Conversely, those studies suggested different mechanisms for the enhancement of innate immunity. Lactobacillus pentosus S-PT84 has been reported to activate

IL-12 production by dendritic cells and to induce IFN-γ production by NK or NKT cells in an IL-12-dependent manner in murine FK228 mouse splenocytes (Koizumi et al., 2008), whereas Shida et al. (2006a) demonstrated that Lactobacillus casei Shirota induced IFN-γ production by T cells through IL-12 secretion by monocytes in human peripheral blood mononuclear cells (PBMCs). It would be important for the clinical application of LAB to understand the mechanisms of the immunomodulating effects by LAB, and thus, further investigation is needed. In the present study, a Lactobacillus paracasei strain, MoLac-1, which strongly induces IL-12, was selected. We investigated the in vitro effects and mechanisms of heat-killed MoLac-1 on IFN-γ production and NK cells and the in vivo effects of oral administration of heat-killed MoLac-1 on NK cells. Further, we evaluated the effectiveness of MoLac-1 in ameliorating

IFV infection using a mouse model. Bacterial strains used in this study are listed in Fig. 1 and were originally isolated from human intestine, intestine of adult, intestine of infant, or dairy. Proteasome inhibition These strains, which were originally isolated mainly from human intestine and dairy source, were obtained from the Morinaga Culture Collection (MCC; Morinaga Milk Industry Co. Ltd, Zama, Japan), the American Type Culture Collection (ATCC; Manassas, VA), and the Japan Collection of Microorganisms (JCM; Riken, Wako, Japan). The MoLac-1 (MCC1375) strain was isolated from the feces of healthy adults and identified as L. paracasei by carbohydrate fermentation patterns using the API 50 CH kit (bioMérieux, Marcy l’Etoile, France), 16S rRNA gene nucleotide sequences, and DNA–DNA hybridization technique. For bacterial Amylase culture, MRS broth (Becton Dickinson, Cockeysville, MD) was used for the strains belonging to Lactobacillus, MRS broth supplemented with 0.05% l-cysteine was used for the strains belonging to Bifidobacterium,

and M-17 broth (OXOID Ltd., Hampshire, UK) supplemented with 1% glucose was used for the strains belonging to Lactococcus, Streptococcus, or Enterococcus. Bacteria were cultured at 37 °C for 16 h, washed twice with phosphate-buffered saline (PBS), and then washed twice with distilled water. The bacteria were heat-killed at 100 °C for 30 min and lyophilized. One microgram of lyophilized MoLac-1 contained approximately 1.9 × 106 microorganisms as enumerated using bacterial counting chamber. Specific pathogen-free BALB/c mice were obtained from Japan SLC (Hamamatsu, Japan). All experiment protocols involving mice were performed according to the guidelines of the Prime Minister’s Office in Japan (no. 6, March 27, 1980). IFV A/PR/8/34 (H1N1) adapted to mice was stored at Japan Biological Science Inc.