In vitro effects of apigenin, curcumin, and fumaric acid on intracellular ROS and TNF-α responses in Escherichia coli-stimulated canine PBMCs.

BACKGROUND: Chronic inflammatory enteropathy (CIE) is a common gastrointestinal disorder in dogs and is associated with dysregulated inflammatory and oxidative responses, potentially contributing to the development of endotoxemia. METHODS: The present study investigated the effects of apigenin, curcumin, and fumaric acid on inflammatory and oxidative responses in canine peripheral blood mononuclear cells (PBMCs) under endotoxin- and bacteria-associated stimulation. PBMCs obtained from three healthy donor dogs were exposed to Escherichia coli O111:B4 or O127:B8 lipopolysaccharide (LPS), or to enrofloxacin-treated E. coli, and subsequently treated with the tested compounds. Cellular metabolic activity, LDH release, intracellular reactive oxygen species (ROS), H2O2 production, and TNF-α secretion were assessed. Minimum inhibitory concentration (MIC), mutant prevention concentration (MPC), and fractional inhibitory concentration (FIC) indices were determined to evaluate antimicrobial activity and interactions with enrofloxacin, amoxicillin-clavulanic acid, and gentamicin. RESULTS: None of the tested compounds significantly altered metabolic activity or LDH release under the applied conditions. Curcumin consistently reduced intracellular ROS and H2O2 production under selected stimulated conditions and decreased TNF-α secretion. Apigenin increased DCF-derived intracellular ROS under basal conditions and following lower-intensity O111 LPS stimulation, while reducing H2O2 and TNF-α production under several conditions, indicating a stimulus- and assay-dependent effect on cellular redox responses. Fumaric acid showed more limited, stimulus-dependent effects on oxidative and inflammatory parameters. Antimicrobial activity was also evaluated against four canine septicemic E. coli isolates. Due to limited solubility, the MIC and MPC values of the tested compounds were ≥1,024 μg/mL and could not be reliably determined above this concentration. Curcumin showed synergistic interaction with amoxicillin-clavulanic acid only against isolate No. 863, whereas fumaric acid exhibited concentration-dependent antagonism with enrofloxacin. CONCLUSION: Overall, the findings demonstrate compound- and stimulus-dependent modulation of oxidative and inflammatory responses in canine PBMCs and support further investigation of these compounds in disease-relevant models of endotoxemia and canine CIE.