In addition, we have also reported an abundant intracellular expression of TLR3 INK 128 in a porcine intestinal epithelial (PIE) cell line [22], which is in line with findings of Liu et al. [8] that demonstrated that the non-transformed porcine jejunum epithelial cell line (IPEC-J2) expresses TLR3 constitutively. We characterized the immune response triggered by poly(I:C) challenge in PIE cells and in PIE-immune cell co-cultures, and demonstrated that these systems are valuable tools for studying in vitro the immune response triggered by TLR3/RIG-I on IECs and the interaction between IECs and immune cells [22,

23]. In this study, we therefore aimed to use these porcine in vitro systems to gain insight into the mechanisms involved in the immunomodulatory effect of CRL1505 strain, and concentrated our attention in the crosstalk between L. rhamnosus CRL1505, PIE cells and APCs in order to deepen our knowledge about the mechanisms, through which this strain may help preventing viral diarrhoea episodes. Methods Microorganisms Lactobacillus rhamnosus CRL1505 (Lr1505) and L. rhamnosus CRL1506 (Lr1506) belong to CERELA Culture Collection and were originally isolated from goat milk [11]. These strains were grown in Man-Rogosa-Sharpe (MRS) broth at 37°C. For immunomodulatory assays, overnight cultures

PCI 32765 were harvested by centrifugation, washed three times with sterile PBS, counted in a Petroff-Hausser counting chamber, selleck screening library and re-suspended in DMEM until use. PIE cell monocultures A non-transformed porcine intestinal

epithelial cell line (PIE), characterized by its ability to build a monolayer with a cobblestone and epithelial-like morphology and close contacts between cells was used as described before [22, 23]. Briefly, PIE cells were grown on type I collagen-coated dishes using DMEM (Gibco, Japan) supplemented with 10% fetal calb serum (FCS, Sigma). PIE cells were incubated at 37°C and 5% CO2. Passages were done by treating the monolayer with sucrose/EDTA for 4 min and detaching the cells with 0.04% trypsin. Isolation of adherent population from swine Peyer’s patches (PPs) Suspensions of porcine PP immunocompetent cells were prepared from adult swine intestine. This study was carried out in strict accordance with the recommendations in the Guide for the Care and Use of Laboratory Animals of the Guidelines for Animal Experimentation of Tohoku University, Sendai, Japan. The present study was approved by the Institution Animal Care and Use Committee of Tohoku University with a permitted No. 2011-noudou-5 and all efforts were made to minimize suffering. Swine PPs were cut into small pieces and gently pressed through a nylon mesh to prepare single immune cell suspensions.