Abstract
BackgroundThe ovine rumen is involved in host defense responses and acts as the immune interface with the environment. The ruminal mucosal epithelium plays an important role in innate immunity and secretes antimicrobial innate immune molecules that have bactericidal activity against a variety of pathogens. Defensins are cationic peptides that are produced by the mucosal epithelia and have broad-spectrum antimicrobial activity. Sheep ÎČ-defensin-1 (SBD-1) is one of the most important antibacterial peptides in the rumen. The expression of SBD-1 is regulated by the probiotic, Saccharomyces cerevisiae (S.c); however, the regulatory mechanism has not yet been elucidated. In the current study, the effects of S.c on the expression and secretion of SBD-1 in ovine ruminal epithelial cells were investigated using quantitative real-time PCR (qPCR) and enzyme-linked immunosorbent assay (ELISA). In addition, specific inhibitors were used to block the nuclear factor kappa-light-chain enhancer of activated B cells (NF-ÎșB), p38, JNK, and ERK1/2 signalling pathways separately or simultaneously, to determine the regulatory mechanism(s) governing S.c-induced SBD-1 upregulation.ResultsIncubation with S.c induced release of SBD-1 by ovine ruminal epithelial cells, with SBD-1 expression peaking after 12 h of incubation. The highest SBD-1 expression levels were achieved after treatment with 5.2 Ă 107 CFUâmLâ 1 S.c. Treatment with S.c resulted in significantly increased NF-ÎșB, p38, JNK, ERK1/2, TLR2, and MyD88 mRNA expression. Whereas inhibition of mitogen-activated protein kinases (MAPKs) and NF-ÎșB gene expression led to a decrease in SBD-1 expression.ConclusionsS.c was induced SBD-1 expression and the S.c-induced up-regulation of SBD-1 expression may be related to TLR2 and MyD88 in ovine ruminal epithelial cells. This is likely simultaneously regulated by the MAPKs and NF-ÎșB pathways with the p38 axis of the MAPKs pathway acting as the primary regulator. Thus, the pathways regulating S.c-induced SBD-1 expression may be related to TLR2-MyD88-NF-ÎșB/MAPKs, with the TLR2-MyD88-p38 component of the TLR2-MyD88-MAPKs signalling acting as the main pathway.
Highlights
The ovine rumen is involved in host defense responses and acts as the immune interface with the environment
S.c induced sheep ÎČ-defensin-1 (SBD-1) mRNA expression in ovine ruminal epithelial cells In the current study, results of quantitative real-time PCR (qPCR) indicated that S.c promoted SBD-1 mRNA transcription in a time- and dose-dependent manner (Fig. 2a)
When the incubation time was constant, the expression of SBD-1 mRNA was highest when cells were stimulated with 5.2 Ă 107 CFUâmLâ 1 S.c, and was significantly higher than compared with cells stimulated with other S.c concentrations and compared to the control group (P < 0.01)
Summary
The ovine rumen is involved in host defense responses and acts as the immune interface with the environment. A group of broad-spectrum antimicrobial agents secreted by epithelial cells in response to microbial infection and stimulation from the main components of microbial pathogens, have recently garnered a lot of attention. SBD-2 expression was found to be confined to the tongue, ileum, and colon, while SBD-1 expression was identified in the entire digestive tract (from the tongue to the colon), with highest expression located in the rumen [30, 31]. This suggests that ÎČ-defensins constitute an important component of ovine rumen innate immunity
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