Derangement of intestinal epithelial cell monolayer by dietary cholesterol oxidation products

The emerging role of the diet in the incidence of intestinal inflammatory diseases has stimulated research on the influence of eating habits with pro-inflammatory properties in inducing epithelial barrier disturbance. Cholesterol oxidation products, namely oxysterols, have been shown to promote and sustain oxidative/inflammatory reactions in human digestive tract. This work investigated in an in vitro model the potential ability of a combination of dietary oxysterols representative of a hyper-cholesterol diet to induce the loss of intestinal epithelial layer integrity. The components of the experimental mixture were the main oxysterols stemming from heat-induced cholesterol auto-oxidation, namely 7-ketocholesterol, 5α,6α-and 5β,6β-epoxycholesterol, 7α- and 7β-hydroxycholesterol. These compounds added to monolayers of differentiated CaCo-2 cells in combination or singularly, caused a time-dependent induction of matrix metalloproteinases (MMP)-2 and -9, also known as gelatinases. The hyperactivation of MMP-2 and -9 was found to be associated with decreased levels of the tight junctions zonula occludens-1 (ZO-1), occludin and Junction Adhesion Molecule-A (JAM-A). Together with such a protein loss, particularly evident for ZO-1, a net perturbation of spatial localization of the three tight junctions was observed. Cell monolayer pre-treatment with the selective inhibitor of MMPs ARP100 or polyphenol (-)-epicathechin, previously shown to inhibit NADPH oxidase in the same model system, demonstrated that the decrease of the three tight junction proteins was mainly a consequence of MMPs induction, which was in turn dependent on the pro-oxidant property of the oxysterols investigated. Although further investigation on oxysterols intestinal layer damage mechanism is to be carried on, the consequent - but incomplete - prevention of oxysterols-dependent TJs alteration due to MMPs inhibition, avoided the loss of scaffold protein ZO-1, with possible significant recovery of intestinal monolayer integrity.