Transcriptional Modulation of the Nrf2 Antioxidant Pathway and Tight Junctions by a Plant Sterol Food Supplement in an Intestinal Inflammation Coculture Model

There is growing evidence supporting the anti-inflammatory activity of plant sterols (PS) in preclinical models of intestinal inflammation. However, the underlying mechanisms driving these beneficial effects remain incompletely understood. This study investigates the transcriptional modulatory activity of a PS food supplement (PS-FS) on genes associated with the cellular antioxidant defense system and tight junction (TJ) proteins. A double-chamber coculture model with differentiated Caco-2 cells (apical) and RAW264.7 macrophages (basolateral) was used. Bioaccessible fraction (BF) of PS-FS was obtained following the INFOGEST 2.0 simulated gastrointestinal digestion. A 1/20 dilution of BF was applied to the apical chamber for 90 min, followed by lipopolysaccharide (LPS) stimulation (1 µg/mL, 24 h) on the basolateral side. The interaction between PS-FS and budesonide was also evaluated. The gene expression of Nrf2 (NFE2L2), glutathione S-transferase A1 (GSTA1), NAD(P)H quinone dehydrogenase 1 (NQO1), and heme oxygenase 1 (HMOX-1) was quantified. Additionally, gene expression of TJ-related genes (CLDN1, CLDN3, CLDN4, OCLN, and TJP1) was quantified. Pretreatment with PS-FS significantly increased expression of GSTA1 (26.8%) and HMOX-1 (124.4%) compared to LPS + digestion blank. Furthermore, there was a trend toward increased expression of NFE2L2 and NQO1. Similarly, PS-FS enhanced expression of CLDN3 (33.0%), CLDN4 (211.7%), OCLN (57.1%), and TJP1 (98.9%). However, co-treatment of budesonide and PS-FS resulted in antagonistic interactions. These findings indicate that PS-FS modulates gene expression associated with antioxidant responses and intestinal barrier integrity-related pathways, suggesting to be further developed as a dietary strategy to support the management of inflammatory bowel disease. Practical Applications: This research supports the potential use of plant sterol-based food supplements in products aimed at supporting intestinal health. The results suggest that these supplements could help strengthen the gut barrier and support the body's natural antioxidant defenses during intestinal inflammation.