The International Journal of Biochemistry & Cell Biology 54 (2014) 122–136 Contents lists available at ScienceDirect The International Journal of Biochemistry & Cell Biology jo u r n al homep ag e: www.elsevier.com/locate/biocel Dual blockade of the A 1 and A 2A adenosine receptor prevents amyloid beta toxicity in neuroblastoma cells exposed to aluminum chloride Salvatore Giunta a , Violetta Andriolo b , Alessandro Castorina a,∗ a Department of Bio-Medical Sciences, Section of Anatomy and Histology, University of Catania, Italy b Department of Pediatrics and Public Health Sciences, University of Turin, Italy a r t i c l e i n f o Article history: Received 6 March 2014 Received in revised form 25 June 2014 Accepted 11 July 2014 Available online 21 July 2014 Keywords: Aluminum Neurotoxicity Adenosine receptor Caffein Alzheimer’s disease a b s t r a c t In a previous work we have shown that exposure to aluminum (Al) chloride (AlCl 3 ) enhanced the neuro- toxicity of the amyloid beta 25-35 fragment (Abeta 25-35 ) in neuroblastoma cells and affected the expression of Alzheimer’s disease (AD)-related genes. Caffein, a compound endowed with beneficial effects against AD, exerts neuroprotection primarily through its antagonist activity on A 2A adenosine receptors (A 2A R), although it also inhibits A 1 Rs with similar potency. Still, studies on the specific involvement of these receptors in neuroprotection in a model of combined neurotoxicity (Abeta 25-35 + AlCl 3 ) are missing. To address this issue, cultured SH-SY5Y cells exposed to Abeta 25-35 + AlCl 3 were assessed for cell viability, morphology, intracellular ROS activity and expression of apoptosis-, stress- and AD-related proteins. To define the role of A 1 R and A 2A Rs, pretreatment with caffein, specific receptor antagonists (DPCPX or SCH58261) or siRNA-mediated gene knockdown were delivered. Results indicate that AlCl 3 treatment exacerbated Abeta 25-35 toxicity, increased ROS production, lipid peroxidation, -secretase-1 (BACE1) and amyloid precursor protein (APP). Interestingly, SCH58261 successfully prevented toxicity associated to Abeta 25-35 only, whereas pretreatment with both DPCPX and SCH58261 was required to fully avert Abeta 25-35 + AlCl 3 -induced damage, suggesting that A 1 Rs might also be critically involved in protection during combined toxicity. The effects of caffein were mimicked by both N-acetyl cysteine, an antioxi- dant, and desferrioxamine, likely acting through distinct mechanisms. Altogether, our data establish a novel protective function associated with A 1 R inhibition in the setting of combined Abeta 25-35 + AlCl 3 neurotoxicity, and expand our current knowledge on the potential beneficial role of caffein to prevent AD progression in subjects environmentally exposed to aluminum. © 2014 Elsevier Ltd. All rights reserved. 1. Introduction Aluminum (Al) has been suggested as a potential neurotoxic metal implicated in the progression of a number of neurodegen- erative diseases, including Alzheimer’s disease (AD) (Gorell et al., 1999; Nakamura et al., 2000; Rondeau et al., 2000). Its causative relationship with AD remains to be fully outlined, although numer- ous evidences have proposed that Al might modify some of the pathogenetic mechanisms that contribute to the development of this neurodegenerative disease (Chen et al., 2011; Cuajungco et al., 2000; Flaten, 2001; Lovell et al., 1993; Lukiw and Pogue, 2007; Rondeau et al., 2009). ∗ Corresponding author at: Department of Bio-Medical Sciences, Section of Anatomy and Histology, University of Catania, Via S. Sofia, 87, 95123 Catania, Italy. Tel.: +39 095 3782042; fax: +39 095 3782046. E-mail address: alecasto@unict.it (A. Castorina). One of the main mechanisms of Al toxicity is the disruption of the intracellular redox environment and involves alterations in metal homeostasis (Becaria et al., 2006; Harris et al., 1996; Kawahara and Kato-Negishi, 2011). Imbalance in metal homeosta- sis seems to be caused by the similarity of the physical and chemical properties of Al with other metals, which enables Al to aberrantly mimic their biological functions to drive biochemical abnormali- ties. Specifically, Al-mediated alterations in iron (Fe) homeostasis have been indicated as pivotal factors that render this metal toxic (Peto, 2010; Ward et al., 2001; Wu et al., 2012). Indeed, the inter- action between Al and Fe contributes to the formation of reactive oxygen species (ROS). To support this, elevated levels of ROS have been shown in various models of Al toxicity (Bhasin et al., 2012; Lemire et al., 2011; Nayak et al., 2010; Yuan et al., 2012) and oxidative damage has been systematically observed in the brain and other organs of animals exposed to the metal (Li et al., 2012; Nehru and Anand, 2005; Praticò et al., 2002). Other evidences have suggested that Al can induce toxicity in primary human neural http://dx.doi.org/10.1016/j.biocel.2014.07.009 1357-2725/© 2014 Elsevier Ltd. All rights reserved.