A blend of botanicals protects cultured enterocytes from damages induced by an inflammatory challenge

Botanicals represent a wide class of complex compounds that naturally contain active principles with numerous biological functions. Among them, the anti-inflammatory and antioxidant actions are of key interest to support intestinal health during stressful phases. The aim of this study was to investigate the ability of a thymol-based blend of botanicals (BOT) to protect intestinal Caco-2 cells from the damages induced by an inflammatory challenge in vitro. Caco-2 cells were differentiated on porous inserts in basal medium, then treated with (CTR+) or without (CTR-) LPS+cytokines challenge, or LPS+cytokines challenge and BOT (BOT+). Transepithelial electrical resistance was daily measured for 7 d and, at the end, cells were harvested for gene expression analysis. Cells were also cultured on 96 well plates or glass coverslips, and treated for 24 h as previously described, to measure reactive oxygen species and perform immunofluorescence staining. The BOT treatment was effective in protecting enterocytes from inflammatory damages because transepithelial electrical resistance was significantly ameliorated in BOT+ group compared with CTR+ (P < 0.05). This outcome was supported by the increased levels of ZO-1, ZO-2, CLDN-1, CLDN-3, and OCCL observed when BOT was added to challenged Caco-2 cells. Moreover, immunofluorescence staining showed improved ZO-1 localization in BOT-treated cells. The BOT mechanism of action was related to a significant reduction in Caco-2 inflammatory response, as demonstrated by the decreased IL-6, IL-8 (P < 0.05), and TNFα (P < 0.1) levels. Furthermore, BOT improved the oxidative status of challenged enterocytes by significantly scavenging cellular reactive oxygen species. This effect was mediated by BOT ability to enhance the expression of antioxidant enzymes such as GPX-1 (P < 0.05) and numerically increase the mRNA levels of SOD. BOT was able to maintain epithelial integrity in vitro by helping enterocytes to face oxidative stress and inflammatory stimuli. BOT has the potential to be further investigated in vivo as a nutritional supplement to support animals’ transition through stressful phases.

Key Words: botanicals, inflammation, oxidative stress, intestinal integrity.