This experiment was conducted to reveal the mitigating effect of dulcitol(DUL)on lipopolysaccha-ride(LPS)induced cellular barrier and inflammatory injury of porcine small intestinal epithelial cells(IPEC-J2 cells)and its possible molecular mechanism by establishing a IPEC-J2 cells damage model after stimulated with LPS.The IPEC-J2 cell viability was detected by the cell count kit-8(CCK-8),to determine the c LPS model-ing concentration(150 μg/mL)and the appropriate DUL treatment concentration(200 μmol/L).Three groups were setup in the experimental group:control(CON)group,LPS group and DUL group.After IPEC-J2 cells adhesion,the CON group was firstly treated with complete culture medium for 24 hours,then treated with DMEM medium for 24 hours;the LPS group firstly treated with complete culture medium for 24 hours,then treated with DMEM medium which contained 150 μg/mL LPS for 24 hours;the DUL group firstly treated with complete culture medium which contained 200 μmol/L DUL for 24 hours,then treated with DMEM me-dium which contained 150 µg/mL LPS for 24 hours.The IPEC-J2 cells growth state in each group was ob-served,the migration and proliferation ability of the cells was detected by wound healing assay,and the mRNA and protein relative expression levels of the barrier and inflammatory related genes was detected by quantitative real-time PCR(RT-qPCR)and Western blot.The results showed as follows:1)in IPEC-J2 cells,the appro-priate treatment concentration of DUL was 200 μmol/L,and the effective inflammatory concentration of LPS was 150 µg/mL.2)Compared with the LPS group,the DUL pretreatment alleviated the decreased cell densi-ty,increased cell vacuoles and poorly defined cell edges.3)Compared with the LPS group,the DUL pretreat-ment significantly increased the cell migration distance and cell migration area(P＜0.05).4)Compared with the LPS group,the DUL pretreatment significantly increased the mRNA and protein relative expression levels of the barrier-associated genes zonula occludens-1(ZO-1),occludin(OCLN)and claudin-1(CLDN1)and the mRNA relative expression levels of mucin 2(MUC2),large tumor suppressor kinase 1(LATS1)and YES-associated protein 1(YAPI)(P＜0.05).5)Compared with the LPS group,the DUL pretreatment signif-icantly decreased the mRNA and protein relative expression levels of myeloid differentiation factor 88(Myd88),inhibitor protein-α of nuclear factor-κB(IκB-α),nuclear factor-κB(NF-κB)and interleukin-1β(IL-1β)and protein relative expression level of Toll-like receptor 4(TLR4)and mRNA relative expression levels of nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3)and interleukin-18(IL-18)(P＜0.05).It is concluded that DUL regulates the secretion of inflammatory factors and enhances the ex-pression of tight junction-related genes by down-regulating the expression of proteins related to the TLR4/Myd88/NF-KB signaling pathway,and to alleviate the LPS induced IPEC-J2 cell barrier damage and inflam-matory response.[Chinese Journal of Animal Nutrition,2024,36(6):3952-3962]
万方数据
维普
