, trolox; VLCFA, very long chain fatty acids; X-ALD, X-linked adrenoleukodystrophy. ∗ Corresponding authors. Tel.: +55 51 33598011; fax: +55 51 33598010. E-mail addresses: desireepmarchetti@gmail.com (D.P. Marchetti), donida.bruna@gmail.com (B. Donida), htdarosa@gmail.com (H.T. da Rosa), maninipaula@gmail.com (P.R. Manini), dinjamoura@gmail.com (D.J. Moura), jenifer.saffi@gmail.com (J. Saffi), marion deon@yahoo.com.br (M. Deon), carolmescka@yahoo.com.br (C.P. Mescka), dcoelho@hcpa.ufrgs.br (D.M. Coelho), ljardim@hcpa.ufrgs.br (L.B. Jardim), crvargas@hcpa.ufrgs.br (C.R. Vargas). 1. Introduction X-linked adrenoleukodystrophy (X-ALD) is the most frequent inherited peroxisomal disease, with an estimated incidence of 1:21,000 for hemizygotes men and of 1:14,000 for heterozygotes women (Bezman et al., 2001). This disorder is caused by mutations in ABCD1 gene (located at chromosome Xq28) which encodes the peroxisomal ATP-binding cassette (ABC) transporter, ABCD1 pro- tein (formerly adrenoleukodystrophy protein, ALDP) responsible for transporting very long chain fatty acids (VLCFA) into the perox- isomes for degradation by -oxidation (Moser, 1997; Moser et al., 2001, 2005). This inborn error of metabolism (IEM) is characterized by progressive demyelination of the white matter, adrenal insuffi- ciency and VLCFA accumulation, mainly hexacosanoic (C26:0) and http://dx.doi.org/10.1016/j.ijdevneu.2015.03.004 0736-5748/© 2015 Elsevier Ltd. All rights reserved.