Reuterin is well-known for its broad-spectrum antimicrobial ability, while the other potential bioactivity is not yet clear. The present study aims to investigate the immunomodulatory activity of reuterin on chicken macrophage HD11 cells for the first time and evaluate whether reuterin is able to regulate the lipopolysaccharide-stimulated inflammatory response. The results showed that the safe medication range of reuterin was less than 250 μM. Reuterin treatment for 6 h decreased the transcriptional ofCD86,IL-1βandiNOSand increased the expression ofCD206in a dose-dependent way, but reuterin treatment for 12 h contrary increased the expression ofIL-1β,IL-6andIL-10. However, it was noticed that reuterin treatment for 12 h significantly decreased the production of reactive oxygen species (ROS) and suppressed the phagocytosis activity of HD11 macrophages against bacteria. Further, the results showed that preincubation or coincubation with reuterin significantly attenuated the promotive effects of lipopolysaccharide (LPS) on transcription of proinflammatory cytokines (includingIL-1β,IL-6andTNF-α) and obviously inhibited nitric oxide (NO) production as well as the protein expression of inducible nitric oxide synthase (iNOS). Meanwhile, Mechanism studies implied that reuterin might exert an anti-inflammatory effect on LPS-stimulated cells by downregulating the expression ofTLR4/MyD88/TRAF6and blocking the activation of NF-κB as well as MAPKs signaling pathways. Additionally, it was found that both pretreatment and cotreatment with reuterin remarkably inhibited the oxidative stress induced by LPS stimulation by activating the Nrf2/HO-1 signaling pathway and enhancing the activities of antioxidative enzymes. These findings suggested the immunoregulatory function of reuterin and indicated this bacterial metabolite was able to inhibit the inflammation and oxidative stress of HD11 macrophages once exposed to LPS stimulation.Keywords:reuterin;HD11 cell line;lipopolysaccharide (LPS);inflammation response;oxidative stress
Reuterin, a potent antimicrobial compound derived from Limosilactobacillus reuteri , was investigated for its mechanism of action against Staphylococcus aureus biofilm and its efficacy in eliminating biofilms on four distinct milk-contact materials. Results showed that reuterin exhibited a minimum biofilm inhibitory concentration of 0.16 mg/mL against S. aureus ATCC 29213 after treatment for 48 h at 37 °C. Reuterin significantly reduced bacterial hydrophobicity, auto-aggregation and biofilm biomass by 39.36%, 25.49%, and 93.46%, respectively, effectively impeding biofilm formation and bacterial adhesion activity. For the mature biofilm, the minimum biofilm eradication concentration of reuterin incubated for 8 h at 37 °C was 0.31 mg/mL. At this concentration, reuterin significantly decreased the levels of extracellular proteins, polysaccharides and DNA of the biofilm as assayed by the BCA method, the phenol-sulfuric acid test and the spectrophotometric assay. Metabolic activity within the biofilm was reduced by 47.95% determined by the resazurin method, while the viable bacteria was not detected by plate counting. Laser confocal microscopy further confirmed the disruptive effects of reuterin on biofilm architecture. Moreover, RT-qPCR analysis showed that reuterin inhibited the relative expression of the accessory regulator ( sarA ) and bacterial adhesion factors ( clfA, clfB, fnbA, fnbB ). Additionally, reuterin was also found to interfere quorum-sensing genes within S. aureus , particularly targeting the LuxS/AI-2 system ( icaR, icaA, icaB, icaC, icaD ) and the agr system ( agrA, agrB, RNAIII ). Finally, reuterin eliminated biofilms formed on four milk-contact materials, including glass, stainless steel, polyvinyl chloride, and acrylonitrile-butadiene-styrene. Conclusively, reuterin emerges as a highly promising anti-biofilm agent in the food industry.
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