Enhancement of Anti-Tumoral Immunity by β-Casomorphin-7 Inhibits Cancer Development and Metastasis of Colorectal Cancer

β-Casomorphin-7 (BCM) is a degradation product of β-casein, a milk component, and has been suggested to affect the immune system. However, its effect on mucosal immunity, especially anti-tumor immunity, in cancer-bearing individuals is not clear. We investigated the effects of BCM on lymphocytes using an in vitro system comprising mouse splenocytes, a mouse colorectal carcinogenesis model, and a mouse orthotopic colorectal cancer model. Treatment of mouse splenocytes with BCM in vitro reduced numbers of cluster of differentiation (CD) 20⁺ B cells, CD4⁺ T cells, and regulatory T cells (Tregs), and increased CD8⁺ T cells. Administration of BCM and the CD10 inhibitor thiorphan (TOP) to mice resulted in similar alterations in the lymphocyte subsets in the spleen and intestinal mucosa. BCM was degraded in a concentration- and time-dependent manner by the neutral endopeptidase CD10, and the formed BCM degradation product did not affect the lymphocyte counts. Furthermore, degradation was completely suppressed by TOP. In the azoxymethane mouse colorectal carcinogenesis model, the incidence of aberrant crypt foci, adenoma, and adenocarcinoma was reduced by co-treatment with BCM and TOP. Furthermore, when CT26 mouse colon cancer cells were inoculated into the cecum of syngeneic BALB/c mice and concurrently treated with BCM and TOP, infiltration of CD8⁺ T cells was promoted, and tumor growth and liver metastasis were suppressed. These results suggest that by suppressing the BCM degradation system, the anti-tumor effect of BCM is enhanced and it can suppress the development and progression of colorectal cancer.     

Inactivation of Bacillus spores by the combination of moderate heat and low hydrostatic pressure in ketchup and potage

The combination effect of moderate heat and low hydrostatic pressure (MHP) on the reduction of Bacillus subtilis, Bacillus coagulans and Geobacillus stearothermophilus spores in food materials (potage and ketchup) was investigated. These bacterial spores were suspended in potage (pH 7), acidified potage (pH 4), neutralized ketchup (pH 7) and ketchup (pH 4). The suspensions were treated with and without pressure (100 MPa) and temperatures of 65-85 degrees C for 3 to 12 h. The bacterial spores were inactivated by 4-8 log cycles during MHP treatment in potage, acidified potage and ketchup, whereas the spores were highly resistant to long time heat treatment in potage and neutralized ketchup. The degrees of spore destruction were mostly dependent on pH and medium composition during MHP treatment. The inactivation effect in MHP treatment was higher at the pH 7 than at pH 4 both in ketchup and potage. The bacterial spores showed higher inactivation in potage than ketchup during MHP treatment.     

Characterization of monofunctional catalase KatA from radioresistant bacterium Deinococcus radiodurans

Catalase plays a key role in protecting cells against toxic reactive oxygen species. Here we report on the cloning, purification and characterization of a catalase (KatA, DR1998) from the extremely radioresistant bacterium Deinococcus radiodurans. The size of purified D. radiodurans KatA monomer was 65 kDa while gel filtration revealed that the size of the enzyme was 240 kDa, suggesting that KatA formed a homotetramer in solution. Purified KatA displayed a final specific activity of 68,800 U/mg of protein. The catalase activity of KatA was inhibited by sodium azide, sodium cyanide and 3-amino-1,2,4-triazole. The absorption spectrum of KatA exhibited a Soret band at 408 nm. The position of the spectral peak remained unchanged following reduction of KatA with dithionite. No peroxidase activity was found for KatA. These results demonstrate that D. radiodurans KatA is a typical monofunctional heme-containing catalase. The stability of KatA with respect to H2O2 stress was superior to that of commercially available Aspergillus niger and bovine liver catalases. The relative abundance of KatA in cells in addition to the H2O2 resistance property may play a role in the survival strategy of D. radiodurans against oxidative damage.     

Single cell-based analysis of torenia petal pigments by a combination of ArF excimer laser micro sampling and nano-high performance liquid chromatography (HPLC)-mass spectrometry

The molecular constituents of the petal pigments of the Torenia plant (Torenia hybrida) were analyzed on a single-cell basis by a combination of newly developed laser-microsampling and nano-flow liquid chromatography-electro spray ionization mass spectrometry (LC-ESIMS) techniques. Our method should provide a facile method for obtaining precise metabolic profiles of each cell in a single plant tissue.     

Dendrimer-based nanoprobe for dual modality magnetic resonance and fluorescence imaging

A novel PAMAM dendrimer-based nanoprobe for dual magnetic resonance and fluorescence imaging modalities was synthesized. Fluorescence studies revealed that Gd(III) complexation to the probe has no effect on the quantum yield; however, increases in the dye content resulted in partial quenching. The potential of the new nanoprobe, G6-(Cy5.5)(1.25)(1B4M-Gd)(145), as a dual modality imaging agent was demonstrated in vivo by the efficient visualization of sentinel lymph nodes in mice by both MRI and fluorescence imaging modalities.     

Engineering of Escherichia coli L-serine O-acetyltransferase on the basis of crystal structure: desensitization to feedback inhibition by L-cysteine

L-Serine O-acetyltransferase (SAT) from Escherichia coli catalyzes the first step of L-cysteine synthesis in E.coli and is strictly inhibited by the second step product, L-cysteine. To establish a fermentation process to produce L-cysteine, we embarked on a mutational study of E.coli SAT to desensitize the feedback inhibition by L-cysteine. The crystal structure and the reaction mechanism of SAT from E.coli have shown that the substrate L-serine and the inhibitor L-cysteine bind to the identical region in the SAT protein. To decrease the affinity for only L-cysteine, we first built the structure model of L-serine-binding SAT on the basis of the crystal structure with bound L-cysteine and compared these two structures. The comparison showed that the Calpha of Asp92 underwent a substantial positional change upon the replacement of L-cysteine by L-serine. We then introduced various amino acid substitutions at positions 89-96 around Asp92 by randomized, fragment-directed mutagenesis to change the position of the Asp92. As a result, we successfully obtained mutant SATs which have both extreme insensitivity to an inhibition by L-cysteine (the concentration that inhibits 50% activity; IC(50) = 1,100 micromol/l, the inhibition constant; K(i) = 950.0 micromol/l) and extremely high emzymatic activities.     

Flexible ultrasonic transducers

Flexible ultrasonic transducers (UTs) consisting of a metal foil, a piezoelectric ceramic film, and a top electrode have been developed. The flexibility is realized owing to the porosity of piezoelectric film and the thinness of metal foil. In this paper, the stainless steel (SS), lead-zirconate-titanate (PZT)/PZT composite and silver paste were chosen as metal foil, piezoelectric film, and top electrode materials, respectively. The SS foil serves as both substrate and bottom electrode. The PZT/PZT piezoelectric composite film is made by the sol-gel spray technique. PZT/PZT films of thicknesses from 40 to 70 microm were fabricated onto SS foils. The capability of these flexible sensors operated in the pulse-echo mode for nondestructive testing on flat and curved surfaces of different materials at room temperature and 160 degrees C has been demonstrated. Numerical simulations of the effects of the metal foil thickness on the ultrasonic performance of flexible UTs also were carried out, and the results are in reasonable agreement with experimental data. In addition, a PZT/PZT flexible transducer showed a signal strength comparable with that obtained by a commercial room temperature broad bandwidth transducer.     

Room-temperature miscibility gap in LixFePO4

The rechargeable lithium-ion cell is an advanced energy-storage system. However, high cost, safety hazards, and chemical instability prohibit its use in large-scale applications. An alternative cathode material, LiFePO(4), solves these problems, but has a kinetic problem involving strong electron/hole localization. One reason for this is believed to be the limited carrier density in the fixed monovalent Fe(3+)PO(4)/LiFe(2+)PO(4) two-phase electrode reaction in LixFePO4. Here, we provide experimental evidence that LixFePO4, at room temperature, can be described as a mixture of the Fe(3+)/Fe(2+) mixed-valent intermediate LialphaFePO4 and Li1-betaFePO4 phases. Using powder neutron diffraction, the site occupancy numbers for lithium in each phase were refined to be alpha=0.05 and 1-beta=0.89. The corresponding solid solution ranges outside the miscibility gap (0相似文献   

Xanthine oxidase inhibitory activity of alkyl gallates

A series (C1-C12) of alkyl gallates was examined for their effects on the activity of xanthine oxidase. Octyl (C8), decyl (C10), and dodecyl (C12) gallates competitively inhibited uric acid formation generated by xanthine oxidase, and the inhibition increased upon increasing the alkyl chain length. Interestingly, neither menthyl nor bornyl gallates inhibited uric acid formation. These data indicate that the hydrophobic alkyl portion is associated with the xanthine-binding site in the Mo-binding domain. It is likely that the linear alkyl portion interacts with the hydrophobic domain close to the binding site, and the hydrophobic interaction is crucial to inhibit the xanthine oxidase reaction. On the other hand, all of gallic acid and its esters equally suppress superoxide anion generation catalyzed by xanthine oxidase at low concentration. The suppression is not due to scavenging activity of these gallates but due to reduction of xanthine oxidase by these gallates. The reduced enzyme catalyzes the reaction to generate hydrogen peroxide and uric acid.     

Mechanism of Ca2+ increase in myoblasts derived from chicken embryos

The mechanism of intracellular calcium ions (Ca(2+)) increase in chicken myoblasts was studied using histological, immunohistochemical, immunoblotting and Ca(2+) imaging techniques. Mononuclear myoblasts at embryonic day 12 (E12) contained myofibrils in the peripheral cytoplasm, and the sarcoplasmic reticulum was observed in the cytoplasm. Several Ca(2+)-related receptors, namely acetylcholine (ACh) receptors, dihydropyridine receptors (DHPRs) and ryanodine receptors (RyRs), were detected in the tissue as early as E12. Western blotting analyses detected one band corresponding to RyR subtype 3 (RyR3) at E12 and two bands corresponding to RyR1 and RyR3 after E13. Ca(2+) imaging of mononuclear myoblasts in vitro revealed an intense Ca(2+)-increase response to ACh stimulation, and this effect was abolished after EGTA addition to the culture medium. Nifedipine treatment also led to a lack of Ca(2+) increase in response to ACh stimulation, while ryanodine treatment led to a weak Ca(2+)-increase response. On the other hand, multinuclear myoblasts showed a Ca(2+)-increase response to ACh stimulation in the presence of not only EGTA but also nifedipine, although ryanodine treatment led to a lack of Ca(2+) increase. These results suggest that the mechanism of Ca(2+) increase in mononuclear myoblasts involves extracellular Ca(2+) entry through DHPRs, which is amplified by Ca(2+) release from the cytoplasmic Ca(2+) store, while multinuclear myoblasts mainly depend on Ca(2+) release from the cytoplasmic Ca(2+) store.