Contents lists available at ScienceDirect Aquatic Toxicology journal homepage: www.elsevier.com/locate/aqtox Sub-lethal doses of polybrominated diphenyl ethers affect some biomarkers involved in energy balance and cell cycle, via oxidative stress in the marine fish cell line SAF-1 Cristobal Espinosa Ruiz a , Simona Manuguerra a , Alberto Cuesta b , Maria Angeles Esteban b , Andrea Santulli a,c , Concetta M. Messina a, ⁎ a University of Palermo, Dept of Earth and Marine Science DISTEM, Laboratory of Marine Biochemistry and Ecotoxicology, Via Barlotta 4, 91100, Trapani, Italy b Fish Innate Immune System Group, Department of Cell Biology and Histology, Faculty of Biology, Campus Regional de Excelencia Internacional “Campus Mare Nostrum”, University of Murcia, 30100, Murcia, Spain c Consorzio Universitario della Provincia di Trapani, Marine Biology Institute, Via Barlotta 4, 91100, Trapani, Italy ARTICLE INFO Keywords: PBDEs Oxidative stress Energetic balance Cell cycle Sparus aurata fibroblast ABSTRACT Polybrominated diphenyl ethers (PBDEs) are a class of persistent contaminants which are found all over the world in the marine environment. Sparus aurata fibroblast cell line (SAF-1) was exposed to increasing con- centrations of PBDEs 47 and 99, until 72 h to evaluate the cytotoxicity, reactive oxygen species (ROS) production and the expression of some selected molecular markers related to cell cycle, cell signaling, energetic balance and oxidative stress (p53, erk-1, hif-1α and nrf-2), by real-time PCR. Furthermore, SAF-1 cells were exposed for 7 and 15 days to sub-lethal concentrations, in order to evaluate the response of some biomarkers by immunoblotting (p53, ERK-1, AMPK, HIF-1α and NRF-2). After 48 and 72 h, the cells showed a significant decrease of cell vitality as well as an increase of intracellular ROS production. Gene expression analysis showed that sub-lethal con- centrations of BDE-99 and 47, after 72 h, up-regulated cell cycle and oxidative stress biomarkers, although exposure to 100 μmol L −1 down-regulated the selected markers related to cell cycle, cell signaling, energetic balance. After 7 and 15 days of sub-lethal doses exposure, all the analyzed markers resulted affected by the contaminants. Our results suggest that PBDEs influence the cells homeostasis first of all via oxidative stress, reducing the cell response and defense capacity and affecting its energetic levels. This situation of stress and energy imbalance could represents a condition that, modifying some of the analyzed biochemical pathways, would predispose to cellular transformation. 1. Introduction Polybrominated diphenyl ethers (PBDEs) are a group of chemical compounds that are used in synthetic polymers and plastics with the aim of diminishing the flammability of combustible materials (Reynier et al., 2001; Teuten et al., 2009). These organobromide compounds are designed to be very stable for many years, which means they are able to remain in the environment for a considerable time (Eljarrat and Barceló, 2011). Many studies on PBDEs have shown that these com- pounds are ubiquitous, toxic, persistent and bioaccumulated in the environment (Boer et al., 2001; Hong et al., 2010; Horri et al., 2018; Hu et al., 2010; Kierkegaard et al., 2004; Kim and Stapleton, 2010; Oberg et al., 2002; Sellstrom and Jansson, 1995; Sjödin et al., 2001; Zhu and Hites, 2006). Unfortunately, they accumulate also in aquatic environ- ments by different ways, and the 2, 2′, 4, 4′-tetrabromodiphenyl ether (BDE-47) and the 2, 2′, 4, 4′, 5-pentabromodiphenyl ether (BDE-99) are among the most abundant found in this ambience (Bi et al., 2007; Leung et al., 2006). Because of these properties, some flame retardants have been prohibited in the USA and European Union (ATSDR, 2004; Directive 76/769/EEC, 1986; Off. J. Eur. Union, 2003; World Health Organization, 2003). In the organism, PBDEs may be metabolized to more polar com- pounds, such as hydroxylated, methoxylated and/or conjugated meta- bolites (OH-, MeO- and GS-PBDEs), via phase I and phase II metabolic https://doi.org/10.1016/j.aquatox.2019.02.014 Received 16 December 2018; Received in revised form 18 February 2019; Accepted 18 February 2019 Abbreviations: AREs, antioxidant response elements; AMPK, adenosine 5′-monophosphate-activated protein kinase; CYP, cytochrome P450; ERK-1, extracellular signal–regulated kinase 1; HIF-1, hypoxia inducible factor 1; NRF-2, nuclear factor (erythroid-derived 2)-like 2; PBDEs, polybrominated diphenyl ethers; ROS, reactive oxygen species ⁎ Corresponding author. E-mail address: concetta.messina@unipa.it (C.M. Messina). Aquatic Toxicology 210 (2019) 1–10 Available online 19 February 2019 0166-445X/ © 2019 Elsevier B.V. All rights reserved. T