Molecular and cellular pharmacology Anti-inflammatory activity of anatabine via inhibition of STAT3 phosphorylation Daniel Paris n , David Beaulieu-Abdelahad, Laila Abdullah, Corbin Bachmeier, Ghania Ait-Ghezala, Jon Reed, Megha Verma, Fiona Crawford, Michael Mullan The Roskamp Institute, 2040 Whitfield Avenue, Sarasota, Florida 34243, USA article info Article history: Received 2 April 2012 Received in revised form 31 October 2012 Accepted 1 November 2012 Available online 21 November 2012 Keywords: Anatabine Inflammation Cytokine STAT3 NFkB LPS Microglia Alzheimer abstract Previous investigations have demonstrated the anti-inflammatory effects of cholinergic agonists, such as nicotine. In the present study, we investigated the potential anti-inflammatory activity of anatabine, a minor tobacco alkaloid also present in plants of the Solanacea family which displays a chemical structural similarity with nicotine. Our data show that anatabine prevents STAT3 and NFkB phosphor- ylation induced by lipopolysaccharide (LPS) or TNF-a in SH-SY5Y, HEK293, human microglia and human blood mononuclear cells. Using human whole blood, we found that anatabine prevents IL-1b production induced by LPS. We assessed anatabine’s anti-inflammatory activity in vivo using an acute model of inflammation by challenging wild-type mice with LPS. We observed that anatabine reduces pro-inflammatory cytokine production (IL-6, IL-1b and TNF-a) in the plasma, kidney and spleen of the animals following the injection of LPS and concomitantly opposes STAT3 phosphorylation induced by LPS in the spleen and kidney. We also investigated the impact of anatabine on neuroinflammation using a transgenic mouse model of Alzheimer’s disease (Tg APPsw) that displays elevated cytokine levels in the brain. Following a chronic oral treatment with anatabine, a reduction in brain TNF-a and IL-6 levels compared to untreated Tg APPsw mice was observed. Moreover, an increased STAT3 phosphorylation was detected in the brains of Tg APPsw mice compared to wild-type littermates and was inhibited by anatabine treatment. Overall our data show that the anti-inflammatory activity of anatabine in vitro and in vivo is mediated in part via an inhibition of STAT3 phosphorylation. & 2012 Elsevier B.V. All rights reserved. 1. Introduction Inflammation is present in many pathological conditions ranging from cancer, obesity, immune disorders, type 2 diabetes, psoriasis, asthma, rheumatoid arthritis, gastrointestinal track disorders, neurodegenerative conditions as well as cardiovascular disorders (Brooks-Worrell and Palmer, 2012; Chen and Nunez, 2011; Dunn et al., 2012; Khandelwal et al., 2011; Schett et al., 2011; Zheng et al., 2011). The inflammatory system is also hyper- activated during sepsis leading to a massive increase in pro- inflammatory cytokines resulting in vascular leakage and impairment of the function of vital organs (Liu and Malik, 2006). Several studies have demonstrated the anti-inflammatory action of cholinergic agonists, such as nicotine, and identified a physiological mechanism that inhibits cytokines production which is referred to as ‘‘the cholinergic anti-inflammatory pathway’’ (Johnston and Webster, 2009; Pavlov and Tracey, 2005; Thayer, 2009). There is considerable evidence that nicotine can reduce peripheral inflammation and it is thought that the a7-nicotinic acetylcholine receptor subtype is a key mediator of this process (Ulloa, 2005). In addition, nicotine is also efficient in reducing central nervous system inflammation (Hao et al., 2011). Both acetylcholine (which is released by stimulation of the vagus nerve) or nicotine stimulate a7-nicotinic acetylcholine receptor and suppress TNFa production by LPS- stimulated macrophages/monocytes (de Jonge et al., 2005; Yoshikawa et al., 2006). To the contrary, vagotomy or a7-nicotinic acetylcholine receptor deficiency exacerbate sensitivity to endo- toxin and abolish the anti-inflammatory activity of nicotine (Li et al., 2010; Pavlov et al., 2009; Yoshikawa et al., 2006). Recent data have also highlighted that cholinergic modulation of inflam- mation is not solely dependent on the a7-nicotinic acetylcholine receptor but likely involves several nicotinic acetylcholine recep- tor subtypes (Hao et al., 2011; Hosur and Loring, 2011). Overall, the mechanisms responsible for the anti-inflammatory activity of cholinergic agonists such as nicotine are complex and appear to be mediated in part by a modulation of Signal Transducer and Activator of Transcription 3 (STAT3) and NFkB signaling pathways (de Jonge et al., 2005; Joe et al., 2011; Kox et al., 2009; Park et al., 2008; Yoshikawa et al., 2006). The transcription factors NFkB and STAT3 are key regulators of cytokine production and are also Contents lists available at SciVerse ScienceDirect journal homepage: www.elsevier.com/locate/ejphar European Journal of Pharmacology 0014-2999/$ - see front matter & 2012 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.ejphar.2012.11.017 n Corresponding author. Tel.: þ1 941 752 2949; fax: þ1 941 752 2948. E-mail address: dparis@rfdn.org (D. Paris). European Journal of Pharmacology 698 (2013) 145–153