Luteolin: A Phytochemical to Mitigate S. Typhimurium Flagellin-Induced Inflammation in a Chicken In Vitro Hepatic Model

SIMPLE SUMMARY: The inadequate use of antibiotics has resulted in the emergence of resistant microbes which imply a major threat in human and veterinary medicine. Therefore, natural alternatives improving the productivity of farm animals should be investigated to replace the extensive antibiotic application. The primary goal of the study was to prove the anti-inflammatory and antioxidant activity of luteolin (a common phytochemical of vegetables with a flavonoid structure) in a chicken hepatic cell culture. To investigate the effect of luteolin, a model was established which could recapitulate the Salmonella enterica serovar Typhimurium-induced hepatic inflammation of chickens. The inflammatory response was triggered with the elementary unit of the organ of bacterial motility, flagellin, and freshly isolated, primary hepatic cell cultures were applied containing both hepatocytes, the functional cells of the liver, and non-parenchymal, inflammatory cells. Luteolin at a concentration of 4 µg/mL did not alter the viability and the membrane integrity of the cells and therefore proved applicable to counteract flagellin. In combination with flagellin exposure, luteolin reduced the elevated IL-8 release of the cultured cells. Moreover, it reduced the concentration of IFN-α, H(2)O(2) and malondialdehyde and restored the level of IL-10 and the ratio of IFN-γ/IL-10. In conclusion, luteolin had an anti-inflammatory and antioxidant effect in a chicken hepatic cell culture mimicking Salmonella enterica-associated inflammation. ABSTRACT: The use of natural feed supplements is an alternative tool to diminish the damage caused by certain bacteria, improving animal health and productivity. The present research aimed to investigate the proinflammatory effect of flagellin released from the bacterial flagellum of Salmonella enterica serovar Typhimurium and to attenuate the induced inflammation with luteolin as a plant-derived flavonoid on a chicken primary hepatocyte–non-parenchymal cell co-culture. Cells were cultured in a medium supplemented with 250 ng/mL flagellin and 4 or 16 µg/mL luteolin for 24 h. Cellular metabolic activity, lactate dehydrogenase (LDH) activity, interleukin-6, 8, 10 (IL-6, IL-8, IL-10), interferon-α, γ (IFN-α, IFN-γ), hydrogen peroxide (H(2)O(2)) and malondialdehyde (MDA) concentrations were determined. Flagellin significantly increased the concentration of the proinflammatory cytokine IL-8 and the ratio of IFN-γ/IL-10, while it decreased the level of IL-10, indicating that the model served adequate to study inflammation in vitro. Luteolin treatment at 4 µg/mL did not prove to be cytotoxic, as reflected by metabolic activity and extracellular LDH activity, and significantly reduced the flagellin-triggered IL-8 release of the cultured cells. Further, it had a diminishing effect on the concentration of IFN-α, H(2)O(2) and MDA and restored the level of IL-10 and the ratio of IFN-γ/IL-10 when applied in combination with flagellin. These results suggest that luteolin at lower concentrations may protect hepatic cells from an excessive inflammatory response and act as an antioxidant to attenuate oxidative damage.