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ABSTRACT: The oral intake of Lactobacillus spp. can provide beneficial effects to the host by modulating the immune response. Atopic dermatitis (AD) is an allergic inflammatory disease mediated by various immune responses. In this study, we examined the effect of a Lactobacillus strain, Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 (OLL1073R-1), on AD development in a murine model of AD that was developed by the topical application of mite antigen in NC/Nga mice. The oral intake of heat-killed OLL1073R-1 cells inhibited both the development of dermatitis and the elevation of an acute inflammation marker, serum amyloid A. Another bacterial strain, Lactobacillus rhamnosus OLL2984, exerted no inhibitory effects on dermatitis. The oral intake of heat-killed OLL1073R-1 cells also attenuated secretion of IL-6 from lymph node cells in response to mite antigen and reduced IL-6 levels in inflamed tissues, such as auricles. Production of IFN-γ or IL-4 was not influenced by OLL1073R-1 intake. We also found that inhibition of IL-6 signaling by gp130-Fc (a fusion protein consisting of the extracellular portion of glycoprotein 130 fused to the Fc region of human IgG1) markedly decreased the severity of dermatitis in NC/Nga mice. Moreover, secretion of IL-6 by lymph node cells was augmented in NC/Nga mice compared with that in BALB/c mice. These results indicate that IL-6 plays an essential role in the development of dermatitis in the NC/Nga mouse model of AD, and that OLL1073R-1 inhibits dermatitis, at least in part, by suppressing the IL-6 response.
Journal of Dairy Science 03/2013; · 2.56 Impact Factor
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Naoya Takanashi,
Yohsuke Tomosada,
Julio Villena,
Kozue Murata,
Takuya Takahashi,
Eriko Chiba,
Masanori Tohno,
Tomoyuki Shimazu,
Hisashi Aso,
Yoshihito Suda, Shuji Ikegami,
Hiroyuki Itoh,
Yasushi Kawai,
Tadao Saito,
Susana Alvarez,
Haruki Kitazawa
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ABSTRACT: BACKGROUND: Previously, a bovine intestinal epithelial cell line (BIE cells) was successfully established. This work hypothesized that BIE cells are useful in vitro model system for the study of interactions of microbial- or pathogen-associated molecular patterns (MAMPs or PAMPs) with bovine intestinal epithelial cells and for the selection of immunoregulatory lactic acid bacteria (LAB). RESULTS: All toll-like receptor (TLR) genes were expressed in BIE cells, being TLR4 one of the most strongly expressed. We demonstrated that heat-stable PAMPs of enterotoxigenic Escherichia coli (ETEC) significantly enhanced the production of IL-6, IL-8, IL-1alpha and MCP-1 in BIE cells by activating both NF-kappaB and MAPK pathways. We evaluated the capacity of several lactobacilli strains to modulate heat-stable ETEC PAMPs-mediated inflammatory response in BIE cells. Among these strains evaluated, Lactobacillus casei OLL2768 attenuated heat-stable ETEC PAMPs-induced pro-inflammatory response by inhibiting NF-kappaB and p38 signaling pathways in BIE cells. Moreover, L. casei OLL2768 negatively regulated TLR4 signaling in BIE cells by up-regulating Toll interacting protein (Tollip) and B-cell lymphoma 3-encoded protein (Bcl-3). CONCLUSIONS: BIE cells are suitable for the selection of immunoregulatory LAB and for studying the mechanisms involved in the protective activity of immunobiotics against pathogen-induced inflammatory damage. In addition, we showed that L. casei OLL2768 functionally modulate the bovine intestinal epithelium by attenuating heat-stable ETEC PAMPs-induced inflammation. Therefore L. casei OLL2768 is a good candidate for in vivo studying the protective effect of LAB against intestinal inflammatory damage induced by ETEC infection or heat-stable ETEC PAMPs challenge in the bovine host.
BMC Microbiology 03/2013; 13(1):54. · 3.04 Impact Factor
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Julio Villena,
Rie Suzuki,
Hitomi Fujie,
Eriko Chiba,
Takuya Takahashi,
Yohsuke Tomosada,
Tomoyuki Shimazu,
Hisashi Aso,
Shyuichi Ohwada,
Yoshihito Suda, Shuji Ikegami,
Hiroyuki Itoh,
Susana Alvarez,
Tadao Saito,
Haruki Kitazawa
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ABSTRACT: Previously, we demonstrated that Lactobacillus jensenii TL2937 attenuates the inflammatory response triggered by activation of Toll-like receptor 4 (TLR-4) in porcine intestinal epithelial cells. In view of the critical importance of antigen-presenting cell (APC) polarization in immunoregulation, the objective of the present study was to examine the effect of strain TL2937 on the activation patterns of APCs from swine Peyer's patches (PPs). We demonstrated that direct exposure of porcine APCs to L. jensenii in the absence of inflammatory signals increased expression of interleukin-10 (IL-10) and transforming growth factor β in CD172a(+) APCs and caused them to display tolerogenic properties. In addition, pretreatment of CD172a(+) APCs with L. jensenii resulted in differential modulation of the production of pro- and anti-inflammatory cytokines in response to TLR4 activation. The immunomodulatory effect of strain TL2937 was not related to a downregulation of TLR4 but was related to an upregulation of the expression of three negative regulators of TLRs: single immunoglobulin IL-1-related receptor (SIGIRR), A20, and interleukin-1 receptor-associated kinase M (IRAK-M). Our results also indicated that TLR2 has an important role in the anti-inflammatory activity of L. jensenii TL2937, since anti-TLR2 antibodies blocked the upregulation of SIGIRR and IRAK-M in CD172a(+) APCs and the production of IL-10 in response to TLR4 activation. We performed, for the first time, a precise functional characterization of porcine APCs from PPs, and we demonstrated that CD172a(+) cells were tolerogenic. Our findings demonstrate that adherent cells and isolated CD172a(+) cells harvested from swine PPs were useful for in vitro study of the inflammatory responses in the porcine gut and the immunomodulatory effects of immunobiotic microorganisms.
Clinical and vaccine immunology: CVI 05/2012; 19(7):1038-53. · 2.37 Impact Factor
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ABSTRACT: Yogurt fermented with Lactobacillus delbrueckii ssp. bulgaricus OLL1073R-1 (1073R-1) has been shown to reduce the risk of catching cold in the healthy elderly (Makino et al., Br. J. Nutr., 104, 998-1006, 2010). In addition, the exopolysaccharides (EPS) produced by 1073R-1 were also reported to exert immunostimulatory effects in mice such as the augmentation of NK cell activity (Makino et al., J. Dairy Sci., 89, 2873-81, 2006). So, we investigated anti-influenza virus effects of this yogurt and EPS in mice. The yogurt (0.4 ml/day) and EPS (20 μg/day) were orally administered to BALB/c mice for 21 days prior to intranasal infection with influenza virus A/PR/8/34 (H1N1). As a result, the survival periods were prolonged in both yogurt- and EPS-treated groups compared to the water-treated group. Moreover, in these groups, we observed significant decrease of influenza virus titer and significant increase of anti-influenza virus antibodies (IgA, IgG(1)) in the bronchoalveolar lavage fluid at 4 days post infection NK cell activity of splenocytes in both groups was also increased significantly. EPS was further fractionated into neutral EPS (NPS) and acidic EPS (APS), and the NPS (20 μg/day) or the APS (20 μg/day) was orally administered to mice for 21 days prior to the intranasal infection. The survival periods were prolonged in APS-treated group, but not in NPS-treated group. Consequently, we concluded that the yogurt fermented with 1073R-1 exerted anti-influenza virus effects in mice by its immunopotentiating activity, and suggested that the APS produced by 1073R-1 was one of active ingredients.
International immunopharmacology 12/2011; 11(12):2246-50. · 2.21 Impact Factor
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Tomoyuki Shimazu,
Julio Villena,
Masanori Tohno,
Hitomi Fujie,
Shoichi Hosoya,
Takeshi Shimosato,
Hisashi Aso,
Yoshihito Suda,
Yasushi Kawai,
Tadao Saito,
Seiya Makino, Shuji Ikegami,
Hiroyuki Itoh,
Haruki Kitazawa
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ABSTRACT: The effect of Lactobacillus jensenii TL2937 on the inflammatory immune response triggered by enterotoxigenic Escherichia coli (ETEC) and lipopolysaccharide (LPS) in a porcine intestinal epitheliocyte cell line (PIE cells) was evaluated. Challenges with ETEC or LPS elicited Toll-like receptor 4 (TLR4)-mediated inflammatory responses in cultured PIE cells, indicating that our cell line may be useful for studying inflammation in the guts of weaning piglets. In addition, we demonstrated that L. jensenii TL2937 attenuated the expression of proinflammatory cytokines and chemokines caused by ETEC or LPS challenge by downregulating TLR4-dependent nuclear factorκB (NF-κB) and mitogen-activated protein kinase (MAPK) activation. Furthermore, we demonstrated that L. jensenii TL2937 stimulation of PIE cells upregulated three negative regulators of TLRs: A20, Bcl-3, and MKP-1, deepening the understanding of an immunobiotic mechanism of action. L. jensenii TL2937-mediated induction of negative regulators of TLRs would have a substantial physiological impact on homeostasis in PIE cells, because excessive TLR inflammatory signaling would be downregulated. These results indicated that PIE cells can be used to study the mechanisms involved in the protective activity of immunobiotics against intestinal inflammatory damage and may provide useful information for the development of new immunologically functional feeds that help to prevent inflammatory intestinal disorders, including weaning-associated intestinal inflammation.
Infection and immunity 11/2011; 80(1):276-88. · 4.21 Impact Factor
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Shoichi Hosoya,
Julio Villena,
Tomoyuki Shimazu,
Masanori Tohno,
Hitomi Fujie,
Eriko Chiba,
Takeshi Shimosato,
Hisashi Aso,
Yoshihito Suda,
Yasushi Kawai,
Tadao Saito,
Susana Alvarez, Shuji Ikegami,
Hiroyuki Itoh,
Haruki Kitazawa
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ABSTRACT: ABSTRACT: This study analyzed the functional expression of TLR3 in various gastrointestinal tissues from adult swine and shows that TLR3 is expressed preferentially in intestinal epithelial cells (IEC), CD172a+CD11R1high and CD4+ cells from ileal Peyer's patches. We characterized the inflammatory immune response triggered by TLR3 activation in a clonal porcine intestinal epitheliocyte cell line (PIE cells) and in PIE-immune cell co-cultures, and demonstrated that these systems are valuable tools to study in vitro the immune response triggered by TLR3 on IEC and the interaction between IEC and immune cells. In addition, we selected an immunobiotic lactic acid bacteria strain, Lactobacillus casei MEP221106, able to beneficially regulate the anti-viral immune response triggered by poly(I:C) stimulation in PIE cells. Moreover, we deepened our understanding of the possible mechanisms of immunobiotic action by demonstrating that L. casei MEP221106 modulates the interaction between IEC and immune cells during the generation of a TLR3-mediated immune response.
Veterinary Research 11/2011; 42(1):111. · 4.06 Impact Factor
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ABSTRACT: Although oral ovabumin (OVA) administration suppressed the antibody (Ab) response in OVA-immunized mice, Lactococcus lactis increased OVA-specific IgG2a in these mice. L. lactis increased the casein-specific IgG level in NC/Nga mice fed on a casein diet. The percentage of CD4(+)CD25(+) cells was increased in DO11.10 mice orally given OVA, but this increase of CD4(+)CD25(+) cells were suppressed in L. lactis-fed DO11.10 mice.
Bioscience Biotechnology and Biochemistry 03/2011; 75(3):599-601. · 1.28 Impact Factor
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ABSTRACT: Increasing evidence suggests that the aryl hydrocarbon receptor (AhR) pathway has an important role in the regulation of inflammatory responses. Most recently, we have shown that the activation of the AhR pathway by a potent AhR agonist inhibits the development of dextran sodium sulfate (DSS)-induced colitis, a model of human ulcerative colitis, by the induction of prostaglandin E2 (PGE2) in the large intestine. Because several strains of probiotic lactic acid bacteria have been reported to inhibit DSS-induced colitis by unidentified mechanisms, we hypothesized that particular strains of lactic acid bacterium might have the potential to activate the AhR pathway, thereby inhibiting DSS-induced colitis. This study investigated whether there are specific lactic acid bacterial strains that can activate the AhR pathway, and if so, whether this AhR-activating potential is associated with suppression of DSS-induced colitis. By using AhR signaling reporter cells, we found that Lactobacillus bulgaricus OLL1181 had the potential to activate the AhR pathway. OLL1181 also induced the mRNA expression of cytochrome P450 family 1A1 (CYP1A1), a target gene of the AhR pathway, in human colon cells, which was inhibited by the addition of an AhR antagonist, α-naphthoflavon (αNF). In addition, mice treated orally with OLL1181 showed an increase in CYP1A1 mRNA expression in the large intestine and amelioration of DSS-induced colitis. Thus, OLL1181 can induce activation of the intestinal AhR pathway and inhibit DSS-induced colitis in mice. This strain of lactic acid bacterium has therefore the potential to activate the AhR pathway, which may be able to suppress colitis.
Immunology and Cell Biology 02/2011; 89(7):817-22. · 3.66 Impact Factor
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ABSTRACT: Immune senescence potentially leads to an increased risk of infections. It is desirable to augment the immune system and protect against infections by daily consumption of immunostimulatory food. The present study evaluated whether the intake of yoghurt fermented with Lactobacillus delbrueckii ssp. bulgaricus (L. bulgaricus) OLL1073R-1 has an effect on resistance to the common cold. We conducted two independent studies, in which fifty-seven (median age 74.5 years) and eighty-five healthy elderly individuals (median age 67.7 years) were participants. In each study, the subjects were divided into two groups based on age and sex and instructed to eat 90 g yoghurt or drink 100 ml milk once per d over an 8- or 12-week period. A meta-analysis of the results of these two independent studies showed the risk of catching the common cold was about 2.6 times lower (OR 0.39; P = 0.019) in the yoghurt group than in the milk group and the increase of natural killer cell activity was significantly higher in the yoghurt group than in the milk group (P = 0.028). In addition, the quality of life score for the 'eye/nose/throat' system after intake was significantly higher in the yoghurt group than in the milk group and the improvement of the score was correlated with the promotion of natural killer cell activity. In conclusion, consumption of yoghurt fermented with L. bulgaricus OLL1073R-1 augmented natural killer cell activity and reduced the risk of catching the common cold in elderly individuals.
The British journal of nutrition 10/2010; 104(7):998-1006. · 3.45 Impact Factor
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ABSTRACT: A randomized, double-blind, placebo-controlled clinical trial was conducted to determine whether oral administration of heat-killed Lactobacillus gasseri OLL2809 would affect the immune response and reduce the symptoms of Japanese cedar pollinosis (JCP) in subjects with JCP. Following a 1-week pre-observation period, the subjects were randomly divided into two groups and were orally administered a placebo or tablets containing 100 mg of L. gasseri OLL2809 per d for 8 weeks during the pollen season in 2007. The results showed no obvious differences between the groups. Supplementary subgroup analysis revealed that the OLL2809 subgroups with CAP-RAST scores of 4 or 5 exhibited improvement in nasal symptoms scores and serum allergy-related items, including Japanese cedar pollen-specific IgE levels. L. gasseri OLL2809 was found to be effective in reducing symptoms in subjects with a high predisposition to allergies by modulating systemic immune systems.
Bioscience Biotechnology and Biochemistry 10/2009; 73(9):1971-7. · 1.28 Impact Factor
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ABSTRACT: Lactobacillus gasseri OLL2809 strongly stimulates the production of interleukin (IL)-12 (p70) by innate immune cells. Thus, it is expected to ameliorate allergic diseases. We investigated whether the oral administration of heat-killed L. gasseri OLL2809 suppressed eosinophilia in cedar pollen antigen-challenged mice.
BALB/c mice sensitized with Japanese cedar pollen extract were intraperitoneally challenged with the same extract. The mice were orally given heat-killed L. gasseri OLL2809 at doses of 0.5, 1, or 2mg/day throughout the experimental period (21 d). After 24 hours of the challenge, the eosinophil number and cytokine levels in the peritoneal lavage fluid and the serum antigen-specific IgG levels were determined.
On administering varying amounts of heat-killed L. gasseri OLL2809, the number of eosinophils among the total number of cells was significantly reduced in all groups. In addition, the eosinophil number significantly decreased, and the eosinophil-suppression rate significantly increased by 44% in the 2-mg group. Although the serum immunoglobulin (Ig) G2a and IgG1 levels were not affected, the IgG2a/IgG1 ratio increased significantly in the 2-mg group compared with that of the control group. Furthermore, the administration of heat-killed L. gasseri OLL2809 resulted in the induction of IL-2 and reduction in granulocyte-macrophage colony-stimulating factor levels in peritoneal lavage fluid.
We demonstrated that the oral administration of heat-killed L. gasseri OLL2809 suppresses eosinophilia via the modulation of Th1/Th2 balance. These observations suggested that heat-killed L. gasseri OLL2809 might potentially ameliorate the increased number of eosinophils in patients with Japanese cedar pollinosis.
Allergology International 12/2008; 57(4):397-403.
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ABSTRACT: Lactobacillus gasseri OLL2809, a probiotic lactic acid bacterium, strongly stimulates interleukin (IL)-12 (p70) production by murine splenocytes; therefore, it is expected to ameliorate allergic diseases. Although many studies have investigated characteristics of the immunostimulatory activity of probiotics, little is known about how bacterial growth conditions affect the activity. In this study, we investigated the effects of the growth conditions of L. gasseri OLL2809 on the stimulation of IL-12 (p70) production. L. gasseri OLL2809 was grown under various culture conditions including different cultivation periods, media, and culture pH, and IL-12 (p70) production by murine splenocytes stimulated with these bacterial cells was determined. The results revealed that IL-12 (p70) production (i) increased with the growth of the bacterial cells and was higher in stationary-phase cells than in logarithmic-phase cells; (ii) it was higher in the cells grown in acidic pH; and (iii) it decreased when the cells were incubated in a buffer at neutral pH prior to heat treatment. These observations indicated that stimulation of IL-12 (p70) production is affected by culture medium pH. In addition, the observations of a difference in the stimulation of IL-12 (p70) production by L. gasseri OLL2809 grown under various conditions are consistent with the characteristics of autolysis. Therefore, it was deduced that the integrity of the bacterial cells is necessary for the stimulatory effect on IL-12 (p70) production and that acidic pH and heat treatment contributed to the stimulation by inhibiting the activity of autolysins indigenous to the bacteria. Our result suggests that cultivation until the stationary phase under acidic pH is required for the effective production of probiotics with immunostimulatory activity.
International Journal of Food Microbiology 01/2008; 120(3):274-81. · 3.33 Impact Factor
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ABSTRACT: Toll-like receptor 2 (TLR2) is a receptor for a variety of microbial components, and it also mediates activation signals in the cell relating to the innate immune system. In order to evaluate the precise molecular immunoregulation by various strains of lactic acid bacteria (LAB) via TLR2, the swine TLR2 (sTLR2)-expressing transfectant was constructed using human embryonic kidney (HEK) 293 cells. It is demonstrated that intact immunobiotic LAB can induce immune responses through TLR2, and that different nuclear factor-κB (NF-κB) activities of various strains can be accurately detected by sTLR2-expressing HEK293 cells. Furthermore, cellular activation of NF-κB via TLR2 is reflected in enhanced binding and uptake of LAB. The sTLR2-expressing HEK293 cells were also useful for characterizing the expression pattern of type I helper T (Th1) and type II helper T (Th2) cytokines by the stimulation of immunobiotic LAB. These results suggest that sTLR2-expressing HEK293 cells may be useful in certain molecular immunoassay systems for producing new physiologically functional foods with intestinal immunomodulatory abilities, such as the maintenance of Th1/Th2 polarization.
Animal Science Journal 03/2007; 78(2):195 - 205. · 0.86 Impact Factor
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ABSTRACT: After organ transplantation, severe osteoporosis is occasionally seen, and the use of immunosuppressants is thought to be one of the causes of such osteoporosis. In the present study, we investigated the effects of FK506 monotherapy on bones and determined the mechanism of onset of osteoporosis, both by assessing chronological changes in bone metabolism and by identifying factors that facilitate bone resorption. In 8-week-old male Sprague-Dawley rats, FK506 (1 mg/kg) was injected intraperitoneally every day for 5 weeks (FK506-treated group), and for comparison, physiological saline was administered in the same manner in a control group of rats. Serum and urine samples were collected at weeks 0, 1, 3, and 5 of administration. The femur and tibia were collected within 24 h of the final administration. When compared to the control group, findings on three-dimensional micro-computed tomography of the femur for the FK506-treated group showed a significant decrease in trabecular bone volume. The level of serum osteocalcin in the FK506-treated group at week 1 of administration was significantly higher than the control. Throughout the administration period, the sum of urinary pyridinoline (PYD) and deoxypyridinoline (Dpd) was significantly higher in the FK506-treated group. Of the various bone resorption factors tested, the level of serum parathyroid hormone (PTH) in the FK506-treated group was significantly higher than the control at week 3 of administration. The results of the present study confirmed that FK506 monotherapy in rats induced high-turnover osteoporosis. Soon after the start of FK506 administration, bone formation and resorption were elevated, and PTH appeared to have been involved in the maintenance of the elevated bone resorption.
Journal of Bone and Mineral Metabolism 02/2004; 22(6):554-60. · 2.27 Impact Factor
