Molecular Mechanism of Combination of Fumaric Acid and Cinnamic Aldehyde in Regulating in Oxidative Stress Induced by Enterotoxigenic Escherichia coli in Porcine Intestinal Epithelial Cells
This study was conducted to investigate the molecular mechanism by which the combination of fu-maric acid(FA)and cinnamic aldehyde(CA)modulated the oxidative stress induced by enterotoxigenic Esch-erichia coli(ETEC)K88 in porcine small intestinal epithelial cells IPEC-J2.IPEC-J2 cells were selected to es-tablish the oxidative damage model,and treated with different concentrations of FA and CA for 12 and 24 h.The cell viability was detected by CCK-8 method to determine the optimal treatment concentration and time.The cells were pretreated with optimal treatment concentrations of FA and CA,and ETEC K88 stimulated the cells for 3,6,12 and 24 h.The viable bacterial adhesion rate was detected,the cytokine contents and antioxi-dant indices were detected by enzyme-linked immunosorbent assay(ELISA),and the mRNA expression levels of heat shock protein 70(Hsp70)and genes related to nuclear-factor κB(NF-κB)signalling pathways by re-al-time fluorescence quantitative PCR.The results showed as follows:1)the optimal treatment concentrations of FA and CA were 1.00 mg/mL and 1.00 μL/mL,respectively,and the optimal incubation time was 12 h.2)The addition of FA and CA effectively inhibited the adhesion of ETEC K88 to IPEC-J2 cells,the adhesion rate of ETEC K88 was significantly decreased when it stimulated the cells for 3 h(P＜0.05),and extremely significantly decreased when it stimulated the cells for 6,12 and 24 h(P＜0.01).3)The addition of FA and CA extremely significantly decreased the contents of pro-inflammatory cytokines as interleukin-8(IL-8)and tumor necrosis factor-α(TNF-α)in ETEC K88-infected IPEC-J2 cells(P＜0.01),and extremely significantly increased the contents of anti-inflammatory cytokines as interleukin-10(IL-10)and transforming growth factor-β(TGF-β)(P＜0.01).4)ETEC K88 induced oxidative damage in IPEC-J2 cells by activating the NF-κB signalling pathway,and the addition of FA and CA up-regulated Hsp70 and inhibited the NF-κB signal-ling pathway to alleviate the oxidative stress in IPEC-J2 cells.5)The addition of FA and CA significantly in-creased the superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)activities in ETEC K88-infec-ted IPEC-J2 cells(P＜0.05),and extremely significantly reduced the malondialdehyde(MDA)content(P＜0.01).In conclusion,the combination of FA and CA can regulate the balance of inflammatory cytokines and oxidative stress proteins and inhibit the adhesion of ETEC K88 to IPEC-J2 cells,which may enhance the an-tioxidant and anti-inflammatory capacity of IPEC-J2 cells and alleviate the oxidative stress induced by ETEC K88 in IPEC-J2 cells by inhibiting the NF-κB signalling pathway through the up-regulation of Hsp70.[Chinese Journal of Animal Nutrition,2024,36(3):1904-1915]
cinnamic aldehydefumaric acidporcine small intestinal epithelial cellsenterotoxigenic Esche-richia colioxidative stress
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