Isolation of food-derived bacteria inducing interleukin-22 in B cells.

Recently, we found a novel function of the lactic acid bacterium Tetragenococcus halophilus derived from miso, a fermented soy paste, that induces interleukin (IL)-22 production in B cells preferentially. IL-22 plays a critical role in barrier functions in the gut and skin. We further screened other bacteria species, namely, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Weissella, Pediococcus, and Bacillus, in addition to Tetragenococcus and found that some of them possessed robust IL-22-inducible function in B cells in vitro. This process resulted in the augmented expression of activation markers CD86 and CD69 on B and T cells, respectively. However, these observations were not correlated with IL-22 production. We isolated Bacillus coagulans sc-09 from miso and determined it to be the best strain to induce robust IL-22 production in B cells. Furthermore, feeding B. coagulans sc-09 to mice augmented the barrier function of the skin regardless of gut microbiota.

Isolation of immune-regulatory Tetragenococcus halophilus from miso.

Tetragenococcus halophilus is a halophilic lactic acid bacterium that exists in the traditional Japanese seasoning miso-a fermented soy paste. Considering the popularity of miso as a component of healthy diet, we attempted to evaluate the immunoregulatory functions of T. halophilus spices isolated from miso. We screened 56 strains that facilitated the upregulation of activation markers such as CD86 and CD69 on B cells and T cells in vitro. Of these, 7 strains (Nos. 1, 3, 13, 15, 19, 30, and 31) were found to preferentially induce the CD86 expression on B cells. Furthermore, DNA microarray analysis revealed that T. halophilus strain No. 1 significantly augmented the gene expressions of CD86, CD70, IL-10, INF-γ, and IL-22 in B cells. We confirmed these results at the protein level by flow cytometry. Mice feeding diet containing 1% T. halophilus No. 1 exhibited significantly greater IgA production in the serum. Furthermore, a diet containing 1% T. halophilus No. 1 augmented ovoalbumin (OVA)-specific IgG titer in mice upon OVA/alum immunization. Thus, we demonstrated that T. halophilus No. 1 is a strong immunomodulatory strain with potential as a probiotic.

Colorimetric inorganic pyrophosphate assay using a double cycling enzymatic method.

Pyruvate phosphate dikinase (PPDK, EC 2.7.9.1) from the hyperthermophile Thermotoga maritima was biochemically characterized with the aim of establishing a colorimetric assay for inorganic pyrophosphate (PPi). When heterologously expressed in Escherichia coli, T. maritima PPDK (TmPPDK) was far more stable any other PPDK reported so far: it retained >90% of its activity after incubation for 1 h at 80°C, and >80% of its activity after incubation for 20 min at pHs ranging from 6.5 to 10.5 (50°C). In contrast to PPDKs from protozoa and plants, this TmPPDK showed very long-term stability at low temperature: full activity was retained even after storage for at least 2 years at 4°C. TmPPDK was successfully applied to a novel colorimetric PPi assay, which employed (i) a PPi cycling reaction using TmPPDK and nicotinamide mononucleotide adenylyltransferase (EC 2.7.7.1) from Saccharomyces cerevisiae and (ii) a NAD cycling reaction to accumulate reduced nitroblue tetrazolium (diformazan). This enabled detection of 0.2 µM PPi, making this method applicable for preliminary measurement of PPi levels in PCR products in an automatic clinical analyzer.

Intake of fermented soybean (natto) increased locomotor activity in mice.

We fed mice food granules containing fermented soybean (natto in Japanese) powder (hereafter "natto granules") for 14 d to investigate whether natto granules had any effects on mouse behavior. We noted an enhancement of locomotor activity in natto-granule-fed mice compared to control and soybean-pellet-fed mice. This enhanced locomotor activity was blocked by a low dose of haloperidol (1 microg/kg i.p.), a dopamine receptor antagonist, but not by methysergide, a serotonin 5-HT(1/2) receptor antagonist. The results suggest that the enhanced locomotor activity induced by continuous intake of natto granules in mice is sensitive to haloperidol.

Effects of soybean food pellets on m-CPP-induced anxiety model of mice.

In this study, we gave the soybean powder-added food pellets (soybean pellets) to investigate anti-anxious effects of soybean in male mice. Twenty eight days after feeding control pellets or soybean pellets, we observed the behavioral changes in the elevated plus maze. There was no significant difference on the time spent in the open arms (%) between mice fed the control and soybean pellets. When we administered m-chlorophenylpiperazine (m-CPP, 2.5 mg/kg, i.p.) to mice, the mice fed control pellets showed the decrease in the time spent in the open arms, suggesting that anxiety-like behavior was induced by m-CPP. On the other hand, we could not observe the m-CPP-induced anxiety-like behavior in mice fed soybean pellets in this test. These results suggest that soybean pellets may attenuate anxiety-like behavior in mice.