Evaluation of anti-inflammatory immunobiotics in porcine intestinal epithelial cells

HARUKI KITAZAWA; TOMOYUKI SHIMAZU; MASANORI TOHNO; JULIO VILLENA; SYOICHI HOSOYA; HITOMI FUJIE; YOSHIHITO SUDA; TAKESHI SHIMOSATO; HISASHI ASO; SEIYA MAKINO; SHUNJI IKEGAMI; YASUSHI KAWAI; TADAO SAITO

Košice

Congreso; International Scientific Conference on Probiotics and Prebiotics 2010; 2010

Introduction Intestinal epithelial cells (IECs) are exposed to a variety of antigens including foods, pathogens and commensal bacteria, and are involved in the regulation of the mucosal immune response to pathogenic bacteria. Immunobiotics have been proposed as probiotics that regulate mucosal immune responses, resulting in both local and systemic immunity. Although immunobiotics are thought to confer protection from pathogenic infection in IECs, the precise mechanism of the anti-inflammatory effects of immunobiotics remains unclear. In the present study, we analyzed the anti-inflammatory effect of immunobiotics on the inflammatory response induced by Enterotoxigenic Escherichia coli (ETEC), a major pathogen of neonatal swine, in porcine intestinal epitheliocyte (PIE) cells [1]. Methods　 Twenty strains of lactic acid bacteria (LAB) which have been selected by the response to porcine Toll-like receptor (TLR) transfectant [2] were studied. PIE cells were stimulated with LAB for 48 hrs before the stimulation by ETEC for 12 hrs. The mRNA expression of selected inflammatory cytokines (IL-6, 8) was analyzed by real-time PCR method. Inflammatory signaling (MAP and I kappa B) via TLRs was evaluated by western blot and real-time PCR analysis. Cytokine induction on PIE cells and T cell subsets in co-culture with porcine Peyer’s patch immune cells was analyzed by flow-cytometry. Results Pretreatment of PIE cells with Lactobacills jensenii strain significantly reduced expression of IL-6 and IL-8 without augmentation of TLR 4 expression and secretion of anti-inflammatory factors. The strain also inhibited the activation of p38. Furthermore a negative regulator of TLR signaling was significantly increased following the addition of L. jensenii. The anti-inflammatory action was also confirmed on PIE cells in co-culture with porcine Peyer’s patch immune cells. Interestingly, IL-10 production in regulatory T cells related to anti-inflammation was augmented by L. jensenii in co-culture system. Discussion In this "new world" of food and feed immunology much remains unknown about the underlying mechanisms of intestinal mucosal immunity. Accordingly, many details remain unclear about the effects of food and feed components and their receptors on intestinal immune responses. The developed evaluation system for immunobiotics in our research activities may provide important clues at the molecular level on PRR signal transduction pathways and recognition mechanisms through immunobiotics in innate and adaptive immune responses, and hold promise in basic research and development of “immunobiotic foods and feeds” to prevent specific diseases in humans and animals [3]. [1] Moue et al., BBA, 1780, 134-144 (2008). [2] Tohno et al., Anim. Sci. J., (2007) [3] Kitazawa et al., Anim. Sci. J., (2008).