The author started a research on xenobiotic metabolism at Graduate School of Pharmaceutical Sciences, Kyushu University in 1965. In 1968, an epidemic of a “strange disease”, called Yusho, occurred in western Japan. The epidemic was soon identified to be a food poisoning caused by the ingestion of commercial Kanemi rice bran oil which had been accidentally contaminated with large amounts of polychlorinated biphenyls (PCBs) and their related compounds such as polychlorinated dibenzofurans (PCDFs.) At first, in this review, our toxicological studies on Yusho during the early thirty years were briefly described. Next, the studies on aldehyde oxidase, a molybdenum hydroxylase, which is involved in the lactam formation reaction such as 1-phenyl-2-(2-oxopyrrolidine)pentane(oxoprolintane) from 1-phenyl-2-pyrrolidinopentane(prolintane) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP) lactam from 1-methyl-4-phenyl-2,3-dihydropyridinium ion (MPDP+) were also presented. Finally, we investigated how the xenobiotic metabolism of endocrine disrupting chemicals such as bisphenol A (BPA) and some isoflavones affects their estrogenic activities. In this study, we demonstrated that BPA is converted to 4-methyl-2,4-bis(4-hydroxyphenyl)pent-1-ene (MBP), an active metabolite as estrogen, by rat liver S9. In the cases of isoflavones, although genistein was inactivated, biochanin A, 4'-methoxy analogue of genistein, was activated to genistein by O-demethylation with rat liver S9.
The aim of our study was to produce a new diabetic model by a single i.p. injection of various doses of streptozotocin (STZ) to 8-week-old male Institute of Cancer Research (ICR) mice. When STZ was i.p. injected at doses rainging from 75 to 200 mg/kg, in the group injected 200 mg/kg STZ, the non-fasting serum glucose levels markedly rose from 1 week after STZ administration and the high glucose levels were maintained for 9 weeks. The serum glucose levels in the group administered 100 mg/kg STZ were within a normal range at 1 week after STZ administration, but thereafter continued to increase gradually till 9 weeks. In contrast with the serum glucose levels, a marked reduction in the percentage of the relative numbers of β-cells in the islets of pancreas from 1 week in mice injected 200 mg/kg STZ was observed. On the other hand, in 100 mg/kg STZ mice, the number of β-cells was almost normal percentage at 1 week and then continued to gradually decrease as the time went on throughout 24-week-observation. These results indicate that only the 100 mg/kg STZ injection produces slowly progressive diabetes mellitus in mice. Low molecular weight (LMW) chitosan (chitosan lactate, average of molecular weight: 20000) was administered as drinking water from prediabetic stage (from 2 weeks after 100 mg/kg STZ injection). The 0.2% or 0.8% LMW chitosan administration prevented the progression of 100 mg/kg STZ-induced slowly progressive diabetes mellitus. The mechanisms, which LMW chitosan is effective in this model may be discussed on β-cells.
This review documents my research for the past 29 years in the work of bacterial sporulation. The Gram-positive bacterium Bacillus subtilis forms spores when conditions are unsuitable for growth. The mature spores remain for long periods of starvation and are resistant to harsh environment. This property is attributed mainly to the unique figures of spore's outer layers, spore coat. The protein composition of the spores was comprehensively analyzed by a combination of SDS-PAGE and LC-MS/MS. The total of 154 proteins were identified and 69 of them were novel. The expression of the genes encoding them was dependent on sporulation-specific sigma factors, σF, σE, σG and σK. The expression of a coat protein gene, cotS, was dependent on σK and GerE. CotE is essential for the assembly of CotS in the coat layer. Many coat genes were identified by reverse genetics and the regulation of the gene expression was studied in detail. Some cot genes are functioned in the resistance to heat and lysozyme, and some of the coat proteins are involved in the specificity of germinants. The yrbA is essential in spore development, yrbA deficient cells revealed abnormal figures of spore coat structure and changed the response to germinants. The location of 16 coat proteins was determined by the observation of fluorescence microscopy using fluorescence-labelled proteins. One protein was assigned to the cortex, nine to the inner coat, and four to the outer coat. In addition, CotZ and CgeA appeared in the outermost layer of the spore coat.
In this review, we would outline the possible signaling system for three types of amino acids including glutamate (Glu), γ-aminobutyric acid (GABA) and D-serine (D-Ser) to play a role as an extracellular signal mediator in mechanisms underlying maintenance of the cellular homeostasis in skeletal tissues. Although Glu and GABA has been thought to be an excitatory/inhibitory amino acid neurotransmitter in the mammalian central nervous system, our molecular biological analyses give rise to a novel function for Glu and GABA as an autocrine and/or paracrine factor in three types of distinct cell types including osteoblasts, osteoclasts and chondrocytes in bone tissues. Moreover, D-Ser plays a pivotal role in osteoclastogenesis through a mechanism related to the incorporation of serine enantiomers in osteoclasts after the synthesis and subsequent release from adjacent osteoblasts. Accordingly, bone formation and maintenance seems to be under control by amino acid signaling in skeletal tissues as seen with neurotransmission in the brain.
Free radicals, reactive oxygen species (ROS), and redox status are known to be involved in cell signaling, proteomics, etc. in homeostasis and oxidative disease. Electron spin resonance (ESR) spectroscopy is a very sensitive and selective technique for detecting free radicals, and we have developed an in vivo ESR spectroscopy/imaging system combined with a nitroxyl probe for noninvasive assessment of free radical reactions in various disease models such as diabetes, cancer, liver/lung damage, gastric lesions, brain injury, etc. The systematic measurement with probes having different degrees of membrane permeability make it possible to clarify the location of free radical generation in blood vessels, cell membranes, and the cytoplasm. Our findings strongly indicate the great advantage of the in vivo ESR spectroscopy/imaging/nitroxyl probe technique in biomedical sciences. However, the spatial resolution of the ESR spectroscopy imaging system is insufficient due to the short T2 of probes. Proton magnetic resonance imaging (MRI) has significant clinical utility in the diagnosis of disease. Dynamic nuclear polarization (DNP)-MRI yields highly resolved images of free radical distribution in small animals by enhancing the water proton signal intensity via the Overhauser effect. We developed a new sequence for DNP-MRI and succeeded in obtaining simultaneous dual images using radicals labeled with 14N and 15N nuclei. This technique can visualize individual redox processes and the individual redox status of inner and/or outer cells in a dose-dependent manner. More recently, we have developed a novel DNP-MRI scanner and enables the use of DNP-MRI in different disease models.
The gastrointestinal tract is the organ that absorbs nutrients and water from foods and drinks. This organ is often exposed to various harmful xenobiotics, and therefore possesses various detoxification/barrier systems, including metabolizing enzymes and efflux transporters. Intestinal epithelial cells express ATP-binding cassette (ABC) efflux transporters such as P-glycoprotein, multidrug resistance-associated proteins (MRPs) and breast cancer resistance protein, in addition to various solute carrier (SLC) influx transporters. These transporters are expressed site- and membrane-specifically in enterocytes, which affects the bioavailability of ingested substrate drugs. Expression and/or function of transporters can be modulated by various compounds, including therapeutic drugs, herbal products, some foods, and by disease states. The modulation of transporters could cause unexpectedly higher or lower blood concentrations, marked inter- and intra-individual variations in pharmacokinetics, and unreliable pharmacological actions in association with toxicities of substrates. Recently, we found that hyperbilirubinemia, which occurs in some disease states, increased intestinal accumulation and toxicity of methotrexate, an MRP substrate, because of the suppression of MRP function by high plasma concentrations of conjugated bilirubin. We also attempted to ameliorate the intestinal toxicity of irinotecan hydrochloride by modulating the hepatic and intestinal functions of MRP2. This review summarizes our findings regarding the role of ABC transporters, especially MRPs, in oral bioavailability and in drug-induced intestinal toxicity. Our approach to treat intestinal toxicity using an MRP2 modulator is also described.
Nucleoside phosphoramidites (PAs) are the most widely used building blocks in the contemporary solid-phase synthesis of oligonucleotides. Accurate molecular weight measurements of such acid-labile compounds may be easily determined in a matrix system comprising triethanolamine (TEOA)-NaCl by liquid secondary ionization mass spectrometry (LSIMS) or fast atom bombardment mass spectrometry (FABMS), using a double-focusing mass spectrometer. The present method rapidly and easily measures the accurate MWs of various PAs as adduct ions [M+Na]+, affording molecular formulas in place of elemental analysis. Further, the intensities by LSIMS of [M+Na]+ were found to be enhanced to the highest degree by adjustment of the mole ratio of PA and NaCl, fixing the amount of TEOA. In addition, the effects of the metal ion in the matrix were investigated. The present method is a powerful tool for the mass spectral identification of polyfunctionalized nucleoside PAs.