Molecular responses to clove and oregano essential oils are associated with reduced inflammation and improved gut barrier function in broiler chickens

In vitro tests were conducted to characterize the host-mediated responses of chickens to Clove Essential Oil (CEO) and Oregano Essential Oil (OEO). Chicken macrophage cells (CMCs), chicken intestinal epithelial cells (IECs), quail muscle cells (QMCs), and chicken embryonic muscle cells (EMCs) were utilized in these assays. EMCs were collected from the 13-day-old embryo during egg incubation and all cell lines were seeded at 2 × 10⁵/mL in a 24-well plate. In CMCs, an inflammatory response was induced by stimulating with 1.0 µg/mL of Lipopolysaccharide (LPS). To induce muscle cell differentiation, 0.5 % FBS was used in QMCs and 2.0 % FBS in EMCs. Three different concentrations (1.0, 10.0, and 100 µg/mL) of CEO and OEO were administered. qRT-PCR was used to measure gene expression levels of IL-1β and IL-8 from CMCs, occludin, ZO-1, and MUC2 from IECs, and Pax7 and MyoG from QMCs and EMCs. Cytotoxic effects of CEO and OEO were determined using an MTT assay; CEO and OEO did not show cytotoxicity at concentrations below 0.1 mg/mL in CMCs, IECs, QMCs, and EMCs. CEO reduced (P < 0.05) the LPS-induced increase of IL-1β and IL-8 in CMCs and increased (P < 0.05) ZO-1 and MUC2 in IECs. OEO suppressed (P < 0.05) the release of IL-8, increased ZO-1, and Pax7. Both CEO and OEO demonstrated microbicidal activity against sporozoite of E. tenella and C. perfringens bacteria, but only at doses 10-100 × higher than those that would be used in feed. These findings support our previous findings on other phytochemicals; both CEO and OEO are promising candidates for improved resilience in chickens not due to their direct antimicrobial effects, but due to gut physiological responses that take place at the level of the host.