Selection of anti-inflammatory Lactobacillus plantarum strains through their abitlity to stimulate TLR2, NOD1 and NOD2 receptors

JULIO VILLENA; KYOKO MORIE; HORTENSIA ZELAYA; YO MUROFUSHI; MASANORI TOHNO; HARUKI KITAZAWA

San Miguel de Tucumán

Simposio; IV International Symposium on Lactic Acid Bacteria - Food, Health and Applications; 2013

Centro de Referencia para Lactobacilos. CERELA-CONICET

We have previously demonstrated that the NF-kB-reporter assay using a porcine Toll-like receptor 2-expressing transfectant (HEKTLR2 cells) is highly effective for selecting lactic acid bacteria with immunoregulatory capacities. In the present work, we established porcine nucleotide-binding oligomerization domain-1 (NOD1)- and NOD2-expressing transfectants (HEKNOD1 and HEKNOD2 cells). Expression of NOD1 and NOD2 receptors in transfectants was confirmed by flow cytometric analysis. In addition, by using confocal laser microscopy, we showed that both NOD1 and NOD2 were not expressed on the cell surface, but were located in the intracellular compartment, closely to the plasma membrane. A total of 31 strains of Lactobacillus planatrum were tested for their capacity to stimulate NF-kB pathway in HEKTLR2, HEKNOD1 and HEKNOD2 cells and the mitogenic assay using porcine Peyer?s patches immunocompetent cells. Three functional groups were defined according to the capacity to stimulate HEK cells and the mitogenic activity. Then L. plantarum strains were classified into high, moderate and low activities groups. Strains MY1 (low), MY7 (moderate) and MY27 (high) were selected for further experiments. We next evaluated whether the three selected strains were able to attenuate the inflammatory response triggered by activation of Toll-like receptor 4 (TLR4) in porcine intestinal epithelial (PIE) cells. Challenge of PIE cells with lipopolysaccharide (LPS) significantly increased IL-8 and IL-6 levels. L. plantarum MY27 significantly reduced IL-8 and IL-6 in LPS-challenged PIE cells (p