Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
35 巻 , 3 号
選択された号の論文の24件中1~24を表示しています
Review
  • Rieko Nakata, Satoru Takahashi, Hiroyasu Inoue
    原稿種別: Review
    2012 年 35 巻 3 号 p. 273-279
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Appropriate long-term drinking of red wine is associated with a reduced risk for lifestyle-related diseases such as cardiovascular disease and cancer, making resveratrol, a constituent of grapes and various other plants, an attractive compound to be studied. Historically, resveratrol has been identified as a phytoalexin, antioxidant, cyclooxygenase (COX) inhibitor, peroxisome proliferator-activated receptor (PPAR) activator, endothelial nitric oxide synthase (eNOS) inducer, silent mating type information regulation 2 homolog 1 (SIRT1) activator, and more. Despite scepticism concerning the biological availability of resveratrol, a growing body of in vivo evidence indicates that resveratrol has protective effects in several stress and disease models. Here, we provide a review of the studies on resveratrol, especially with respect to COX, PPAR, and eNOS activities, and discuss its potential for promoting human health.
Regular Articles
  • Ting Zhao, Yan-Na Cheng, Hai-Ning Tan, Jin-Feng Liu, Huan-Li Xu, Guang ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 280-288
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Human serum albumin (HSA) is used as an important plasma volume expander in clinical practice. However, the infused HSA may extravasate into the interstitial space and induce peripheral edema in treating the critical illness related to marked increase in capillary permeability. Such poor intravascular retention also demands a frequent administration of HSA. We hypothesize that increasing the molecular weight of HSA by PEGylation may be a potential approach to decrease capillary permeability of HSA. In the present study, HSA was PEGylated in a site-specific manner and the PEGylated HSA carrying one chain of polyethylene glycol (PEG) (20 kDa) per HSA molecule was obtained. The purity, PEGylated site and secondary structure of the modified protein were characterized by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), thiol group blockage method and circular dichroism (CD) measurement, respectively. In addition, the pharmacokinetics in normal mice was investigated, vascular permeability of the PEGylated HSA was evaluated in lipopolysaccharide (LPS)-induced lung injury mouse model and the pharmacodynamics was investigated in LPS-induced sepsis model with systemic capillary leakage. The results showed that the biological half-life of the modified HSA was approximately 2.3 times of that of the native HSA, PEG-HSA had a lower vascular permeability and better recovery in blood pressure and haemodilution was observed in rats treated with PEG-HSA. From the results it can be inferred that the chemically well-defined and molecularly homogeneous PEGylated HSA is superior to HSA in treating capillary permeability increase related illness because of its longer biological half-life and lower vascular permeability.
  • Young Sun Hwang, Kwang-Kyun Park, Won-Yoon Chung
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 289-300
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    An inability to control cancer cell invasion and metastasis is the leading cause of death in patients with cancer. The present study was performed to determine the anti-invasive effect of Kalopanaxsaponin A (KPS-A) on matrix metalloproteinase-9 (MMP-9)-meidated invasion in phorbol 12-myristate 13-acetate (PMA)-stimulated human oral squamous cell carcinoma (OSCC) cells and a murine xenograft model of human OSCC. KPS-A, isolated from Kalopanax pictus, inhibited PMA-induced proliferation and invasion as well as PMA-induced MMP-9 expression and secretion at non-cytotoxic doses. KPS-A treatment reduced the stability of PMA-induced MMP-9 mRNA and inhibited the PMA-induced cytoplasmic translocation of HuR. In PMA-treated cells, KPS-A treatment resulted in the intracellular accumulation of MMP-9 and suppressed Ras-associated binding 1A (Rab1A) expression. KPS-A treatment suppressed PMA-induced phosphorylation of extracellular signal regulated kinase (ERK)1/2 and Akt. Furthermore, the oral administration of KPS-A led to substantial inhibition of tumor growth and the expression of proliferating cell nuclear antigen (PCNA), MMP-9, tissue inhibitor of metalloproteinase-1 (TIMP-1), HuR, and Rab1A in the tumor tissues of mice inoculated with YD-10B OSCC cells. Collectively, KPS-A inhibits the invasiveness of oral cancer by reducing HuR-mediated MMP-9 mRNA stability and Rab1A-mediated MMP-9 secretion via ERK1/2 and phosphatidylinositide 3-kinase (PI3K)/Akt. Therefore, KPS-A is a promising anti-invasive agent.
  • Mika Ikehata, Kumiko Ueda, Seigo Iwakawa
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 301-307
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    The purpose of this study on the involvement of epigenetic control of the expression of solute carrier (SLC) transporters by DNA methylation and histone deacetylation in 4 colon cancer cells is to find the epigenetic control mechanisms of drug transporters in colon cancers. Human colon cancer cell lines (HCT116, HT29, SW48, SW480) were treated with 5-aza-2′-deoxycytidine (DAC), as a DNA methyltransferase inhibitor, followed by trichostatin A (TSA), as a histone deacetylase inhibitor. The mRNA expression and DNA methylation of several SLC transporters were analyzed by real-time polymerase chain reaction (PCR) and methylation-specific PCR, respectively. Among 12 SLC transporters possessing cytosine-phosphate-guanine (CpG) islands, thiamine transporter 2 (THTR2) (SLC19A3) gene showed a correlation between its mRNA expression level and DNA methylation status. TSA treatment increased histone H3 acetylation of THTR2 promoter region in all 4 colon cancer cell lines examined. HCT116 and SW48 cells showed a lack of THTR2 mRNA expression and methylation of its promoter, and DAC treatment induced its re-expression. In addition, the co-treatment with DAC and TSA increased THTR2 mRNA expression more markedly than DAC treatment in HCT116 and SW48 cells. In HT29 and SW480 cells that showed little methylation of THTR2 promoter, TSA treatment induced THTR2 mRNA expression markedly, but DAC treatment did not. In the 4 colon cancer cells examined, THTR2 mRNA expression is down-regulated by DNA methylation and/or histone deacetylation.
  • Wenjun He, Lingling Ye, Shichong Li, Hong Liu, Qiwei Wang, Xiaobing Fu ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 308-316
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Embryonic stem cells (ESCs) can propagate unlimitedly in vitro and differentiate into cardiomyocytes, which have been proposed as unlimited cell sources for cardiac cell therapy. This was limited by difficulties in large-scale generation of pure cardiomyocytes. In this study, we used stirred bioreactors to optimize the differentiation condition for mass production of embryoid bodies (EBs) derived from genetically modified mouse ESCs. Stirred suspension culture could more efficiently produce EBs and have a more uniform EB population without large necrotic centers, compared with the conventional static culture. Importantly, the cardiac-specific gene expressions (GATA binding protein 4, α-cardiac myosin heavy chain and myosin light chain-2v) were increased within EBs cultured in stirred bioreactor. Stirred suspension culture significantly increased the proportion of spontaneously contracting EBs, yielded a greater percentage of α-sarcomeric actinin-positive cells detected via flow cytometry, and harvested relatively more cardiomyocytes after G418 selection. Stirred suspension culture provided a more ideal culture condition facilitating the growth of EBs and enhancing the cardiogenic differentiation of genetically modified ESCs, which may be valuable in large-scale generation of pure cardiomyocytes.
  • Naoki Matsuzawa, Katsunori Nakamura, Masayuki Matsuda, Fumihiro Ishida ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 317-320
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Purpose: Cytochrome P450 (CYP)2C19 polymorphisms may partly explain the variability of thalidomide concentration and adverse drug effects by altering its metabolism. To compare the genetic and clinical factors responsible for the adverse effects and efficacy of thalidomide treatment, we investigated CYP2C19 genetic polymorphisms in Japanese subjects. Materials and Methods: Variations in the CYP2C19 gene in 6 patients treated with thalidomide were analyzed. The dosage of thalidomide, concentrations of (R)- and (S)-thalidomide in whole blood, and clinical laboratory test results were used as pharmacokinetic and pharmacodynamic indices. Using genomic DNA, CYP2C19*2 and *3 allele frequencies were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assays. Results: The frequencies of CYP2C19 PM and hetero EM (hetEM) genotypes in Japanese patients taking thalidomide were 2 (33.3%) and 4 (66.7%), respectively. The areas under the curve (AUC) of (R)-thalidomide were 3.42 and 5.33 μg·h/L, and those of (S)-thalidomide were 1.64 and 2.46 μg·h/L for hetEM and PM, respectively. Conclusions: This study provided new insights regarding the contribution of CYP2C19 gene variations to adverse responses to thalidomide. Genotyping of CYP2C19*2 and *3 can be considerably simplified by using KOD FX as a polymerase for prediction of adverse effects to thalidomide by the PCR-RFLP method. CYP2C19 PM patients tend to have high serum thalidomide concentrations.
  • Mamoru Naruse, Ken-ichi Ogawara, Toshikiro Kimura, Ryoji Konishi, Kazu ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 321-328
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    The aim of this study was to investigate the feasibility of percutaneous absorption of CNS5161, a novel N-methyl-D-aspartate (NMDA) receptor antagonist developed as a potential treatment for neuropathic pain and other neurological disorders. Six pressure-sensitive adhesives (PSA) with different physicochemical properties, namely, styrene–isoprene–styrene (1) (SIS(1)), styrene–isoprene–styrene (2) (SIS(2)), silicone, acrylate with a hydroxyl group (acrylate(OH)), acrylate without a functional group (acrylate(none)) and acrylate with a carboxyl group (acrylate(COOH)), were investigated for their release of CNS5161 and its subsequent skin permeability. Among the adhesives examined, silicone PSA provided the highest value of transdermal flux of CNS5161, which could be attributable to the highest release rate from it due to its very high thermodynamic activity. Although CNS5161 was also in the supersaturated state in SIS(1) and SIS(2) PSAs, the release and transdermal permeation from these adhesives were slower than those from silicone PSA. As for the acrylic PSAs, the highest release rate and permeability of CNS5161 were observed for acrylate(OH) PSA, followed by acrylate(none) and acrylate(COOH) PSAs, but none of them was better in terms of either the release or the permeability of CNS5161 than silicone PSA. These results clearly indicated that silicone PSA would be the most suitable for transdermal delivery of CNS5161 and silicone PSA containing 10% CNS5161 would be suitable for clinical use in humans.
  • Katsuyoshi Sunaga, Kenichi Ohkawa, Kenichi Nakamura, Atsuko Ohkubo, So ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 329-334
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    This study investigates whether tomato juice can inhibit cytochrome P450 (CYP) 3A4-mediated drug metabolism. Three commercially available, additive-free tomato juices, along with homogenized fresh tomato, were analyzed for their ability to inhibit testosterone 6β-hydroxylation activity using human recombinant CYP3A4. Results were compared to that of grapefruit juice. Ethyl acetate extracts of the tomato juices moderately reduced residual activity of CYP3A4 testosterone 6β-hydroxylation activity by 19.3—26.2% with 0-min preincubation. Residual activity was strongly reduced by 69.9—83.5% at 20-min preincubation, a reduction similar to that of grapefruit juice extract, known to contain constituents of mechanism-based inhibitors. One juice extract (tomato juice C) showed irreversible dose- and preincubation time-dependent and partial nicotinamide adenine dinucleotide phosphate (NADPH)-dependent inhibition of CYP3A4 activity. Furthermore, we examined whether the CYP3A4 inhibitory effect of tomato juice was substrate dependent by examining midazolam 1′-hydroxylation activity and nifedipine oxidation activity, in addition to testosterone 6β-hydroxylation activity. Tomato juice showed a potent inhibitory effect on nifedipine oxidation activity, which was comparable to that on testosterone 6β-hydroxylation activity; however, it showed a weak inhibitory effect on midazolam 1′-hydroxylation activity. We conclude that tomato juice contains one or more mechanism-based and competitive inhibitor(s) of CYP3A4. Additionally, significant CYP3A4 inhibitory activity did not result from lycopene, a major compound in tomato. Although the active compound was uncertain, a strong CYP3A4 inhibitory activity was observed in other solanaceous plants, i.e., potato, eggplant, sweet pepper, and capsicum. Therefore, responsible compounds in tomato are likely commonly shared among solanaceous vegetables.
  • Shanshan Zhao, Lan Yuan, Jiancheng Wang, Xuan Zhang, Zhonggui He, Qian ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 335-345
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Although the cellular endocytosis or uptake research using confocal laser scanning microscopy (CLSM) and Transwell inserts was frequently reported in experimental cell research and pharmaceutical research, there is little report on cellular transport process based on the same techniques. One of main reasons is that fluorescence of most fluorescent reagents could be clearly and definitely seen under CLSM only when they are internalized into and concentrated within cells. Here, Madin-Darby Canine Kidney (MDCK) cells and Coumarin 6 labeled nanoparticles (C6-NPs) was used as models, a new system was developed to image C6-NPs transport across Transwell filter grown cell monolayer by adding free cells into the basolateral medium and making the Transwell insert semi-permeable membrane visible under CLSM. The transport process could be clearly imaged in real-time and in situ, based on the visualization of Transwell membrane indicating the relative position of cells and nanoparticles, and the free cells in the basolateral medium serving as collector and indicator for the transported nanoparticles. The method was also applied in cell migration study. We believe that the novel approach developed here will be certainly useful not only in exploration of nanoparticle cellular transport mechanism, but also in other cell biological sciences based on CLSM and Transwell.
  • Satoshi Numazawa, Makiko Takase, Tomomi Ahiko, Masakazu Ishii, Shun-ic ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 346-354
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Endogenously produced reactive oxygen species reportedly stimulate insulin secretion from islet β-cells. However, the molecular machinery that governs the oxidant-induced insulin secretion has yet to be determined. The present study demonstrates, using rat islet β-cell-derived RINm5F cells, the involvement of the transient receptor potential (TRP) cation channels in the insulin secretion induced by the lipid peroxidation product 4-hydroxy-2-nonenal. Short-term (1 h) exposure of 4-hydroxy-2-nonenal induced a transient increase in intracellular Ca2+ concentration and subsequent insulin secretion in a concentration-dependent manner. The increase in intracellular Ca2+ concentration seemed to be due to an influx through the L-type voltage-dependent Ca2+ channel, since it was not observed when extracellular Ca2+ was absent and was inhibited almost completely by diltiazem or nifedipine. Ruthenium red, a non-specific inhibitor of TRP channels, inhibited the Ca2+ influx and insulin secretion evoked by 4-hydroxy-2-nonenal. Among the TRP channels, TRPA1 was found to be predominantly expressed, not only in RINm5F cells, but also rat islets. TRPA1 agonists, allylisothiocyanate and 15-deoxy-Δ12,14-prostaglandin J2, significantly induced Ca2+ influx, and a specific inhibitor TRPA1, HC-030031, blocked the effects elicited by 4-hydroxy-2-nonenal. These results suggest that 4-hydroxy-2-nonenal induces Ca2+ influx via the activation of TRP channels, including TRPA1, which appears to be coupled with the L-type voltage-dependent Ca2+ channel, and ultimately insulin secretion in RINm5F cells.
  • Yilu Ye, Chunyan Huang, Lili Jiang, Xiangdi Shen, Shanyong Zhu, Yan Ra ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 355-361
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Huanglian-Jie-Du-Tang (HJDT) is a traditional Chinese herbal formula which is widely used clinically. In this study, we investigated the effects of an aqueous (HJDTaq) and an ethanolic (HJDTet) extract of HJDT on chronic brain injury after focal cerebral ischemia in mice. The ischemia was induced by occlusion of the right middle cerebral artery for 30 min. HJDTaq (4 g/kg) and HJDTet (200, 400, 800 mg/kg) were orally administered for 21 d from day 7 before ischemia to day 14 after ischemia. The survival rate decreased to less than 50% at 35 d after ischemia. HJDTet at 400 mg/kg increased the survival rate. HJDTaq (4 g/kg) and HJDTet (400, 800 mg/kg) significantly attenuated the neurological dysfunction, brain atrophy and infarct volume after ischemia. There were few cells positive for CD31, hypoxia-inducible-factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and Flk-1 in the sham control. After ischemia, the number increased. HJDTaq (4 g/kg) and HJDTet (400 or 800 mg/kg) further increased the numbers of CD31, HIF-1α, VEGF and Flk-1-positive cells in the ischemic hemisphere. We conclude that HJDTaq and HJDTet have neuroprotective effects on chronic brain injury after focal cerebral ischemia and lead to accelerated angiogenesis by HIF-1α-regulated VEGF signaling.
  • Tomohiro Hikima, Noriaki Kaneda, Kyouhei Matsuo, Kakuji Tojo
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 362-368
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    The objective of this study is to establish a relationship of the skin penetration parameters between the three-dimensional cultured human epidermis LabCyte EPI-MODEL (LabCyte) and hairless mouse (HLM) skin penetration in vitro and to predict the skin penetration and plasma concentration profile in human. The skin penetration experiments through LabCyte and HLM skin were investigated using 19 drugs that have a different molecular weight and lipophilicity. The penetration flux for LabCyte reached 30 times larger at maximum than that for HLM skin. The human data can be estimated from the in silico approach with the diffusion coefficient (D), the partition coefficient (K) and the skin surface concentration (C) of drugs by assuming the bi-layer skin model for both LabCyte and HLM skin. The human skin penetration of β-estradiol, prednisolone, testosterone and ethynylestradiol was well agreed between the simulated profiles and in vitro experimental data. Plasma concentration profiles of β-estradiol in human were also simulated and well agreed with the clinical data. The present alternative method may decrease human or animal skin experiment for in vitro skin penetration.
  • In-uk Koh, Hye-Seung Jun, Joo Sun Choi, Joo Hyun Lim, Won Ho Kim, Jong ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 369-375
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Fenretinide (FEN), a ligand of retinol binding protein 4 (RBP4), has been suggested as a measure to reduce insulin resistance and its associated disorders such as obesity, and fatty liver by reducing serum RBP4. We investigated whether there is another possible mechanism by which fenretinide reduces insulin resistance and fatty liver in genetically obese (ob/ob) mice. Male obese mice fed a high-fat diet (45% of calories from fat) were divided into two groups (n=13 each). One (FEN) received fenretinide (20 mg/kg body weight, intraperitoneally) and the other (O) received vehicle three times weekly for 24 d. C57BL/6J mice fed a normal-fat diet (16% of calories from fat) were used as a control (C; n=13). No changes in fat weight and serum leptin level could be observed in FEN mice. Lower plasma RBP4 was observed in FEN mice compared with O mice. Fenretinide improved whole-body insulin sensitivity based on glucose and insulin tolerance tests and the homeostasis model assessment of insulin resistance. Fenretinide decreased the plasma lipid (triglyceride, cholesterol, and free-fatty acid) levels, hepatic TG level, and histological steatosis score. The mechanism by which fenretinide prevents fatty liver may be explained by an increased plasma adiponectin level, increased activation of hepatic AMP-activated protein kinase, and the expression of peroxisome proliferator-activated protein-α and peroxisomal acyl-CoA oxidase, which promote fat oxidation. FEN alleviated insulin resistance and fatty liver in obese mice and thus may act as an anti-lipidemic and anti-diabetic drug.
  • Nobuaki Egashira, Ryoko Okuno, Atsunori Shirakawa, Masaki Nagao, Kenic ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 376-379
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    We examined the role of 5-hydroxytryptamine2C (5-HT2C) receptors in marble-burying behavior in mice. When administered alone, the selective 5-HT2C agonist WAY161503 (3 mg/kg) inhibited marble-burying behavior. Moreover, the selective 5-HT2C antagonist SB242084 (3 mg/kg) reversed the inhibition of marble-burying behavior by 2,5-dimethoxy-4-iodoamphetamine (DOI) (1 mg/kg) or WAY161503 (3 mg/kg). Similarly, SB242084 (1 mg/kg) reversed the inhibition of marble-burying behavior by fluvoxamine (30 mg/kg) or paroxetine (3 mg/kg). These results suggest that 5-HT2C receptors play a role in marble-burying behavior in mice.
  • Masae Ohno, Mayu Fujita, Makoto Nishizuka, Shigehiro Osada, Masayoshi ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 380-384
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    We previously identified Ku proteins and interleukin enhancer binding factor 3 (ILF3) as cofactors for the nuclear receptor farnesoid X receptor and liver receptor homolog-1, respectively. Here we provide further evidence that these cofactors modulate the promoter activity of the nuclear receptor thyroid hormone receptor (TR) target gene, thyroid-stimulating hormone alpha (TSHα), which is negatively regulated by the TR ligand triiodothyronine (T3). Ku proteins suppressed TSHα promoter activity independent of T3, whereas ILF3 enhanced TSHα activity, especially in the presence of T3. Taken together, our results suggest that Ku proteins and ILF3 function as co-regulators for TR-mediated TSHα expression.
  • Kenzo Ohtsuki, Sayaka Miyai, Akira Yamaguchi, Kouhei Morikawa, Tetsuro ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 385-393
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    The stimulatory and inhibitory effects of several compounds and lignans isolated from the water extract of Taxus yunnanensis on the phosphorylation of three functional brain proteins (bovine myelin basic protein (bMBP), recombinant human tau protein (rhTP) and rat collapsin response mediator protein-2 (rCRMP-2)) by glycogen synthase kinase-3β (GSK-3β) were quantitatively compared in vitro, using (−)-epigallocatechin-3-gallate [(−)EGCG] as a positive control. We found that (i) three selected Taxus lignans [(3S,4R)-4′-hydroxy-6,3′-dimethoxyisoflavan-4-ol,(7R)-7-hydroxytaxiresinol and tanegool] highly stimulated the autophosphorylation of GSK-3β and the GSK-3β-mediated phosphorylation of two basic brain proteins [bMBP (pI=11.3) and rhTP (pI=8.2)], but inhibited dose-dependently the phosphorylation of an acidic protein (rCRMP-2, pI=6.0) by the kinase; (ii) these three Taxus lignans showed binding-affinities with bMBP as well as rhTP, but had low affinities with rCRMP-2; (iii) the binding of tanegool and (7R)-7-hydroxytaxiresinol to these two basic proteins induced their novel potent phosphorylation sites for GSK-3β; and (iv) these three Taxus lignans, but not EGCG, induced Tyr-phosphorylation of GSK-3β in vitro. These results provided here suggest that (i) these three Taxus lignans act as novel effective activators for GSK-3β and the GSK-3β-mediated phosphorylation of their binding basic proteins (rhTP and bMBP); and (ii) tanegool (IC50=1 μM) is an effective inhibitor for the phosphorylation of rCRMP-2 by the kinase in vitro.
  • Songhee Han, Seunghye Choi, Young-Jin Chun, Chul-Ho Yun, Chang Hoon Le ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 394-399
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Cytochrome P450 2A6 (CYP2A6) catalyzes important metabolic reactions of many xenobiotic compounds, including coumarin, nicotine, cotinine, and clinical drugs. Genetic polymorphisms of CYP2A6 can influence its metabolic activities. This study analyzed the functional activities of six CYP2A6 allelic variants (CYP2A6*5, *7, *8, *18, *19, and *35) containing nonsynonymous single-nucleotide polymorphisms. Recombinant variant enzymes of CYP2A6*7, *8, *18, *19, and *35 were successfully expressed in Escherichia coli and purified. However, a P450 holoenzyme spectrum was not detected for the CYP2A6*5 allelic variant (G479V). Structural analysis shows that the G479V mutation may alter the interaction between the A helix and the F—G helices. Enzyme kinetic analyses indicated that the effects of mutations in CYP2A6 allelic variants on drug metabolism are dependent on the substrates. In the case of coumarin 7-hydroxylation, CYP2A6*8 and *35 displayed increased Km values whereas CYP2A6*18 and *19 showed decreased kcat values, which resulted in lower catalytic efficiencies (kcat/Km). In the case of nicotine 5-oxidation, the CYP2A6*19 variant exhibited an increased Km value, whereas CYP2A6*18 and *35 showed much greater decreases in kcat values. These results suggest that individuals carrying these allelic variants are likely to have different metabolisms for different CYP2A6 substrates. Functional characterization of these allelic variants of CYP2A6 can help determine the importance of CYP2A6 polymorphisms in the metabolism of many clinical drugs.
  • Yohei Kawano, Masashi Nagata, Takafumi Kohno, Akihiro Ichimiya, Tomomi ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 400-407
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Caffeine is thought to increase the antitumor effect of cisplatin or DNA-damaging agents because it is known that caffeine inhibits DNA repair. Caffeine-assisted chemotherapy has been used in the treatment of osteosarcomas. In addition, there are several reports about combination chemotherapy with caffeine for certain malignancies other than osteosarcomas. However, there are no reports that show the utility of combination chemotherapy with caffeine for hepatocellular carcinoma (HCC). We examined the combined effects of caffeine and cisplatin in human HCC cell lines, and screened for a more effective administration method of caffeine in vitro. Human HCC cell lines (HepG2, HLF, HuH-7, and Li-7) were exposed to caffeine (0—0.5 mM) and cisplatin (0—1.2 μg/mL) for 72 h, either alone or in combination. Cell numbers were measured by WST-8 assay, and cell apoptosis was determined by annexin V-fluorescein isothiocyanate (FITC)/propidium iodide (PI) binding assay. As a result, caffeine increased the antitumor effect of cisplatin on cell proliferation and cell apoptosis in the HCC cell lines. Moreover, this effect was dependent on the amount of exposure to caffeine. These results suggest that caffeine-assisted chemotherapy is useful for HCC treatment.
  • Hirotsugu Takano, Kazuyuki Furuta, Kazuhito Yamashita, Mariko Sakanaka ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 408-412
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Appropriate culture models for tissue mast cells are required to determine how they are involved in regulation of local immune responses. We previously established a culture model for cutaneous mast cells, in which bone marrow-derived immature mast cells were co-cultured with Swiss 3T3 fibroblasts in the presence of stem cell factor. In this study, we focused on the roles of hyaluronan, which is produced by the feeder fibroblasts and forms the extracellular matrix during the co-culture period. Hyaluronan synthesis was found to be mediated by hyaluronan synthase 2 (HAS2) expressed in Swiss 3T3 cells. A decreases in the amount of hyaluronan, which was achieved by retroviral expression of short hairpin RNA for Has2 or by addition of hyaluronidase, significantly enhanced the proliferation of the cultured mast cells without any obvious effects on their maturation. Although we previously demonstrated that CD44 is required for proliferation of cutaneous mast cells, the deficiency of hyaluronan did not affect the proliferation of the cultured mast cells that lack CD44. These findings suggest that the extracellular matrix containing hyaluronan may have a potential to restrict proliferation of cutaneous mast cells in a CD44-independent manner.
  • Toshifumi Shiraga, Kanako Yajima, Takuya Teragaki, Katsuhiro Suzuki, T ...
    原稿種別: Regular Article
    2012 年 35 巻 3 号 p. 413-421
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Darexaban maleate is a novel oral direct factor Xa inhibitor. Darexaban glucuronide (YM-222714) was the major component in plasma after oral administration of darexaban to humans and is the pharmacologically active metabolite. Additionally, YM-222714 N-oxides were detected as minor metabolites in human plasma and urine. It is possible that YM-222714 N-oxides are formed by the N-oxidation of YM-222714 and/or the glucuronidation of darexaban N-oxides (YM-542845) in vivo. The former reaction is the pharmacological inactivation process. In this study, we identified the human enzymes responsible for YM-222714 N-oxidation and the uridine 5′-diphosphate (UDP)-glucuronosyltransferase (UGT) isoforms involved in YM-542845 glucuronidation in vitro. YM-222714 N-oxidation activity was detected in human liver microsomes (HLM), but not in human intestinal microsomes. In HLM, YM-222714 N-oxidation activities were significantly correlated with flavin-containing monooxygenase (FMO) marker enzyme activities (p<0.001) and inhibited by methimazole, a typical inhibitor of FMOs. Recombinant human FMO3 and FMO1 were capable of efficiently catalyzing YM-222714 N-oxidation, but not FMO5 or any recombinant human cytochrome P450 (CYP) isoforms. Considering the mRNA expression levels of FMO isoforms in human liver, these results strongly suggest that YM-222714 N-oxidation in HLM is mainly catalyzed by FMO3. In HLM, YM-542845 glucuronidation was strongly inhibited by typical substrates for UGT1A8, UGT1A9, and UGT1A10. Recombinant human UGT1A7, UGT1A8, UGT1A9, and UGT1A10 were capable of catalyzing YM-542845 glucuronidation, and UGT1A9 exhibited the highest intrinsic clearance. Considered together with the expression levels of UGT isoforms in human liver, these results strongly suggest that YM-542845 glucuronidation in HLM is mainly catalyzed by UGT1A9.
Notes
  • Tomoaki Kawaguchi, Patricia Naomi Nanbu, Mihoko Kurokawa
    原稿種別: Note
    2012 年 35 巻 3 号 p. 422-427
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Prolylhydroxyproline (Pro-Hyp), which is derived from collagen hydrolysate, has been shown to be beneficial for skin and joint health. However, little is known about the distribution of Pro-Hyp in these tissues. In the present study, we investigated the biodistribution of orally administered [14C]Pro-Hyp in rats. Whole-body autoradiography at 30 min after administration of [14C]Pro-Hyp showed that radioactivity is widely distributed in tissues including skin and articular cartilage, with the highest level of radioactivity observed in the gastric and intestinal walls. Incorporation of radioactivity into cells known to respond to Pro-Hyp such as dermal fibroblasts, synovial cells, chondrocytes, osteoblasts, and osteoclasts was observed. The chemical form of [14C]Pro-Hyp-derived radioactivity detected in the tissues was investigated by thin layer chromatography. The radioactive constituents in cartilage extract were two proline-modified peptides (56%), intact Pro-Hyp (5%), and two nonpeptide metabolites (28%). Similar results were obtained for skin and bone marrow. Plasma analysis at 3 to 30 min post-dose suggested that the majority of Pro-Hyp is modified in its proline residue by a first-pass effect without peptide bond hydrolysis. In conclusion, we demonstrated that Pro-Hyp is partly distributed in observed tissues including skin and cartilage in its intact form, which might be responsible for its biological functions.
  • Sooryun Lee, Soon-Chun Chung, So-Hyoung Lee, Wanki Park, Ikhoon Oh, Wo ...
    原稿種別: Note
    2012 年 35 巻 3 号 p. 428-432
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Na+/K+-adenosine triphosphatase (ATPase) inhibitors have considerable therapeutic potential against some heart diseases like congestive heart failure and cardiac arrhythmias. Through bioassay-guided separation of the leaf extract of Laurus nobilis, six acylated kaempferol glycosides (compounds 16) were isolated. Their structures were determined on the basis of spectroscopic analysis and comparison with reported data. All the isolates were subjected to in vitro bioassays to evaluate their inhibitory activities against Na+/K+-ATPase from porcine cerebral cortex and bacterial growth. These studies led to the identification of compounds 16 as potent Na+/K+-ATPase inhibitors, with IC50 values in the range of 4.0±0.1—10.4±0.6 μM. These compounds also exhibited a broad spectrum of antibacterial activity. In particular, compounds 4 and 6 showed potent inhibitory activities against several bacterial strains, except Escherichia coli, with minimum inhibitory concentration (MIC) values in the range of 0.65—2.08 μg/mL. Thus, L. nobilis-derived acylated kaempferol glycosides may have a potential to be leads for the development of Na+/K+ ATPase inhibitors (16) and antibacterial agents (4, 6).
  • Toshimitsu Okamura, Tatsuya Kikuchi, Mika Nodaira, Kenichi Odaka, Kiyo ...
    原稿種別: Note
    2012 年 35 巻 3 号 p. 433-437
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    Phosphorylation of tyrosine residues by protein tyrosine kinases (PTK) and phosphotyrosine/Src homology 2 (SH2) domain interactions are crucial not only for signal transduction but also for regulation of PTK activity. Tyrosine residues also receive nitration and halogenation under oxidative conditions. It has been reported that nitration of tyrosine residue caused peptides to be a poor substrate for PTK and that nitrotyrosine residues could bind to SH2 domains as a phosphotyrosine mimic to activate Src family kinase. However, the effect of halogenation on tyrosine phosphorylation or SH2 domain binding is not well understood. We examined the phosphorylation of model peptides containing 3-halotyrosine or 3-nitrotyrosine using typical receptor tyrosine kinase, epidermal growth factor receptor (EGFR), and nonreceptor tyrosine kinase, lymphocyte-specific protein tyrosine kinase (Lck). The EGFR- and Lck-mediated phosphorylation was markedly inhibited by tyrosine halogenation. Iodination showed the strongest inhibition of the phosphorylation among four types of halogenation, and its inhibitory effect was stronger than that of nitration. We also examined the effect of iodination and nitration of tyrosine residues on binding to the SH2 domain of Lck, using a model peptide containing the phosphoTyr-Glu-Glu-Ile motif, which has a high affinity for the SH2 domain. The relative affinities of the modified peptides whose phosphotyrosine was substituted with unphosphorylated tyrosine, 3-nitrotyrosine, and 3-iodotyrosine, and of the model peptide were 0.024, 0.26, 1, and 16, respectively. These results suggest that tyrosine iodination may have an effect on the phosphorylation or binding to the SH2 domain similar to nitration. Tyrosine iodination possibly modulates signal transduction, with the potential impairment of cell function.
  • Satoshi Takagi, Toshihiro Miura, Takenori Yamashita, Naoki Ando, Haruk ...
    原稿種別: Note
    2012 年 35 巻 3 号 p. 438-443
    発行日: 2012/03/01
    公開日: 2012/03/01
    ジャーナル フリー
    We examined the bone mineral density (BMD) of the proximal region and the mid-diaphysis of the femur using dual energy X-ray absorption (DXA), the blood osteocalcin level and the blood glucose level every five weeks from 8 to 23 weeks old in KK-Ay diabetic mice. The BMD of the proximal region after 18 weeks old was significantly lower when compared with that at 8 weeks old (p<0.05), whereas there was no significant difference in the BMD of the mid-diaphysis at each week. The BMD of the proximal region at 18 weeks old was significantly lower than that in ddY mice, used as controls (p<0.05). The blood osteocalcin level at 18 weeks old was significantly lower than that at 8 weeks old and that in 18-week-old ddY mice (p<0.05). There was significant negative correlation between the blood glucose level and the BMD of the proximal region (r=−0.64, p<0.05). These results suggest that type 2 diabetes exerts an influence only on spongy bone, not on cortical bone, and that the BMD in the proximal region of the femur seems to be affected by blood glucose level, parallel with the progression of diabetes, through the blood osteocalcin level. In the present study, we show the characteristics of diabetic osteopenia in KK-Ay mice, an animal model of type 2 diabetes.
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