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Xenobiotics are usually detoxified by drug-metabolizing enzymes and excreted from the body. The expression of many of drug-metabolizing enzymes is regulated by the aryl hydrocarbon receptor (AHR). Some substances in vegetables have the potential to be AHR ligands. To search for vegetable components that exhibit AHR-mediated transcriptional activity, we assessed the activity of vegetable extracts and identified the active compounds using the previously established stable AHR-responsive HepG2 cell line. Among the hot water extracts of vegetables, the highest activity was found in ginger. The ethyl acetate fraction of the ginger hot water extract remarkably induced AHR-mediated transcriptional activity, and the major active compound was found to be 6-shogaol. Subsequently, the mRNA levels of AHR-targeting drug-metabolizing enzymes (CYP1A1, UGT1A1, and ABCG 2) and the protein level of CYP1A1 in HepG2 cells were shown to be increased by 6-shogaol. This is the first report that 6-shogaol can regulate the expression of detoxification enzymes by AHR activation.

Vibrio parahaemolyticus in uncooked seafood causes acute gastroenteritis. The microorganism has two sets of type III secretion systems and two hemolysins. When it injects its effector proteins into a host cell via type III secretion system 1, one of the type III secretion systems induces secretion of interleukin (IL)-8, a proinflammatory chemokine, through the phosphorylation of ERK 1/2 and p38 MAPK. Although probiotics have beneficial effects on hosts and can help control some infectious diseases, there is little research on the efficacy of probiotics in V. parahaemolyticus infection. Here we pretreated V. parahaemolyticus-infected human intestinal epithelial cells with heat-killed Lactobacillus brevis KB290, a probiotic isolated from fermented vegetables (traditional Japanese pickles) and utilized as an ingredient of beverages and supplementary foods, and demonstrated its efficacy in enhancing IL-8 secretion from V. parahaemolyticus-infected cells. Among the three heat-killed lactic acid bacterial strains we tested, L. brevis KB290 induced the highest level of IL-8 secretions in the infected cells. Relative to control cells (Caco-2 cells pretreated with PBS), V. parahaemolyticus-infected Caco-2 cells pretreated with heat-killed L. brevis KB290 secreted IL-8 earlier, although concentrations were similar 450min after infection. Heat-killed L. brevis KB290 pretreatment also induced earlier ERK 1/2 phosphorylation, greater p38 MAPK phosphorylation, and enhanced IL-8 mRNA expression. Heat-killed L. brevis KB290 accelerated IL-8 secretion, a host cell immune response, in V. parahaemolyticus-infected cells. We consider this to be beneficial because IL-8 plays an important defensive role against infection, and would contribute to the repair of injured epithelial cells.

Twenty plant-derived and 18 non-plant-derived strains of Lactobacillus casei were compared for their growth in tryptone – yeast extract – glucose broth containing 0.3% bile by measuring absorbance at a wavelength of 620 nm after 24 h of incubation at 37 °C. Bile tolerance — a fundamental probiotic property — was calculated by dividing the experimental data by control values (growth without bile). We found that bile tolerance was strain specific but that the average bile tolerance of the plant-derived strains was significantly (P < 0.05) lower than that of the non-plant-derived strains tested. All tested strains could not deconjugate sodium taurocholate, indicating that the difference in bile tolerance was not due to the ability to deconjugate bile. The fatty acid compositions of the test strains with and without exposure to 0.3% bile were investigated, and a statistical correlation analysis between these compositions and their bile tolerance was conducted. The fatty acids correlated with bile tolerance differed between plant and non-plant lactobacilli. This is the first report to show that the origin (i.e., growth environment) of lactobacilli affects their fatty acid composition, which in turn, appears to be related to their bile tolerance.

Our purpose was to investigate the safety of the probiotic strain Lactobacillus brevis KB290. The European Qualified Presumption of Safety (QPS) evaluation approach was applied to the strain. We determined the strain's antibiotic resistance, verified it at the genetic level, and determined whether it could be transferred to intestinal microflora. Of 14 antibiotics tested, 11 showed MICs within the limits of the QPS criteria. However, the L. brevis KB290 MICs of ciprofloxacin (a fluoroquinolone), tetracycline, and vancomycin were two, four, and eight times, respectively, the breakpoint MICs suggested by the European Scientific Committee on Animal Nutrition, and the MIC of tetracycline was eight times the breakpoint MIC suggested by the European Scientific Panel on Additives and Products or Substances Used in Animal Feed. Using analysis of gapped-genome sequences, we found no known transferable determinants for tetracycline or vancomycin resistance, and we found no mutations in the quinolone resistance-determining regions of the genes encoding GyrA or ParC for ciprofloxacin resistance associated with insertion sequences, integrons, or transposons. These data were confirmed by using PCR primers specific for the respective genes. We assessed the transferability of the resistance traits in conjugation experiments with enterococci and obtained no transconjugants, strongly suggesting that the resistance traits were not transferable. This study demonstrated that the antibiotic resistance observed in L. brevis KB290 was due not to dedicated mechanisms but to intrinsic resistance. According to the QPS criteria, these results provide safety assurance for the ongoing use of L. brevis KB290 as a probiotic.

We examined the survivability of Lactobacillus brevis KB290 and derivative strain KB392 in artificial digestive juices and bile salts. The strains have similar membrane fatty acids but different amounts of cell-bound exopolysaccharides (EPS). In artificial digestive juices, KB290 showed significantly higher survivability than KB392, and homogenization, which reduced the amount of EPS in KB290 but not in KB392, reduced the survivability only of KB290. In bile salts, KB290 showed significantly higher survivability than KB392, and cell-bound EPS extraction with EDTA reduced the survivability of only KB290. Transmission electron microscopy showed there to be a greater concentration of cell-bound EPS in KB290 than in either KB392 or EDTA-treated or homogenized KB290. We conclude that KB290’s cell-bound EPS (which high performance liquid chromatography showed to be made up of glucose and N-acetylglucosamine) played an important role in bile salt tolerance.

Lactobacillus brevis KB290 (KB290), a plant-derived probiotic lactic acid bacterium, improves gut health and stimulates immune function. Here we extensively investigated the teratogenicity of KB290 in rats and rabbits. We observed no adverse maternal or fetal effects and concluded that the no observable adverse effect level for maternal general toxicity, maintenance of pregnancy, and teratogenicity should be ≥ 10(10) cfu/kg/day. Our results suggest that KB290 would be safe for pregnant females and their offspring.

To elucidate the physiological responses to a social stressor, we exposed mice to an isolation stress and analyzed their hepatic gene expression profiles using a DNA microarray. Male BALB/c mice were exposed to isolation stress for 30 days, and then hepatic RNA was sampled and subjected to DNA microarray analysis. The isolation stress altered the expression of 420 genes (after considering the false discovery rate). Gene Ontology analysis of these differentially expressed genes indicated that the stress remarkably downregulated the lipid metabolism-related pathway through peroxisome proliferator-activated receptor-α, while the lipid biosynthesis pathway controlled by sterol regulatory element binding factor 1, Golgi vesicle transport, and secretory pathway-related genes were significantly upregulated. These results suggest that isolation for 30 days with a mild and consecutive social stress regulates the systems for lipid metabolism and also causes endoplasmic reticulum stress in mouse liver.

Lactobacillus brevis KB290 (KB290), a plant-derived probiotic lactic acid bacterium, reportedly improves gut health and stimulates immune function. Here we extensively investigated the geno-, acute, subacute, and subchronic toxicity of KB290 and its bacterial translocation potential. KB290 was non-mutagenic in the bacterial reverse mutation assay by the preincubation method. In the single oral dose toxicity test, KB290 at 10(9) cfu/ml was nontoxic at maximum capacity (20 ml/kg). When 10(8), 10(9), or 10(10) cfu/kg was administered daily to rats by gavage for 2 weeks (subacute assay), we observed no clear treatment-related effect and no evidence of bacterial translocation from the gastrointestinal tract. When it was administered for 13 weeks (subchronic assay), we again observed no clear treatment-related effect and no significant toxicological effect. Based on those results, we consider 10(10) cfu/kg per day, the highest dose tested, to be the no observed adverse effect level (NOAEL). These results suggest that KB290 is safe for human consumption.

Suckling (12-d-old) rats that were fasted for 15 h received epidermal growth factor (EGF) s.c. (0.5 and 1.0 microgram per rat, i.e. approximately 2 and 4 micrograms/100 g of body weight), together with motility markers 51Cr-EDTA or Poly R-478, and were killed 45 min later. Counts were measured separately in the stomach and the small intestine, which was divided into 12 segments. Administration of EGF delayed gastric emptying. In controls, the stomach contained 26.1 +/- 1.6% (mean +/- SEM); in EGF-treated rats the stomach contained 75.9 +/- 10.2% and 75.7 +/- 2.5% of the total 51Cr-EDTA counts given. EGF had the maximum effect (1.0 microgram) when given simultaneously with 51Cr-EDTA. Significant, but lower, effects of EGF were seen with the administration of EGF preceded by 10 min or followed by 10 and 20 min with the administration of 51Cr-EDTA (65.8 +/- 5.8%, 60.0 +/- 6.4%, and 54.1 +/- 4.2%, respectively). Small intestinal transit was also delayed. Administration of anti-EGF antiserum did not affect gastric emptying, but accelerated small intestinal transit as determined 30 min after administration of 51Cr-EDTA. These studies are the first to demonstrate the effect of EGF on gastrointestinal motility in vivo in suckling mammals.

Six oligosaccharides were first formed from lactinol by a transgalactosylation reaction catalyzed by Aspergillus oryzae β- d -galactosidase. From the results of methylation analysis, MS, and 1 H- and 13 C-NMR studies, it was concluded that these oligosaccharides are O -β- d -galactopyranosyl-(1 → 4)- O -β- d -galactopyranosyl- (1 → 4)- d -glucitol, O -β- d -galactopyranosyl-(1 → 3)- O -β- d -galactopyranosyl-(1 → 4)- d -glucitol, O -β- d -galactopyranosyl-(1 → 4)-[ O -β- d -galactopyranosyl-(1 → 6)]- d -glucitol, O -β- d - galactopyranosyl-(1 → 6)- O -β- d -galactopyranosyl-(1 → 4)- d -glucitol, O -β- d -galactopyranosyl-(1 → 4)-[ O -β- d - galactopyranosyl-(1 → 5)- d -glucitol, and O -β- d -galactopyranosyl-(1 → 4)-[ O -β- d -galactopyranosyl-(1 → 1)]- d -glucitol. The last three are newly observed oligosaccharides.