In Vitro Transcriptome Response to a Mixture of Lactobacilli Strains in Intestinal Porcine Epithelial Cell Line

Food and feed supplements containing microorganisms with probiotic potential are of increasing interest due to their healthy promoting effect on human and animals. Their mechanism of action is still unknown. Using a microarray approach, the aim of this study was to investigate the differences in genome-wide gene expression induced by a mixture of three Lactobacillus strains (L. rhamnosus, L. plantarum, and L. paracasei) in intestinal porcine epithelial cells (IPEC-1) and to identify the genes and pathways involved in intestinal barrier functions.

Undifferentiated IPEC-1 cells seeded at a density of 2.0 × 10⁵/mL in 24-wells culture plates were cultivated at 37 °C and 5% CO₂ until they reached confluence (2⁻3 days). Confluent cells monolayer were then cultivated with 1 mL of fresh lactobacilli (LB) mixture suspension prepared for a concentration of approximately 3.3 × 10⁷ CFU/mL for each strain (1 × 10⁸ CFU/mL in total) for 3 h and analyzed by microarray using Gene Spring GX v.11.5.

The functional analysis showed that 1811 of the genes modulated by LB treatment are involved in signaling (95% up-regulation, 121 genes with a fold change higher than 10). The most enhanced expression was registered for AXIN2 (axis inhibition protein 2-AXIN2) gene (13.93 Fc, p = 0.043), a negative regulator of β-catenin with a key role in human cancer. LB affected the cellular proliferation by increasing 10 times (Fc) the NF1 gene encoding for the neurofibromin protein, a tumor suppressor that prevent cells from uncontrolled proliferation. The induction of genes like serpin peptidase inhibitor, clade A member 3 (SERPINA 3), interleukin-20 (IL-20), oncostatin M(OSM), granulocyte-macrophage colony-stimulating factor (GM-CSF), and the suppression of chemokine (C-X-C motif) ligand 2/macrophage inflammatory protein 2-alpha (CXCL-2/MIP-2), regulator of G-protein signaling 2 (RGS2), and of pro-inflammatory interleukin-18 (IL-18) genes highlights the protective role of lactobacilli in epithelial barrier function against inflammation and in the activation of immune response.

Gene overexpression was the predominant effect produced by lactobacilli treatment in IPEC-1 cells, genes related to signaling pathways being the most affected. The protective role of lactobacilli in epithelial barrier function against inflammation and in the activation of immune response was also noticed.