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Sheep β-Defensin 2 Regulates Escherichia coli F17 Resistance via NF-κB and MAPK Signaling Pathways in Ovine Intestinal Epithelial Cells.

ABSTRACT: Escherichia coli (E. coli) F17 is a member of enterotoxigenic Escherichia coli, which can cause massive diarrhea and high mortality in newborn lambs. β-defensin is mainly produced by the epithelial tissue of the gastrointestinal tract in response to microbial infection. However, the molecular mechanism of sheep β-defensin 2 (SBD-2) against E. coli F17 remains unclear. This study aims to reveal the antibacterial ability of SBD-2 against E. coli F17 infection in sheep. Firstly, we established the culture system of ovine intestinal epithelial cells (OIECs) in vitro, treated with different concentrations of E. coli F17 for an indicated time. Secondly, we performed RNA interference and overexpression to investigate the effect of SBD-2 expression on E. coli F17 adhesion to OIECs. Finally, inhibitors of NF-κB and MAPK pathways were pre-treated to explore the possible relationship involving in E. coli F17 infection regulating SBD-2 expression. The results showed that E. coli F17 markedly (p < 0.01) upregulated the expression levels of SBD-2 mRNA and protein in a concentration- and time-dependent manner. Overexpression of SBD-2 contributed to enhancing E. coli F17 resistance in OIECs, while silencing SBD-2 dramatically improved the adhesion of E. coli F17 to OIECs (p < 0.05 or p < 0.01). Furthermore, E. coli F17 stimulated SBD-2 expression was obviously decreased by pre-treatment with NF-κB inhibitor PDTC, p38 MAPK inhibitor SB202190 and ERK1/2 MAPK inhibitor PD98095 (p < 0.05 or p < 0.01). Interestingly, adhesion of E. coli F17 to OIECs were highly enhanced by pre-treated with PDTC, SB202190 and PD98095. Our data suggested that SBD-2 could inhibit E. coli F17 infection in OIECs, possibly through NF-κB and MAPK signaling pathways. Our results provide useful theoretical basis on developing anti-infective drug and breeding for E. coli diarrhea disease-resistant sheep.

SUBMITTER: Ge L

PROVIDER: S-EPMC8698448 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Diarrhea is one of the most commonly reported diseases in young farm animals. Escherichia coli (E. coli) F17 is one of the major pathogenic bacteria responsible for diarrhea. However, the pathogenicity of diarrhea in lambs involving E. coli F17 strains and how E. coli F17 infection modifies lambs' intestinal microbiota are largely unknown. To evaluate diarrhea in newborn lambs with an infection of E. coli F17, 50 lambs were selected for challenge experiments and divided into four groups, namely, a high-dose challenge group, low-dose challenge group, positive control group, and negative control group. The E. coli F17 challenge experiments caused diarrhea and increased mortality in the experimental lamb population, with a higher prevalence (90%), mortality (35%), and rapid onset time (4-12 h) being observed in the high-dose challenge group than the results observed in the low-dose challenge group (75%, 10%, 6-24 h, respectively). After the challenge experiment, healthy lambs in the high-dose challenge group and severely diarrheic lamb in the low-dose challenge group were identified as lambs sensitive/resistant to E. coli F17 (E. coli F17 -resistant/-sensitive candidate, AN/SE) according to the histopathological detection. Results of intestinal contents bacteria plate counting revealed that the number of bacteria in the intestinal contents of SE lambs was 102~3-fold greater than that of the AN lambs, especially in the jejunum. Then, 16S rRNA sequencing was conducted to profile the intestinal microbiota using the jejunal contents, and the results showed that SE lambs had higher Lactococcus and a lower Bacteroidetes:Firmicutes ratio and intestinal microbiota diversity in the jejunum than AN lambs. Notably, high abundance of Megasphaera elsdenii was revealed in AN lambs, which indicated that Megasphaera elsdenii may serve as a potential probiotic for E. coli F17 infection. Our study provides an alternative challenge model for the identification of E. coli F17-sensitive/-resistant lambs and contributes to the basic understandings of intestinal microbiota in lambs with different susceptibilities to E. coli F17.

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Sheep β-Defensin 2 Regulates Escherichia coli F17 Resistance via NF-κB and MAPK Signaling Pathways in Ovine Intestinal Epithelial Cells.

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