TDK conference 2024

In our society with an increasing demand for meat, improving the productivity of the swine industry is essential. However, the spread of antimicrobial resistance poses a global public and animal health challenge. Due to the risk of developing resistant bacterial strains, it is crucial to seek alternative agents to replace antibiotics. Our aim was to develop cell models suitable for investigating antimicrobial peptides (AMPs), with a focus on their anti-inflammatory and immunomodulatory properties, and to help mitigate enteric diseases of complex etiology in swine farming.

We established primary porcine liver cell and small intestinal epithelial explant cultures to study the gut-liver axis, which plays a key role in systemic inflammatory processes. We selected for treatment pro-inflammatory molecules that are known to cause significant production losses in the swine industry. The cell cultures were treated with lipopolysaccharide (LPS) from Escherichia coli and lipoteichoic acid (LTA) from Staphylococcus aureus (both at 10 µg/ml and 50 µg/ml), flagellin from Salmonella Typhimurium (100 ng/ml and 250 ng/ml), and the viral nucleoside analog polyinosinic-polycytidylic acid (poly I:C) (50 µg/ml and 100 µg/ml). To assess the viability of the cell cultures, the CCK-8 test was applied and the extracellular lactate dehydrogenase (LDH) activity was measured. For monitoring redox homeostasis, Amplex Red assay for hydrogen peroxide measurement was used and malondialdehyde concentrations were also determined. The concentrations of inflammatory cytokines were measured using multiplex Luminex xMAP technology.

According to the viability assays, metabolic changes and LDH activity were more pronounced in the liver cell cultures after treatment with pro-inflammatory molecules compared to the small intestinal epithelial explants. In terms of redox homeostasis, hydrogen peroxide levels increased in the liver cell cultures after all pro-inflammatory treatments. Among the inflammatory cytokines, the concentrations of IL-4, IL-6, IL-8, and TNF-α significantly increased in the hepatic cell cultures compared to the control group after LPS and LTA treatments, while poly I:C significantly elevated IL-6 levels. In the small intestinal epithelial explants, IL-6 concentrations significantly increased after flagellin and poly I:C treatments, while IL-8 concentrations markedly rose following LPS treatment. Based on these findings, we identified pro-inflammatory agents that caused significant increases in certain inflammatory cytokines in both porcine intestinal epithelial and liver cell cultures. Thus, the model system we developed is highly promising for studying the anti-inflammatory and immunomodulatory effects of antimicrobial peptides.