Mustard Meal Extract as an Alternative to Zinc Oxide for Protecting the Intestinal Barrier Against E. coli-Lipopolysaccharide Damage

The present study aimed to investigate the ability of an aqueous extract derived from mustard seed meal to counteract the effects of <i>E. coli</i> endotoxin lipopolysaccharide (LPS) on the intestinal epithelium. Caco-2 cells were cultured together with HT29-MTX and used as a cellular model to analyze critical intestinal parameters, such as renewal, integrity, innate immunity, and signaling pathway. Byproducts of mustard seed oil extraction are rich in soluble polysaccharides, proteins, allyl isothiocyanates, and phenolic acids, which are known as powerful antioxidants with antimicrobial and antifungal properties. Cells were seeded at a ratio of nine (Caco-2) to one (HT29-MXT) and treated for 2 h with mustard meal extract (ME, dilution 1/50) and zinc oxide (ZnO, 50 μM) after reaching 80-100% confluence. Then, they were challenged with 5 μg/mL <i>E. coli</i>-LPS and incubated for another 4 h. The results show that LPS did not alter the cell viability but decreased proliferation compared to the control, ME and ZnO treatments. LPS altered the cell membrane integrity and monolayer permeability by decreasing the transepithelial electrical resistance and tight-junction protein expression. In addition, LPS increased the activity of LDH and the expression of Toll-like receptors. The mechanisms by which LPS induces these disturbances involves the overexpression of PKC, p38 MAPK, and NF-κB signaling molecules. The pretreatment with mustard meal and ZnO succeeded in counteracting the impairment of epithelial renewal, the damage of the membrane integrity and permeability as well as in restoring the gene expression of tight-junction proteins.