P082 Assessment of anti-inflammatory effect of high acetate administration in UC patient-derived epithelial monolayer cultures

Abstract

Abstract Background Human organoid-based intestinal monolayer cultures provide an interesting tool to preclinically assess new therapeutic properties in inflammatory bowel disease (IBD). Recently, short-chain fatty acids (SCFA) such as acetate and butyrate have gained interest for their potential beneficial effects in IBD therapy. Most studies have focused on butyrate, given its beneficial effects on gut microbiome composition, intestinal barrier function and the immune system1. Effects of acetate are less well known, despite its lower toxicity to epithelial cells and its potential to support growth of butyrate-producing bacteria by metabolic cross-feeding2. In fact, a recent report suggested that the probiotic potential of Saccharomyces cerevisiae var. boulardii might be related to its high acetic acid production3. We here studied the effect of acetate on organoid-derived epithelial monolayer cultures from ulcerative colitis (UC) patients. Methods Colonic biopsies were obtained from non-inflamed regions in 3 patients with UC. Crypts were isolated, cultured and expanded as 3D-organoids. These organoids were then dissociated and transferred to transwell inserts as monolayer cultures. Upon confluency of the monolayer, evaluated by Transepithelial electrical resistance (TEER), cells were basolaterally stimulated with control medium (CTRL) or an inflammatory mix (INFL) containing 100 ng/ml TNF-α, 20 ng/ml IL-1β and 1 µg/ml Flagellin (Fig. 1). After 24h, cells were apically stimulated with control medium or high acetate (HA) (100mM). TEER was measured at 0, 24 and 48h stimulation. After 48h, cells were subjected to RNA extraction followed by reverse transcriptase qPCR targeting a selection of key diagnostic marker genes. The apical and basolateral media were collected for cytokine determination by Mesoscale (Proinflammatory panel 1). Data were analyzed using GraphPad Prism 9 (D’Agostino & Pearson test for normality followed by a Friedman test). Results A protective effect (Fig. 2) of high acetate administration on TEER values (HA vs INFL: p=0˒017) was observed, together with a trend towards decrease in IL8 (p=0˒11), TNFA (INFL vs INFL+HA: p=0˒68) and CLDN2 (CTRL vs HA: p=0˒03). Moreover, this was confirmed by a decrease of most proinflammatory cytokines (Fig. 3) in the apical and basolateral media upon HA stimulation e.g., IFN-γ (INFL vs INFL+HA: p=0˒25) and IL 10 (CTRL vs HA: p=0˒11). Conclusion In this patient-derived human epithelial cell culture model, a protective effect of high acetate administration on TEER-values, gene expression and cytokine production was observed. Ongoing experiments are expanding the number of patients to 10. References