Effect of Saccharomyces cerevisiae var. Boulardii and β-galactomannan oligosaccharide on porcine intestinal epithelial and dendritic cells challenged in vitro with Escherichia coli F4 (K88)

Abstract

Probiotic and prebiotics, often called "immune-enhancing" feed additives, are believed to deal with pathogens, preventing the need of an immune response and reducing tissue damage. In this study, we investigated if a recently developed β-galactomannan (βGM) had a similar protective role compared to Saccharomyces cerevisiae var. Boulardii (Scb), a proven probiotic, in the context of enterotoxigenic Escherichia coli (ETEC) infection. ETEC causes inflammation, diarrhea and intestinal damage in piglets, resulting in large economic loses worldwide. We observed that Scb and βGM products inhibited in vitro adhesion of ETEC on cell surface of porcine intestinal IPI-2I cells. Our data showed that Scb and βGM decreased the mRNA ETEC-induced gene expression of pro-inflammatory cytokines TNF-α, IL-6, GM-CSF and chemokines CCL2, CCL20 and CXCL8 on intestinal IPI-2I. Furthermore, we investigated the putative immunomodulatory role of Scb and βGM on porcine monocyte-derived dendritic cells (DCs) per se and under infection conditions. We observed a slight up-regulation of mRNA for TNF-α and CCR7 receptor after co-incubation of DC with Scb and βGM. However, no differences were found in DC activation upon ETEC infection and Scb or βGM co-culture. Therefore, our results indicate that, similar to probiotic Scb, prebiotic βGM may protect intestinal epithelial cells against intestinal pathogens. Finally, although these products may modulate DC activation, their effect under ETEC challenge conditions remains to be elucidated.

Figure 1

Adherence of ETEC on pig intestinal IPI-2I cells in the presence of Scb or βGM. Attachment of ETEC on IPI-2I cells co-cultured with Scb (A) or βGM (B) is inhibited in a dose-dependent manner. Data are presented as mean percentage ± S.D (n = 5) representative of 3 independent assays. Means with different superscripts (a, b, c) are significantly different (p < 0.05).

Figure 2

Effects of Scb and βGM on cytokine and chemokine mRNA expression in IPI-2I cells cultured with ETEC. IPI-2I cells (1 × 10⁶ cells/well) were co-cultured with Scb (3 yeast/cell) or βGM (10 μg/mL) with ETEC (1 × 10⁷ CFU/well) for 3 h. Data are presented as means of mRNA relative expressions ± SD (n = 6). Results are representative of 3 independent experiments. Means with different superscripts (a, b, c, d, e) are significantly different (p < 0.0.5).

Figure 3

Effect of Scb and βGM on IL6 and CXCL8 secretion induced by ETEC. Cytokine IL6 (A) and chemokine CXCL8 (B) concentration in supernatants from IPI-2I cells (1 × 10⁶ cells/well) co-cultured for 24 h with ETEC (1 × 10⁷ CFU/well) is decreased by Scb (3 yeast/cell) or βGM (10 μg/mL). Data were presented as mean ± S.D (n = 6). Data are representative of 3 independent experiments. Means with different superscripts (a, b, c, d) are significantly different (p < 0.05).

Figure 4

ETEC-induced gene expression in porcine DCs co-cultured with Scb or βGM. Relative mRNA expression of proinflammatory cytokines (TNFα, GM-CSF, IL6, IL10), chemokines (CXCL8, CCL17), receptors (CCR7, TLR2, TLR4) and regulatory factors (APRIL, BAFF, TGFβ) in DCs is enhanced by ETEC. Data were presented as mean ± S.D (n = 6). Treatments with different letters (a, b, c, d) mean p < 0.05.