Contents lists available at ScienceDirect Prostaglandins, Leukotrienes and Essential Fatty Acids journal homepage: www.elsevier.com/locate/plefa Protective effect of Rumenic acid rich cow's milk against colitis is associated with the activation of Nrf2 pathway in a murine model P. Bergamo a, ⁎ , E. Cocca b , A. Monaco c , V. Cozzolino a , F. Boscaino a , I. Ferrandino c , F. Maurano a , M. Rossi a a Institute of Food Sciences, National Research Council (CNR-ISA), Avellino, Italy b Institute of Biosciences and Bio-resources, National Research Council (CNR-IBBR), Naples, Italy c Department of Biology, University “Federico II” of Naples, Italy ARTICLE INFO Keywords: Experimental colitis Conjugated Linoleic Acid Fatty acids incorporation Intestinal inflammation Cow milk ABSTRACT Dietary supplementation with pure cis9, trans11 isomer of Conjugated Linoleic Acid –known as Rumenic Acid (RA)– improves cytoprotective defenses downstream through the activation of nuclear factor-E2-related factor- 2(Nrf2). This capability, when Rumenic Acid is consumed in the form of foods, is still unknown. The ability of standard (St) or cow milk naturally-enriched in RA (En) to activate Nrf2 pathway and its impact on dextran sodium sulfate (DSS)-induced colitis was comparatively evaluated. Activity of Nrf2 pathway was investigated in colonic tissue of BALB/c mice, receiving 4-week supplement with skimmed milk (SK), St or St reinforced with pure RA (RSt) providing increasing RA dose (0, 124 or 404 mg RA/kg -1 b.w, respectively). Next, the anti-oxidant/ anti-inflammatory effect produced by St or En treatment (383 mg RA/kg -1 b.w.) was explored. Finally, macroscopic and histomorphologic features of colitis were eval- uated in animals challenged with 5% (w/v) DSS, at the end of St or En treatment. Significant activation of Nrf2 pathway is associated with RSt and En intake (P < 0.05), but not with SK or En treatment. En pre-treatment offers better protection, in comparison with St, against pro-oxidant, pro-in- flammatory signs (P < 0.01) and macroscopic signs triggered by DSS. It can be concluded that Nrf2 activation by higher RA amount contained in En is, at least in part, responsible for the improved protection associated with En intake against DSS-induced colitis. 1. Introduction Colitis induced by dextran sodium sulfate (DSS) represents a useful experimental model, because it produces clinical signs observed in human pathology (diarrhea, bloody faeces, body weight loss, mucosal ulceration and shortening of the large intestine). DSS challenging is accompanied by an enhanced generation of Reactive Oxygen Species (ROS) [1], which induces inflammatory cascade and tissue damage [2]. The role played by redox status alteration in the etiology of DSS-in- duced colitis, is supported by the protective action of some antioxidant food components [3]. Moreover, additional data were obtained by using transgenic mice deficient in nuclear factor-E2-related factor 2 (Nrf2), which is a master regulator of a number of genes coding for antioxidant/detoxifying enzymes (e.g. glutathione S-reductase, GSR; glutathione S–transferase, GST; γ-glutamate cysteine ligase, GCL) (phase-2 enzymes) [3]. The cis9, trans11 isomer of Conjugated Linoleic acid (also known as Rumenic Acid, RA) is the most abundant dietary CLA predominantly found in milk and meat of ruminants, but it is also synthesized from trans-vaccenic acid (trans11-C18; TVA) through the action of liver de- saturases [4]. The beneficial effects of CLA intake (isomeric mixture of trans 10 , cis 12 CLA: RA, 1:1) were demonstrated [5,6], but it has been proved that RA is mainly accountable for anti-inflammatory and anti- oxidant properties [7]. Moreover, although animal supplementation with pure RA enhances antioxidant-detoxifying defenses (through the stimulation of the Nrf2 pathway) [8,9] the retention by RA of such capability when consumed in foods, is still not adequately supported by scientific evidence [10]. The fatty acid profile in milk is largely influenced by the dairy an- imal's diet; in particular, higher RA concentrations were found in milk http://dx.doi.org/10.1016/j.plefa.2017.08.006 Received 17 February 2017; Received in revised form 6 July 2017; Accepted 21 August 2017 ⁎ Correspondence to: Institute of Food Sciences, National Research Council (CNR-ISA), Via Roma 64, 83100 Avellino, Italy. E-mail address: p.bergamo@isa.cnr.it (P. Bergamo). Abbreviations: ALA, α-Linolenic Acid; CLA, Conjugated Linoleic Acid; DAI, Disease Activity Index; DSS, Dextran Sodium Sulfate; En, naturally-enriched RA milk; En+, En+DSS challenge; GCL, γ-Glutamate Cysteine Ligase; GCs, Goblet Cells; GSR, Glutathione-S-reductase; GST, Glutathione-S–transferase; LA, Linoleic Acid; Nrf2, Nuclear factor-E2-related factor 2; RA, Rumenic Acid (cis9, trans11-CLA); St, Standard milk; RSt, RA-reinforced St treatment; St+, St+DSS challenge; SK, Skimmed milk; t10, trans10, cis12-CLA; TVA, trans-vaccenic acid Prostaglandins, Leukotrienes and Essential Fatty Acids 125 (2017) 14–23 0952-3278/ © 2017 Elsevier Ltd. All rights reserved. MARK