Inflammatory mediators in a short-time mouse model of doxorubicin-induced cardiotoxicity Michela Pecoraro a , Mariagiovanna Del Pizzo a , Stefania Marzocco a , Rosalinda Sorrentino a , Michele Ciccarelli b , Guido Iaccarino b , Aldo Pinto a,1 , Ada Popolo a, ⁎ ,1 a Department of Pharmacy, University of Salerno, Fisciano, SA, Italy b Department of Medicine and Surgery, University of Salerno, Baronissi, SA, Italy abstract article info Article history: Received 18 August 2015 Revised 14 December 2015 Accepted 8 January 2016 Available online 11 January 2016 Doxorubicin (DOXO) is commonly used to treat a wide range of malignant tumors, but its clinical use is limited by acute and chronic cardiotoxicity. The precise mechanism underlying DOXO-induced cardiotoxicity is still not completely elucidated, but cardiac inflammation seems to be involved. Effects of DOXO on proinflammatory cy- tokines, inflammatory cell infiltration, and necrosis have been proven only when a functional impairment has al- ready occurred, so this study aimed to investigate the acute effect of DOXO administration in mouse heart. The results of our study demonstrated alterations in cardiac function parameters assessed by ultrasound within 24 h after a single injection of DOXO, with a cumulative effect along the increase of the dose and the number of DOXO administrations. At the same time, DOXO causes a significant production of proinflammatory cytokines (such as TNF-α and IL-6) with a concomitant reduction of IL-10, a well-known antiinflammatory cytokine. Fur- thermore, overexpression of inducible nitric oxide synthase (iNOS) in heart tissue and increased levels of serum nitrite in DOXO-treated mice were detected. Notably, DOXO administration significantly increased nitrotyrosine expression in mouse heart. Our data support the hypothesis that these early events, could be responsible for the later onset of more severe deleterious remodeling leading to DOXO induced cardiomyopathy. © 2016 Elsevier Inc. All rights reserved. Keywords: Doxorubicin Cardiotoxicity Inflammation Cytokines Nitric oxide 1. Introduction Doxorubicin (DOXO) is one of the most widely used anticancer drug (Riad et al., 2009), but its clinical application is hampered by cardiotoxicity. DOXO induces progressive myocardial stress, cardiomy- opathy and heart failure that can be fatal (Wong et al., 2013). Intensive investigations on DOXO-induced cardiotoxicity have con- tinued for decades. Different lines of evidences have provided putative mechanisms, nevertheless the precise mechanism underlying DOXO- induced cardiotoxicity is still not completely elucidated (Octavia et al., 2012). Traditionally, reactive oxygen species (ROS) have received more attention, although basic research has proven other factors (such as sarcomeric structure disruption, iron metabolism, energetic al- terations and inflammation) to be important as well (Salazar- Mendiguchía et al., 2014). Indeed, numerous studies showed that car- diac production of proinflammatory cytokines, inflammatory cell infil- tration, and necrosis are increased in hearts of chronic DOXO treated mouse (Ikegami et al., 2007; Li et al., 2006; Riad et al., 2009). To note, it is believed that the activation of the innate immune system with the ensuing proinflammatory cytokines release are at the basis of the pathogenesis of DOXO-induced cardiotoxicity (Hadi et al., 2012). In support to this hypothesis, it has been demonstrated that DOXO induces release of proinflammatory cytokines such as tumor necrosis factor (TNF)-α, via the activation of nuclear factor kappa-B (NF-κB), in the heart of rats treated with DOXO over a 2- weeks' timeframe (Abd El-Aziz et al., 2012). A relationship between TNF-α release and ROS increase in the failing hearts of patients with dilatative cardiomyopathy was found (Tsutamoto et al., 2001), and this could give an indication of the possible timeline that links in- flammation and oxidative stress. Inflammatory cytokines have been shown to participate in several heart diseases in that they affect heart rate to a negative inotropic effect and induce deleterious left ventricular remodeling (Zhu et al., 2009). In this scenario, higher expression of inducible nitric oxide synthase (iNOS) in cardiomyocytes is responsible of high levels of cytosolic nitric oxide (NO), which lead to proinflammatory mediators release by the innate immune cells (Takimoto et al., 2002). The most of studies report cardiotoxic effects of DOXO after chronic administration, however, a weakness in these studies is that structural cardiac damage is usually detected only when a functional impairment has already occurred, which leaves little room for early, preventive Toxicology and Applied Pharmacology 293 (2016) 44–52 ⁎ Corresponding author at: via Giovanni Paolo II 132, 84084 Fisciano, SA, Italy. E-mail address: apopolo@unisa.it (A. Popolo). 1 These authors equally contributed as last author. http://dx.doi.org/10.1016/j.taap.2016.01.006 0041-008X/© 2016 Elsevier Inc. All rights reserved. Contents lists available at ScienceDirect Toxicology and Applied Pharmacology journal homepage: www.elsevier.com/locate/ytaap