S100A12在胸膜肺炎放线杆菌诱导猪肺泡巨噬细胞炎性细胞因子表达及吞噬中的作用

doi:10.16431/j.cnki.1671-7236.2021.11.036

Abstract

Abstract: The purpose of this study was to explore the role of S100A12 in the expression of inflammatory cytokines and phagocytosis in porcine alveolar macrophages (PAM) induced by Actinobacillus pleuropneumoniae (APP). After PAM cells were inoculated with APP strain, Real-time quantitative PCR was used to detect the mRNA expression levels of inflammatory cytokines and S100A12 at different time points after infection. S100A12 recombinant protein expression vector was constructed and purified, and its cytotoxicity to PAM was detected by MTT method. The effects of different concentrations (0, 5, 50 and 500 ng/mL) of S100A12 recombinant protein on the expression level of inflammatory cytokine mRNA in PAM were detected by Real-time quantitative PCR. S100A12 interference plasmid was constructed, and its interference efficiency and effects on inflammatory cytokine mRNA expression and apoptosis of PAM after APP infection were detected by Western blotting, Real-time quantitative PCR and flow cytometry. Plate counting method was used to detect the influence of S100A12 expression on APP adhesion and invasion and survival in PAM cells. The results showed that APP infection promoted the expression of IL-6, IL-8, IL-18, IL-1β and TNF-α as well as the expression of S100A12 in PAM, and the expression increased significantly or extremely significantly with the increase of APP infection dose and time (P<0.05;P<0.01). 5, 50 and 500 ng/mL recombinant protein S100A12 had no toxicity to PAM. The low concentration of S100A12 recombinant protein significantly promoted the expression of inflammatory cytokines IL-6, IL-8, TNF-α, IL-1β, IL-21 and IL-5 in PAM (P<0.05), while the high concentration of S100A12 recombinant protein had no effect on the expression of the above-mentioned inflammatory cytokines (P>0.05). The interference plasmid could successfully block the expression of S100A12 protein. Compared with control group transfected with shNC, the transcriptional level of the cytokines IL-6, IL-8, IL-1β and IL-5 in PAM transfected with shS100A12 interference plasmid under APP induction were significantly or extremely significantly reduced, and apoptosis rate was also significantly or extremely significantly increased (P<0.05;P<0.01). Further research found that interfering with the expression of S100A12 protein in PAM could significantly enhance the adhesion and invasion ability of APP to PAM and its survival ability in PAM (P<0.05). On the contrary, adding S100A12 recombinant protein in vitro significantly inhibited the adhesion, invasion and survival of APP to PAM cells (P<0.05). The above results showed that S100A12 could enhance the expression of inflammatory cytokines after APP infection in PAM cells, inhibit APP-induced PAM apoptosis, and reduce APP's adhesion and invasion to PAM and its survival in PAM cells, which laid a foundation for further revealing the regulatory effect and mechanism of S100A12 on PAM in the process of APP infection.

Key words: Actinobacillus pleuropneumoniae (APP); S100A12; porcine alveolar macrophages(PAM); cytokines

CLC Number:

S859.7

LIU Haiyao, LI Na, ZHANG Xiaoguang, LI Ziheng, BAO Chuntong, JIANG Hexiang, LIU Baijun, XIAO Jiameng, WANG Jun, LI Fengyang, LEI Liancheng. Role of S100A12 in the Expression of Inflammatory Cytokines and Phagocytosis in Porcine Alveolar Macrophages Induced by Actinobacillus pleuropneumoniae[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(11): 4262-4273.

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Role of S100A12 in the Expression of Inflammatory Cytokines and Phagocytosis in Porcine Alveolar Macrophages Induced by Actinobacillus pleuropneumoniae

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