Immunobiotics beneficially modulate TLR4 signaling triggered by lipopolysaccharide and reduce hepatic steatosis in vitro

Hepatic inflammation and injury may result from the translocation of pathological bacteria and their proinflammatory mediators. Probiotics attenuate hepatic diseases related to inflammation by exhibiting immunoregulatory effects. Therefore, this study was conducted to evaluate lipid reduction and immunoregulatory potentials of probiotic bacteria in vitro. HepG2 cells treated with total cellular fluid (TCF) of LABs reduced lipid accumulation. Moreover, cells responded to lipopolysaccharide (LPS) by producing higher levels of IL-6, IL-8, MCP-1, and TNF-α. TCF of LABs treatment showed remarkably diminished levels of the expression of these cytokines via modulation of the expression of TLR-negative regulators, as well as MAPK and NF-κB pathways. Moreover, heat-killed LABs were able to diminish TGF-β, IL-1β, and IL-6 and to increase IL-10 and TLR4 levels in THP-1 cells. LABs also decreased the protein level of TNF-α. These results demonstrated that immunobiotics exhibit potent immunoregulatory activity and may be used as effective therapeutic agents to alleviate inflammatory response.Fil: Kanmani, Paulraj. Dongguk University; Corea del SurFil: Ansari, AbuZar. Dongguk University; Corea del SurFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Kim, Hojun. Dongguk University; Corea del Su

Immunoregulatory effects triggered by immunobiotic Lactobacillus jensenii TL2937 strain involve efficient phagocytosis in porcine antigen presenting cells

Background: Immunobiotic Lactobacillus jensenii TL2937 modulates porcine mononuclear phagocytes from Peyer?s patches (PPMPs) and induces a differential production of pro- and anti-inflammatory cytokines in response to Toll-like receptor (TLR)-4 activation. Objective: In view of the important role played by phagocytosis in the activation of antigen presenting cells (APCs), the aim of the present work was to examine the interaction of TL2937 with porcine PPMPs focusing on phagocytosis. In addition, this study aimed to investigate whether the effects of L. jensenii TL2937 in porcine blood monocyte-derived dendritic cells (MoDCs) are similar to those found in PPMPs considering that MoDCs do not recapitulate all functions of mucosal APCs. Results: studies showed a high ability of porcine CD172a+ PPMPs to phagocytose L. jensenii TL2937. Interestingly, our results also revealed a reduced capacity of the non-immunomodulatory L. plantarum TL2766 to be phagocytosed by those immune cells. Phagocytosis of L. jensenii TL2937 by porcine PPMPs was partially dependent on TLR2. In addition, we demonstrated that TL2937 strain was able to improve the expression of IL-1, IL-12 and IL-10 in immature MoDCs resembling the effect of this immunobiotic bacterium on PPMPs. Moreover, similarly to PPMPs those immunomodulatory effects were related to the higher capacity of TL2937 to be phagocytosed by immature MoDCs. Conclusions: Microbial recognition in APCs could be effectively mediated through ligand-receptor interactions that then mediate phagocytosis and signaling. For the immunobiotic strain TL2937, TLR2 has a partial role for its interaction with porcine APCs and it is necessary to investigate the role of other receptors. A challenge for future research will be advance in the full understanding of the molecular interactions of immunobiotic L. jensenii TL2937 with porcine APCs that will be crucial for the successful development of functional feeds for the porcine host. This study is a step in that direction.Fil: Tsukida, Kohichiro. Tohoku University; JapónFil: Takahashi, Takuya. Tohoku University; JapónFil: Iida, Hikaru. Tohoku University; JapónFil: Kanmani, Paulraj. Tohoku University; JapónFil: Suda, Yoshihito. Miyagi University; JapónFil: Nochi, Tomonori. Tohoku University; JapónFil: Ohwada, Shuichi. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Ohkawara, Sou. Meiji Seika Pharma Co., Ltd. Agricultural & Veterinary Division; JapónFil: Makino, Seiya. Meiji Co., Ltd. Division of Research and Development; JapónFil: Kano, Hiroshi. Meiji Co., Ltd. Division of Research and Development; JapónFil: Saito, Tadao. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Kitazawa, Haruki. Tohoku University; Japó

Modulation of porcine intestinal epitheliocytes immunetranscriptome response by Lactobacillus jensenii TL2937

In order to evaluate probiotic strains applicable for the beneficial immunomodulation of the porcine gut (immunobiotics), we previously developed a porcine intestinal epitheliocyte cell line (PIE cells). Here, transcriptomic studies using PIE cells were performed considering that this information would be valuable for understanding the mechanisms involved in the protective activity of the immunobiotic strain Lactobacillus jensenii TL2937 against intestinal inflammatory damage in pigs. In addition, those studies would provide criteria for selecting biomarkers for the screening of new immunobiotic strains. We performed microarray analysis to investigate the transcriptomic response of PIE cells to the challenge with heat-stable Enterotoxigenic Escherichia coli (ETEC) pathogen-associated molecular patterns (PAMPs) and, the changes induced by L. jensenii TL2937 in that response. The approach allowed us to obtain a global overview of the immune genes involved in the response of PIE cells to heat-stable ETEC PAMPs. We observed that L. jensenii TL2937 differently modulated gene expression in ETEC PAMPs-challenged PIE cells. Microarray and RT-PCR analysis indicated that the most remarkable changes in PIE cells transcriptomic profile after heat-stable ETEC PAMPs challenge were observed in chemokines, adhesion molecules, complement and coagulation cascades factors. In addition, an anti-inflammatory effect triggered by TL2937 strain in PIE cells was clearly demonstrated. The decrease in the expression of chemokines (CCL8, CXCL5, CXCL9, CXCL10, and CXCL11), complement (C1R, C1S, C3, and CFB), and coagulation factors (F3) by L. jensenii TL2937 supports our previous reports on the immunoregulatory effect of this strain. These results provided clues for the better understanding of the mechanism underlying host-immunobiotic interaction in the porcine host. The comprehensive transcriptomic profiles of PIE cells provided by our analyses successfully identified a group of genes, which could be used as prospective biomarkers for the screening and evaluation of new anti-inflammatory immunobiotics for the prevention of inflammatory intestinal disorders in pigs.Fil: Kobayashi, Hisakazu. Tohoku University; JapónFil: Albarracín, Leonardo Miguel. Tohoku University; Japón. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Sato, Nana. Tohoku University; JapónFil: Kanmani, Paulraj. Tohoku University; JapónFil: Kober, Humayun A.K.M.. Tohoku University; Japón. Chittagong Veterinary and Animal Sciences University. Department of Dairy and Poultry Science; BangladeshFil: Ikeda-Ohtsubo, Wakako. Tohoku University; JapónFil: Suda, Yoshihito. Miyagi University; JapónFil: Nochi, Tomonori. Tohoku University; JapónFil: Aso, Hisashi. Tohoku University; JapónFil: Makino, Seiya. Meiji Co., Ltd. Food Science Research Labs.; JapónFil: Kano, Hiroshi. Meiji Co., Ltd. Food Science Research Labs.; JapónFil: Ohkawara, Sou. Meiji Seika Pharma Co., Ltd. Agricultural and Veterinary Division; JapónFil: Saito, Tadao. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; Japó

Evaluation of the immunoregulatory capacities of feed microbial materials in porcine intestinal immune and epithelial cells.

The establishment of drug-free feeding systems has been required for secure and healthy lives- tock production. Although functional feed materials containing microorganisms as alternatives to enhance intestinal immunity are expected to be beneficial for reducing diarrhoea caused by pathogens in weaned piglets, the effects of such materials on porcine intestinal cells have not been investigated in detail. Therefore, this work evaluated the immunoregulatory functions of microbial feed materials in porcine intestinal immune and epithelial cells. Porcine immune cells isolated from Peyer?s patches and mesenteric lymph nodes were stimulated with six different feed materials containing microorganisms, and evaluated for lymphocyte mitogenicity and cytokine inductions. In addition, porcine intestinal epithelial cells were stimulated with the materials before treatment with heat-killed enterotoxigenic Escherichia coli (ETEC), and analyzed for the proinflammatory cytokine expressions. The material containing Bifidobacterium thermophilum significantly augmented lymphocytes? mitogenicity and also induced a high expression of IL-2, IL-6 and IFN-γ in immune cells, and inhibited ETEC-induced overexpression of IL-6 and IL-8 via regulation of Toll-like receptor signaling. These results suggest that this feed material stimulates intestinal epithelial and immune cells to exert immunoregulation, suggesting that this feed is expected to contribute to promoting the health of piglets without using antimicrobial feed materials.Fil: Kumagae, Naosuke. Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; Japón. Scientific Feed Laboratory Co. Ltd.; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Centro de Referencia para Lactobacilos (i); Argentina. Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; JapónFil: Tomosada, Yohsuke. Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; JapónFil: Kobayashi, Hisakazu . Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; JapónFil: Kanmani, Paulraj. Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; Japón. Japan Society for the Promotion of Science; JapónFil: Aso, Hisashi . Tohoku University. Graduate School of Agricultural Science. Cell Biology Laboratory; JapónFil: Sasaki, Takashi . Scientific Feed Laboratory Co. Ltd.; JapónFil: Yoshida, Motohiko . Scientific Feed Laboratory Co. Ltd.; JapónFil: Tanabe, Hiroshi. Scientific Feed Laboratory Co. Ltd.; JapónFil: Shibata, Isao. Scientific Feed Laboratory Co. Ltd.; JapónFil: Saito, Tadao . Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; JapónFil: Kitazawa, Haruki. Tohoku University. Graduate School of Agricultural Science. Laboratory of Animal Products Chemistry. Food and Feed Immunology Group; Japó

The Respiratory Commensal Bacterium Dolosigranulum pigrum 040417 Improves the Innate Immune Response to Streptococcus pneumoniae

Previously, we demonstrated that the nasal administration of Dolosigranulum pigrum 040417 differentially modulated the respiratory innate immune response triggered by the activation of Tolllike receptor 2 in infant mice. In this work, we aimed to evaluate the beneficial effects of D. pigrum 040417 in the context of Streptococcus pneumoniae infection and characterize the role of alveolar macrophages (AMs) in the immunomodulatory properties of this respiratory commensal bacterium. The nasal administration of D. pigrum 040417 to infant mice significantly increased their resistance to pneumococcal infection, differentially modulated respiratory cytokines production, and reduced lung injuries. These effects were associated to the ability of the 040417 strain to modulate AMs function. Depletion of AMs significantly reduced the capacity of the 040417 strain to improve both the reduction of pathogen loads and the protection against lung tissue damage. We also demonstrated that the immunomodulatory properties of D. pigrum are strain-specific, as D. pigrum 030918 was not able to modulate respiratory immunity or to increase the resistance of mice to an S. pneumoniae infection. These findings enhanced our knowledge regarding the immunological mechanisms involved in modulation of respiratory immunity induced by beneficial respiratory commensal bacteria and suggested that particular strains could be used as next-generation probiotics.Fil: Raya Tonetti, María Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Tomokiyo, Mikado. Tohoku University; JapónFil: Ortiz Moyano, Francisco Ramiro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Quilodrán Vega, Sandra. Universidad de Concepción; ChileFil: Yamamuro, Hikari. Tohoku University; JapónFil: Kanmani, Paulraj. Tohoku University; JapónFil: Melnikov, Vyacheslav. Saint Petersburg State University; RusiaFil: Kurata, Shoichiro. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentin

Peptidoglycan from Immunobiotic Lactobacillus rhamnosus Improves Resistance of Infant Mice to Respiratory Syncytial Viral Infection and Secondary Pneumococcal Pneumonia

Several research works have demonstrated that beneficial microbes with the capacity to modulate the mucosal immune system (immunobiotics) are an interesting alternative to improve the outcome of bacterial and viral respiratory infections. Among the immunobiotic strains with the capacity to beneficially modulate respiratory immunity, Lactobacillus rhamnosus CRL1505 has outstanding properties. Although we have significantly advanced in demonstrating the capacity of L. rhamnosus CRL1505 to improve resistance against respiratory infections as well as in the cellular and molecular mechanisms involved in its beneficial activities, the potential protective ability of this strain or its immunomodulatory cellular fractions in the context of a secondary bacterial pneumonia has not been addressed before. In this work, we demonstrated that the nasal priming with non-viable L. rhamnosus CRL1505 or its purified peptidoglycan differentially modulated the respiratory innate antiviral immune response triggered by toll-like receptor 3 activation in infant mice, improving the resistance to primary respiratory syncytial virus (RSV) infection, and secondary pneumococcal pneumonia. In association with the protection against RSV-pneumococcal superinfection, we found that peptidoglycan from L. rhamnosus CRL1505 significantly improved lung CD3+CD4+IFN-γ+, and CD3+CD4+IL-10+ T cells as well as CD11c+SiglecF+IFN-β+ alveolar macrophages with the consequent increases of IFN-γ, IL-10, and IFN-β in the respiratory tract. Our results also showed that the increase of these three cytokines is necessary to achieve protection against respiratory superinfection since each of them are involved in different aspect of the secondary pneumococcal pneumonia that have to be controlled in order to reduce the severity of the infectious disease: lung pneumococcal colonization, bacteremia, and inflammatory-mediated lung tissue injury

The Ability of Respiratory Commensal Bacteria to Beneficially Modulate the Lung Innate Immune Response Is a Strain Dependent Characteristic

We investigated whether the ability of commensal respiratory bacteria to modulate the innate immune response against bacterial and viral pathogens was a shared or strain-specific characteristic. Bacterial strains belonging to the Corynebacterium pseudodiphtheriticum and Dolosigranulum pigrum species were compared by studying their influence in the Toll-like receptor (TLR)-2- and TLR3-triggered immune responses in the respiratory tract, as well as in the resistance to Respiratory Syncytial Virus (RSV) and Streptococcus pneumoniae infections. We demonstrated that nasally administered C. pseudodiphteriticum 090104 or D. pigrum 040417 were able to modulate respiratory immunity and increase the resistance against pathogens, while other strains of the same species did not influence the respiratory immune responses, demonstrating a clear strain-dependent immunomodulatory effect of respiratory commensal bacteria. We also reported here that bacterium-like particles (BLP) and cell walls derived from immunomodulatory respiratory commensal bacteria are an interesting alternative for the modulation of the respiratory immune system. Our study is a step forward in the positioning of certain strains of respiratory commensal bacteria as next-generation probiotics for the respiratory tract.Fil: Ortiz Moyano, Francisco Ramiro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Raya Tonetti, María Fernanda. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina. Universidad Nacional de Tucumán. Facultad de Medicina; ArgentinaFil: Tomokiyo, Mikado. Tohoku University; JapónFil: Kanmani, Paulraj. Dongguk University; Corea del SurFil: Vizoso Pinto, María Guadalupe. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Instituto Superior de Investigaciones Biológicas. Universidad Nacional de Tucumán. Instituto Superior de Investigaciones Biológicas; Argentina. Universidad Nacional de Tucumán. Facultad de Medicina; ArgentinaFil: Kim, Hojun. Dongguk University; Corea del SurFil: Quilodrán Vega, Sandra. Universidad de Concepción; ChileFil: Melnikov, Vyacheslav. Gabrichevsky Research Institute; RusiaFil: Alvarez, Gladis Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Takahashi, Hideki. Tohoku University; JapónFil: Kurata, Shoichiro. Tohoku University; JapónFil: Kitazawa, Haruki. Tohoku University; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Tohoku University; Japó

Genomic Characterization of Lactobacillus delbrueckii TUA4408L and Evaluation of the Antiviral Activities of its Extracellular Polysaccharides in Porcine Intestinal Epithelial Cells

In lactic acid bacteria, the synthesis of exopolysaccharides (EPS) has been associated with some favorable technological properties as well as health-promoting benefits. Research works have shown the potential of EPS produced by lactobacilli to differentially modulate immune responses. However, most studies were performed in immune cells and few works have concentrated in the immunomodulatory activities of EPS in non-immune cells such as intestinal epithelial cells. In addition, the cellular and molecular mechanisms involved in the immunoregulatory effects of EPS have not been studied in detail. In this work, we have performed a genomic characterization of Lactobacillus delbrueckii subsp. delbrueckii TUA4408L and evaluated the immunomodulatory and antiviral properties of its acidic (APS) and neutral (NPS) EPS in porcine intestinal epithelial (PIE) cells. Whole genome sequencing allowed the analysis of the general features of L. delbrueckii TUA4408L genome as well as the characterization of its EPS genes. A typical EPS gene cluster was found in the TUA4408L genome consisting in five highly conserved genes epsA-E, and a variable region, which includes the genes for the polymerase wzy, the flippase wzx, and seven glycosyltransferases. In addition, we demonstrated here for the first time that L. delbrueckii TUA4408L and its EPS are able to improve the resistance of PIE cells against rotavirus infection by reducing viral replication and regulating inflammatory response. Moreover, studies in PIE cells demonstrated that the TUA4408L strain and its EPS differentially modulate the antiviral innate immune response triggered by the activation of Toll-like receptor 3 (TLR3). L. delbrueckii TUA4408L and its EPS are capable of increasing the activation of interferon regulatory factor (IRF)-3 and nuclear factor κB (NF-κB) signaling pathways leading to an improved expression of the antiviral factors interferon (IFN)-β, Myxovirus resistance gene A (MxA) and RNaseL

Immunobiotics counteract the expression of hepatic profibrotic genes via the attenuation of TGF-/SMAD signaling and autophagy in hepatic stellate cells

RT-PCR, Westernblot dat