MOLECULAR MECHANISMS INVOLVED IN THE IN VITRO PROTECTIVE EFFECTS OF ANTHOCYANINS AGAINST INTESTINAL INFLAMMATION

The inflammatory bowel disease (IBD) is a group of multifactorial pathologies with an unknown etiology, characterized by an alternation of an acute and a remission phase of the intestinal epithelium inflammation. In the last years there was a worldwide increased incidence of IBD, especially in the industrialized and growing countries. In addition, epidemiological studies reported a positive correlation between gut inflammation and obesity. Recent in vivo and in vitro studies have supported the beneficial effects of anthocyanins, a class of flavonoid compounds widely distributed in Mediterranean diet, in various chronic inflammatory diseases, such as IBD, since they possess anti-inflammatory and antioxidant activity. In the first part of this study, we aimed to evaluate the molecular mechanisms involved in the modulation of intestinal epithelial inflammation by using an in vitro model consisting of Caco-2 cells exposed to high concentrations of palmitic acid (PA), and the protective effects exerted by cyanidin-3-O-glucoside (C3G) pre-treatment. For all the experiments, fully differentiated Caco-2 were pretreated for 24h with different concentration of C3G (10 and 20μM), added on the apical side, and then exposed to PA 100μM, added on the basolateral chamber, for 6h. The data obtained demonstrated the C3G anti-inflammatory activity through the modulation of NF-κB pathway induced by PA. In addition, C3G was able to improve the intracellular redox status through the activation of the adaptive cellular response modulated by Nrf2 pathway. Furthermore, it ameliorates the intestinal barrier by reducing the intestinal paracellular activity altered by PA. Since the bioavailability of the anthocyanins seems to be very low mainly due to poor stability during gastrointestinal digestion, in the second part of this study an in vitro simulated gastrointestinal digestion of a purified and standardized bilberry and blackcurrant extract (BBE), rich in anthocyanins, was performed. We further studied the bioactivity of the BBE, after the static simulated digestion, on an in vitro model of intestinal inflammation by using differentiated Caco-2 cells exposed to TNF-α. The outcomes confirmed the high instability of the anthocyanins in mild alkaline environment of the small intestine reporting a 13% of recovery index. However, although the high loss of anthocyanins, the digested BBE maintained part of its bioactivity, proved by the inhibition of the of NF-κB pathway induced by TNF-α, and by the activation of Nrf2 pathway. These data hence confirm that anthocyanins, introduced by diet or food supplements, could represent a possible approach for the prevention of IBD.