Daily Rhythms of P‐glycoprotein Expression in Mice

Recent studies have shown the gene expression of several transporters to be circadian rhythmic. However, it remains to be elucidated whether the expression of P‐glycoprotein, which is involved in the transport of many medications, undergoes 24 h rhythmicity. To address this issue, we investigated daily profiles of P‐glycoprotein mRNA and protein levels in peripheral mouse tissues. In the liver and intestine, but not in the kidney, Abcb1a mRNA expression showed clear 24 h rhythmicity. On the other hand, Abcb1b and Abcb4, the other P‐glycoprotein genes, did not exhibit significant rhythmic expression in the studied tissues. In the intestine, levels of whole P‐glycoprotein also exhibited a daily rhythm, with a peak occurring in the latter half of the light phase and a trough at the onset of the light phase. Consistent with the day‐night change of P‐glycoprotein level, the ex vivo accumulation of digoxin, an Abcb1a P‐glycoprotein substrate, into the intestinal segments at the onset of dark phase was significantly lower than it was at the onset of the light phase. Thus, Abcb1a P‐glycoprotein expression, and apparently its function, are 24 h rhythmic at least in mouse intestine tissue. This circadian variation might be involved in various chronopharmacological phenomena.     

Biosynthetic relationship among aflatoxins B1, B2, G1, and G2.

Aspergillus parasiticus NIAH-26, a UV-irradiated mutant of A. parasiticus SYS-4 (NRRL 2999), produces neither aflatoxins nor precursors. When sterigmatocystin (ST) or O-methylsterigmatocystin was fed to this mutant in YES medium, aflatoxins B1 (AFB1) and G1 (AFG1) were produced. When dihydrosterigmatocystin (DHST) or dihydro-O-methylsterigmatocystin was fed to this mold, aflatoxins B2 (AFB2) and G2 (AFG2) were produced. The reactions from ST to AFB1 and DHST to AFB2 were also observed in the cell-free system and were catalyzed stepwise by the methyltransferase and oxidoreductase enzymes. In the feeding experiments of strain NIAH-26, the convertibility from ST to AFB1-AFG1 was found to be remarkably suppressed by the coexistence of DHST in the medium, and the convertibility from DHST to AFB2-AFG2 was also suppressed by the presence of ST. When some other mutants which endogenously produce a small amount of aflatoxins (mainly AFB1 and AFG1) were cultured with DHST, the amounts of AFB1 and AFG1 produced were significantly decreased, whereas AFB2 and AFG2 were newly produced. In similar feeding experiments in which 27 kinds of mutants including these mutants were used, most of the mutants which were able to convert exogenous ST to AFB1-AFG1 were also found to convert exogenous DHST to AFB2-AFG2. These results suggest that the same enzymes may be involved in the both biosynthetic pathways from ST to AFB1-AFG1 and DHST to AFB2-AFG2. The reactions described herein were not observed when the molds had been cultured in the YEP medium.     

Role of the essential thiol group in the thiol-activated cytolysin from Clostridium perfringens

A hemolysin, 0-toxin, produced by Clostridium perfringens has one cysteinyl residue in the free thiol form which is essential for its hemolytic activity. The cysteinyl residue was shown to be located at a position about 5 kDa from the C terminus of the molecule by the method of cysteine-specific chemical cleavage. Modification of the residue with a thiol-blocking agent, 5,5'-dithiobis(2-nitrobenzoic acid), reduced the binding affinity of the toxin to sheep erythrocytes to 1/100 that of intact toxin, resulting in a failure of binding at low cell concentrations (0.5%). Thus the failure of hemolysis at low cell concentrations is primarily ascribed to a decreased affinity of the toxin for erythrocytes. Effects of the modification on the lytic processes were examined using high cell concentrations where considerable amounts of modified toxin bound to the cells. The modified toxin hemolyzes erythrocytes once it binds to them; however, the efficiency of hemolysis is reduced by the modification. These, and additional results indicating that modification alters the sensitivity of toxin molecules to protease digestion, show that thiol-modification inactivates the toxin by affecting both binding and the subsequent lytic processes, probably through a conformational change introduced in the toxin molecules.     

Enhanced Excretion of Fecal Neutral Sterols and the Hypocholesterolemic Property of 4-O-Methylascochlorin in Mice

The hypocholesterolemic property of experimental agent 4-O-methylascochlorin (MAC) was examined in male mice. MAC exerted hypocholesterolemic activity without hepatomegaly in mice given MAC mixed in a standard laboratory diet for a week. MAC also significantly reduced plasma total cholesterol (p-TC) in mice when it was administered immediately after meals by gastric intubation in controlled feeding. However, the oral administration through gastric tube was completely ineffective on p-TC in mice with empty stomach. These results suggest a close relationship between hypocholesterolemic efficacy and the timing of diet intake. Hypercholesterolemia also induced by a high fat-cholesterol diet was unaffected by MAC when it was given mixed with the diet.

From the isotopic study, three plausible mechanism were proposed for the mode of action: (I) enhanced output of biliary cholesterol, (II) inhibition of intestinal cholesterol absorption followed by an increment of fecal neutral sterols and (III) modulation of cholesterol partition in the plasma.     

Chronotherapy of maxacalcitol on skin inflammation induced by topical 12-O-tetradecanoylphorbol-13-acetate in mice

ABSTRACT

In general, chronotherapy is desirable for a more effective and/or safe dosage regimen. In this study, a daily rhythm of skin vitamin D receptor (VDR) and chronotherapeutic profiles of maxacalcitol, a vitamin D analogue, were evaluated using mice with skin inflammation induced by topical 12-O-tetradecanoylphorbol-13-acetate (TPA). This study showed that skin nuclear VDR expression in TPA-treated mice has a daily rhythm with the peak at the middle of active period. The effects of maxacalcitol were greater after dosing during early to middle of active period than those after dosing during early to middle of inactive period. These data suggest that chronotherapeutic profiles of maxacalcitol partly depend on the daily rhythm of skin nuclear VDR in TPA-treated mice. Because TPA-treated mice are considered as one of animal models of psoriasis, these animal data might be helpful for establishing chronotherapeutic approach of maxacalcitol in clinical practice.     

Extra and Intracellular Synthesis of Nickel Oxide Nanoparticles Mediated by Dead Fungal Biomass

The use of dead biomass of the fungus Hypocrea lixii as a biological system is a new, effective and environmentally friendly bioprocess for the production and uptake of nickel oxide nanoparticles (NPs), which has become a promising field in nanobiotechnology. Dead biomass of the fungus was successfully used to convert nickel ions into nickel oxide NPs in aqueous solution. These NPs accumulated intracellularly and extracellularly on the cell wall surface through biosorption. The average size, morphology and location of the NPs were characterized by transmission electron microscopy, high-resolution transmission electron microscopy, scanning electron microscopy, and energy dispersive X-ray spectroscopy. The NPs were mainly spherical and extra and intracellular NPs had an average size of 3.8 nm and 1.25 nm, respectively. X-ray photoelectron spectroscopy analysis confirmed the formation of nickel oxide NPs. Infrared spectroscopy detected the presence of functional amide groups, which are probable involved in particle binding to the biomass. The production of the NPs by dead biomass was analyzed by determining physicochemical parameters and equilibrium concentrations. The present study opens new perspectives for the biosynthesis of nanomaterials, which could become a potential biosorbent for the removal of toxic metals from polluted sites.     

Latent infection with Corynebacterium kutscheri in mice after peroral inoculation and its provocation by cortisone (author's transl)]

Latent infection with Corynebacterium kutscheri in mice and its provocation by cortisone were studied with a rifampicin-resistant strain of the organism. Mice having been infected perorally began to excrete the organisms in feces within 6 hours, and most of them were found to be carrying the organisms in the intestine, especially in the cecum even 90 days after infection. In such state of latency, however, no organisms were detected in other main organs, and neither visible lesions nor serum agglutinin was detectable. The latent infection with excretion of the organisms in feces after peroral infection was shown to become overt and fatal by cortisone treatment made even 90 days after infection. In infected mice excreting no organisms in feces and having bites on their skin, the wounds became severe ulcers after cortisone treatment resulting in septicemia.