Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
Volume 32 , Issue 9
Showing 1-29 articles out of 29 articles from the selected issue
Analytical Biochemistry
Regular Articles
  • Takeshi Nagata, Asako Nakamura, Toshifumi Akizawa, Hidemitsu Pan-Hou
    Type: Regular Articles
    Subject area: Analytical Biochemistry
    2009 Volume 32 Issue 9 Pages 1491-1495
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    To further enhance the efficiency and potential of plants for phytoremediation of mercury pollution, a genetically engineered tobacco to simultaneously express mercury transporter, mercury transporter (MerT) and mercury chelator, polyphosphate (polyP) was constructed by integrating bacterial merT gene in polyphosphate kinase gene (ppk)-transgenic tobacco, and its ability to phytoremediate mercury was evaluated. Integration of merT gene into ppk-transgenic tobacco did not significantly affect the mercury resistant phenotypes and polyP production. Transgenic expression of MerT in ppk-transgenic tobacco resulted in accelerated and enhanced mercury uptake into tobacco. In addition, tobacco expressing MerT and polyP accumulated significantly more mercury than the ppk-transgenic tobacco from medium containing a wide range of low concentrations of Hg2+. The combination of accelerated mercury uptake and enhanced mercury accumulation mediated by MerT represents one way for shortening the purification completion time, and for improving tobacco plants to be more suitable for use in phytoremediation of low levels of mercury contamination.
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  • Wei Shen, Jiaming Zhang, Genjun Mao, Keyi Jiang, Qing Zhu
    Type: Regular Articles
    Subject area: Analytical Biochemistry
    2009 Volume 32 Issue 9 Pages 1496-1499
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Epoxide hydrolase plays an important role in the detoxification of genotoxic compounds and in the control of physiological signaling molecules. Altered levels of epoxide hydrolase activity are associated with many diseases, such as emphysema, lung cancer, ovarian cancer, and laryngeal carcinoma. We designed and synthesized a resorufin-based fluorogenic probe, 7-(2-(oxiran-2-yl)ethoxy) resorufin, which was hydrolyzed by microsomal epoxide hydrolase to form the corresponding diol, which upon further treatment with sodium periodate released the strongly fluorescent resorufin. The probe exhibits good biological compatibility and photophysical properties, such as long wavelength excitation (571 nm) and emission (585 nm) and a wide working pH range (from 6.0 to 10.0), and thus facilitates the determination of the activity of microsomal epoxide hydrolase.
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Notes
  • Setsuko Tojo, Akira Okazaki, Masako Wakitani, Toyohide Shinkawa, Kazuh ...
    Type: Notes
    Subject area: Analytical Biochemistry
    2009 Volume 32 Issue 9 Pages 1604-1608
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Recent studies have shown that antibodies with low fucose content in their oligosaccharides exhibit highly potent antibody-dependent cellular cytotoxicity (ADCC). However, composites of therapeutic antibodies produced by conventional production systems using cell lines such as Chinese hamster ovary (CHO) and SP2/0 do not necessarily contain sufficient amounts of non-fucosylated antibody species. In this study, we combined two lectin-affinity chromatography techniques, Concanavalin A and Lens culinaris agglutinin, to enrich the non-fucosylated species from therapeutic material using the anti-Her2/neu model antibody. Oligosaccharide analysis by matrix-assisted laser desorption/ionization-time of flight MS following peptide-N-glycosidase F digestion suggested that non-fucosylated antibody could be enriched in the purified fraction with efficient removal of high-mannose species. The ADCC activity of the purified fraction was about 100-fold higher than that of the initial material. The chromatographic strategy presented here can be a useful tool to elevate ADCC activity of antibody materials without concentrating high-mannose oligosaccharides.
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Biochemistry
Regular Articles
  • Meihua Jin, Ju Hye Yang, Eunkyung Lee, Yue Lu, Soonyoul Kwon, Kun Ho S ...
    Type: Regular Articles
    Subject area: Biochemistry
    2009 Volume 32 Issue 9 Pages 1500-1503
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Previously, we reported that an ethanol extract of Ailanthus altissima has antiinflammatory activity in an ovalbumin (OVA)-sensitized murine asthmatic model. To determine the biological compounds from this plant, luteolin-7-O-glucoside (L7G) was isolated and its antiasthmatic activity was evaluated in an in vivo murine asthmatic model. L7G (10 to 100 mg/kg, per os (p.o.)) reduced the amount of eosinophil infiltration in bronchoalveolar lavage (BAL) fluid in a dose-dependent manner. In comparison, dexamethasone (5 mg/kg, p.o.), which was used as a positive control, also strongly inhibited the number of infiltrating eosinophils. L7G inhibited both the prostaglandin E2 (PGE2) and serum immunoglobulin E level in BAL fluid in a dose-dependent manner. In addition, L7G inhibited the transcript profiles of interleukin (IL)-4, IL-5, and IL-13 mRNA expression levels in the murine asthma model, as determined using reverse transcription-polymerase chain reaction (RT-PCR). These results suggest that the antiasthmatic activity of L7G in OVA-induced lung inflammation may occur in part via the downregulation of T helper 2 cytokine transcripts as well as the inhibition of PGE2 production.
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  • Akemi Ryu, Kumi Arakane, Chiharu Koide, Hiroyuki Arai, Tetsuo Nagano
    Type: Regular Articles
    Subject area: Biochemistry
    2009 Volume 32 Issue 9 Pages 1504-1509
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Based on our previous finding (Biochem. Biophys. Res. Commun., 223, 578—582, 1996) of singlet oxygen generation from coproporphyrin excreted on the skin surface from Propionibacterium acnes, we hypothesized that singlet oxygen formed in this way under UV exposure would promote peroxidation of skin surface lipids. We found that squalene was oxidized efficiently by singlet oxygen derived from coproporphyrin under UV exposure, and that the rate constant of squalene peroxidation by singlet oxygen was ten-fold higher than that of other skin surface lipids examined. The reaction was promoted more efficiently by UVA than by UVB. Furthermore, we found that topical application of squalene peroxide induced skin hyperpigmentation through increasing prostaglandin E2 release from keratinocytes in guinea pigs. These results suggest that squalene peroxide formation by singlet oxygen plays a key role in photo-induced skin damage.
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Molecular and Cell Biology
Regular Articles
  • Mai Mukai, Kunihiro Hayakawa, Maro Okamura, Yasuhiro Tagawa, Shotaro N ...
    Type: Regular Articles
    Subject area: Molecular and Cell Biology
    2009 Volume 32 Issue 9 Pages 1510-1515
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Halogenated and polycyclic aromatic hydrocarbons are widely distributed pollutants in environments. These toxic substances activate the aryl hydrocarbon receptor (AhR) and thereby cause a broad spectrum of pathological changes. Development of AhR inhibitors will be useful for prevention of diseases caused by AhR activation. Using the dioxin responsive element (DRE)-based sensing via secreted alkaline phosphatase (DRESSA), we examined effects of Antrodia camphorata, a mycerial extract, on the activation of AhR by halogenated and polycyclic aromatic hydrocarbons. We found that Antrodia camphorata markedly suppressed activation of AhR triggered by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). In contrast, activation of AhR by polycyclic aromatic hydrocarbons (benzo[a]pyrene and 3-methylcholanthrene) was inhibited only modestly by this mycelium. Similarly, Antrodia camphorata only mildly attenuated activation of AhR by cigarette smoke that contains polycyclic aromatic hydrocarbons. Consistent with these results, Northern blot analysis revealed that DRE-driven exogenous and endogenous gene expression triggered by TCDD was abolished by Antrodia camphorata, whereas it did not substantially affect DRE-induced transcription triggered by benzo[a]pyrene, 3-methylcholanthrene or cigarette smoke. We also found that the inhibitory effect of Antrodia camphorata on TCDD-induced AhR activation was ascribed to neither down-regulation of AhR, down-regulation of the AhR nuclear translocator, nor up-regulation of the AhR repressor. These results suggest that Antrodia camphorata preferentially inhibits AhR activation and DRE-dependent gene expression triggered by dioxin.
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Notes
  • Kyoung-Chan Park, So-Young Kim, Dong-Seok Kim
    Type: Notes
    Subject area: Molecular and Cell Biology
    2009 Volume 32 Issue 9 Pages 1609-1613
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Recently, indole-3-acetic acid (IAA) has been introduced as a new cancer therapeutic agent through oxidative decarboxylation by horseradish peroxidase (HRP). The purpose of this study was to determine the therapeutic feasibility of IAA/light combination against liver cancer. SK-HEP-1 cells were irradiated with UVB or visible light (518 nm) in the presence of IAA. Cell viability was measured using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Then, IAA was injected in SK-HEP-1 liver cancer cell-implanted nude mice, and the tumor area was irradiated with intense pulsed light (IPL). Then, tissue was taken for terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assay and immunohistochemical staining for 8-hydroxy-deoxyguanosine (8-OHdG), p53, caspase-3, and proliferating cell nuclear antigen (PCNA). In vitro experiments demonstrated that IAA alone was not cytotoxic, but activated IAA by HRP or light caused cell death. In vivo experiments showed that IAA/IPL treatment caused regression of tumor cells in SK-HEP-1-implanted nude mice. The TUNEL assay showed that IAA/IPL induced cancer cell apoptosis, and this was confirmed by increases in 8-OHdG, p53, and caspase-3 in IAA/IPL-treated mice. In contrast, IPL alone did not induce apoptosis, indicating that the apoptotic effect resulted from activated IAA by light. In summary, we showed that IAA/light induced tumor regression in SK-HEP-1-implanted nude mice. These results suggest the potential use of IAA/light combination in liver cancer.
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Microbiology
Notes
  • Mari Kashiwamura, Kohta Kurohane, Takashi Tanikawa, Aya Deguchi, Daise ...
    Type: Notes
    Subject area: Microbiology
    2009 Volume 32 Issue 9 Pages 1614-1617
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Shiga toxins (Stxs) are major virulence factors produced by enterohemorrhagic Escherichia coli O157:H7 colonizing the human and cattle intestines. We previously demonstrated that recombinant binding subunits (Stx1B) bound to the mucosal epithelium of the distal but not that of the proximal part of the mouse colon. Here we developed a method for isolating colon epithelial cells from the proximal and distal parts separately. Enrichment of epithelial cells was confirmed by the expression of cytokeratin. There was no difference in the epithelial cell purity between the proximal and distal colon preparations. The isolated epithelial cells from the distal colon were found to display binding sites for recombinant Stx1B whereas those from the proximal colon were not. Taking advantage of this single cell isolation, we examined the effect of Stx1 holotoxin on the epithelial cells. Consistent with the expression of the binding sites, Stx1 induced apoptosis of the epithelial cells from the distal but not those from the proximal colon. The results provide direct evidence that mouse colon epithelial cells are susceptible to Stx1 toxicity corresponding to the expression of binding sites for toxins.
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Pharmacology
Regular Articles
  • Min Young Um, Ji Yun Ahn, Suna Kim, Mi Kyung Kim, Tae Youl Ha
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1516-1520
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    This study was designed to investigate the effect of sesaminol glycosides (SG), one of the most abundant lignan glycosides in sesame (Sesamum indicum LINN.) seed, on cognitive deficits and oxidative stress induced by intracerebroventricular (i.c.v.) injection of β-amyloid protein (Aβ)25—35 in mice. Mice were fed diets containing 0%, 0.25%, or 0.5% of SG for six weeks. Dietary SG showed a protective effect against Aβ-induced learning and memory deficits in passive avoidance and the Morris water maze test. Aβ caused significant neuronal loss in the CA1 and CA3 regions of the hippocampus, but SG supplement showed decrease of the Aβ25—35 induced neuronal loss. The SG supplementation significantly decreased thiobarbituric acid reactive substance values and 8-hydroxy-2′-deoxyguanosine (8-OHdG) levels in brain tissue. SG also reversed the activity of glutathione peroxidase (GPx), which is decreased by Aβ. These results suggest that SG protects against cognitive deficits induced by Aβ25—35, in part through its antioxidant activity.
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  • Xuelian Li, Weiming Chu, Jinling Liu, Xiaorong Xue, Yanjie Lu, Hongli ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1521-1526
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Matrine, a monomer of traditional Chinese medicine Sophora flavescens, is a potential drug for treatment of arrhythmia. The aim of the study is to elucidate the protective effects of matrine on arrhythmic rat induced by myocardial infarction (MI) and further explore underlying targets. Experiments were performed to investigate the effects of long-term oral administration of matrine on coronary ligation induced arrhythmia, measured in whole animals, via surface electrocardiogram (ECG). Whole-cell patch-clamp technique was used to record the action potential and potassium ionic currents in myocytes isolated from rat hearts. The cytoplasmic free Ca2+ concentration ([Ca2+]i) was measured using the scanning confocal microscopy. Mortality rate was 19/30 (63%) in MI group and 10/30 (33%) in matrine group (p<0.05). This represented a 1.9-fold reduction in long-term mortality rate. The prolonged action potential duration (APD) induced by MI were significantly shortened by long-term treatment of matrine. Matrine restored Kv4.2/Ito, Kir2.1/IK1 in rat ventricular myocytes after MI. Abnormaly decreased [Ca2+]i mediated by ischemia can be recovered by matrine. Our results suggested that long-term oral administration of matrine reduced arrhythmia and mortality. Electrophysiological experiment revealed that long-term matrine treatment played an important role in anti-arrhythmia through ionic mechanism. Knowledge of matrine from this work may provide insight into the development of new drugs for long-term myocardial infarction treatment.
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  • Phuong Thien Thuong, Yuba Raj Pokharel, Moo Yeol Lee, Sang Kyum Kim, K ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1527-1532
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Atherosclerosis is main cause of arteriosclerosis. The pivotal role of low-density lipoprotein (LDL) oxidation in atherogenesis suggests antioxidants may help prevent cardiovascular disease. Fraxinus rhynchophylla DENCE (Oleaceae) is a traditional medicinal plant from East Asia. During the course of characterizing potential drug candidates from natural products, we isolated two major coumarins, esculetin and fraxetin and found that fraxetin has dual-antioxidative functions. Low concentrations (1—5 μM) of fraxetin potently inhibited LDL oxidation induced by metal and free radicals. Moreover, treatment of vascular smooth muscle cells (VSMCs) with higher concentrations (above 30 μM) of fraxetin significantly increased the protein level of heme oxygenase-1 (HO-1), a key enzyme that inhibits vascular proliferation and atherosclerosis. Subcellular fractionation and reporter gene analysis using an antioxidant response element (ARE) construct revealed that fraxetin increased the level of nuclear factor (NF)-E2-related factor 2 (Nrf2) and reporter activity, and these were associated with the induction of antioxidant enzymes, such as HO-1 and glutathione S-transferase-α. In conclusion, fraxetin has direct protective properties against LDL oxidation at lower concentrations, and higher concentrations of fraxetin induce antioxidant enzymes via Nrf2/ARE activation. These effects suggest potential anti-atherosclerosis effects of Fraxinus rhynchophylla D.
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  • Xu Sun, Xiaodong Zhang, Hui Hu, Yina Lu, Jie Chen, Kazuki Yasuda, Heya ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1533-1537
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Proliferation of hepatic stellate cells (HSCs) is central for the development of fibrosis during liver injury. Our aim in this study was to determine whether berberine could inhibit HSC proliferation in vitro and prevent experimental liver fibrosis in vivo. Activated rat hepatic stellate cells (CFSCs) were incubated with various concentrations (0—20 μg/ml) of berberine. After 48 h incubation, berberine significantly inhibited CFSC proliferation and induced cell cycle arrest in G1 phase. Real-time and Western blotting revealed that both p21 and p27 expression was markedly reduced by berberine. Berberine also decreased Akt phosphorylation and FoxO1 phosphorylation, which led to FoxO1 nuclear translocation. Berberine effectively prevented CCl4-induced liver fibrosis in mice, which was accompanied by a decrease in the number of activated HSCs. Thus, berberine was able to prevent liver fibrosis by inhibition of hepatic stellate cell proliferation.
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  • Kazuhide Hayakawa, Keiichi Irie, Kazunori Sano, Takuya Watanabe, Sei H ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1538-1544
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Cannabidiol decreases cerebral infarction and high-mobility group box1 (HMGB1) in plasma in ischemic early phase. However, plasma HMGB1 levels in ischemic delayed phase reach higher concentration with the progressing brain injury. In this study, we investigated the therapeutic time window of cannabidiol on functional deficits, glial HMGB1 and plasma HMGB1 levels in a 4 h mouse middle cerebral artery (MCA) occlusion model. Cannabidiol-treated mice were divided into 3 groups as follows: group (a) treated from day 1, group (b) treated from day 3, group (c) treated from day 5 after MCA occlusion. Moreover, minocycline, microglia inhibitor, and fluorocitrate, an inhibitor of astroglial metabolism, were used to compare with cannabidiol-treated group. Repeated treatment with cannabidiol from 1 and 3 d at the latest after cerebral ischemia improved functional deficits and survival rates. However, cannabidiol from 5 d could not improve the ischemic damage as well as fluorocitrate-treated group. Moreover, both group (a), group (b) and minocycline but not group (c) and fluorocitrate-treated group had a decrease in the number of Iba1 expressing HMGB1 positive cells and HMGB1 levels in plasma. Cannabidiol may provide therapeutic possibilities for the progressing brain injury via HMGB1-inhibiting mechanism.
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  • Yiguo Sun, Yun Zhang, Ming Yan, Yabin Wu, Xiaoxiang Zheng
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1545-1551
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    B-type natriuretic peptide (BNP) is one peptide hormone released in response to myocyte stretch, whose functions play significant roles in health and disease. Its physiologic effects result in improved loading conditions and have led to the development of recombinant BNP as a therapeutic agent for heart failure. Previous work has identified that BNP protect myocardium against reperfusion injury through mitochondrial pathway. Mitochondria are both essential effectors of cardioprotection and primary targets of cardioprotective signaling. Their role during reperfusion is particularly critical because of the conditions that promote both apoptosis by the mitochondrial pathway and necrosis by irreversible damage to mitochondria in association with mitochondrial permeability transition pores (mPTP). After an episode of myocardial ischemia, opening of mPTP, at the onset of reperfusion, is a critical determinant of myocyte death. The relationship of BNP and mPTP in mediating reperfusion-induced cardiomyocytes injury is a novel investigative area. In this study, our results indicated that the beneficial effect of BNP in cultured cardiomyocytes subjected to reperfusion is associated with attenuation of mPTP opening, resultant mitochondrial dysfunction and apoptosis. Further investigation of underlying mechanisms revealed that these were associated with BNP-mediated repolarization of mitochondrial membrane potential (Δψm), inhibition of reactive oxygen species (ROS) generation, improvement of Bcl-2 level, and inhibition of Bax and second mitochondria-derived activator of caspases/direct inhibitor of apoptosis protein-binding protein with a low isoelectric point (Smac/DIABLO) levels. In summary, we demonstrate that BNP exerts protective actions within reperfusion by inhibiting mPTP opening and these roles of BNP may involve phosphatidylinositol 3-kinase (PI3K) dependent pathway.
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  • Jing-Tian Xie, Chong-Zhi Wang, Bin Zhang, Sangeeta Ram Mehendale, Xiao ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1552-1558
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Panax ginseng roots, including the steamed roots, have been demonstrated to possess anticancer properties. However, there have been limited published studies on the cancer preventive effects of American ginseng. In this study, the in vitro and in vivo anti-colorectal cancer effects of American ginseng berry extracts, and their representative bioactive compounds were evaluated. The ginsenoside content in unsteamed American ginseng berry extract (AGE) and steamed berry extract (S-AGE) were determined by HPLC. In comparison to AGE, S-AGE showed significantly stronger antiproliferative effects on HCT-116, SW-480 and HT-29 human colorectal cancer cells. Antiproliferative effects of representative constituents in AGE and S-AGE, ginsenosides Rb3 and Rg3, were also evaluated, showing that Rg3 had a positive effect. Using flow cytometric analyses, we found that S-AGE arrests cancer cells in G1-phase and significantly induces cell apoptosis. Using xenograft mice, we conducted an in vivo antitumor study using S-AGE after HCT-116 cell inoculation. We observed that 50 mg/kg of S-AGE showed significant antitumor effects. Our results suggested that S-AGE inhibited the colorectal cancer growth both in vitro and in vivo, and this inhibition might be achieved through cell cycle arrest and induced apoptosis in the cells.
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  • Nobuaki Mizutani, Jun-ichi Fuchikami, Maki Takahashi, Takeshi Nabe, Sh ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1559-1564
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Exposure of animals to cigarette smoke for longer than 3 months leads to the development of chronic obstructive pulmonary disease (COPD) showing pulmonary emphysema. We attempted to create a COPD model with emphysema that could be established in a shorter period of time. Guinea pigs were intratracheally treated once a day on days 0—3, 5—8, 10—13 and 15—18 with a cigarette smoke solution (CSS), which was prepared by bubbling a stream of smoke into saline. Additionally, lipopolysaccharide (LPS) was administered intratracheally as an exacerbation factor on days 4, 9 and 14. By day 19, there was a gradual elevation of specific airway resistance (sRaw). In addition, both residual volume and functional residual capacity were found to be significantly higher on day 19. In the lungs, there was a marked increase in leukocytes, especially neutrophils. Histologically, we observed epithelial hyperplasia and emphysema. On the other hand, daily oral administration of theophylline during the administration of CSS and LPS suppressed the sRaw increase and the epithelial hyperplasia, but not other functional structural changes. In conclusion, we established an experimental COPD model in guinea pigs by using intratracheal instillations of CSS and LPS over a considerably shorter term than has been reported for other models.
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  • Xin Chang Wang, Jian Feng, Feng Huang, Yong Sheng Fan, Yan Yan Wang, L ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1565-1570
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    The present study was performed to evaluate the potential protective effects of Shikonin extracted from Zicao on lupus nephritis (LN) using NZB/W F1 mice. Oral administration of Shikonin (24, 40 mg/kg body weight/d) or vehicle was applied to sixty female NZB/W F1 mice of 28-week-old with LN. Treatment with Shikonin for 14 weeks suppressed proteinuria dose-dependently with the mean proteinuria of 274.0 mg/dl and 160.3 mg/dl for low-dose and high-dose Shikonin groups, respectively, compared to 499.2 mg/dl for the vehicle. Also, Shikonin was observed to reduce circulating adhesion molecules significantly and down-regulate intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) mRNA expression in kidney. However, anti-double stranded (ds)DNA antibody in mice with low or high Shikonin dose administration both exhibited no significant elevation, differing from vehicle group. Kidney histological examination showed that renal glomerular lesions were alleviated after Shikonin application. These results suggest that Shikonin has therapeutic effects on LN in NZB/W F1 mice, to which inhibition of anti-dsDNA may be potential contribution, and its part mechanism is related to suppression of mRNA expression of cell adhesion molecules (CAMs) in the kidney.
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  • Takashi Okura, Tadahiro Ozawa, Michiko Ibe, Yuko Taki, Midori Kimura, ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1571-1575
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    The present study was conducted to investigate the effects of repeated treatment with morphine on the drug's antinociceptive effects, intestinal absorption, and transepithelial transport. The antinociceptive effects of morphine in rats were markedly decreased after repeated oral administration of the drug for 5 d, indicating the development of tolerance. In the morphine-tolerant rats, intestinal absorption of morphine was determined using the in situ loop method. Absorption of morphine from the jejunum was significantly decreased after repeated administration. The permeability of human intestinal epithelial Caco-2 cells was increased in the efflux direction after repeated treatment. The repeated administration of morphine also reduced the cellular accumulation and efflux of P-glycoprotein substrates ([3H]vincristine and rhodamine123) from Caco-2 cells, suggesting that it enhances P-glycoprotein-mediated efflux in Caco-2 cells. These results suggest that repeated use enhances the efflux of morphine in the epithelial cells of the small intestine, subsequently decreasing its intestinal absorption.
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  • Akiko Nakatani-Pawlak, Kazumasa Yamaguchi, Yoshimi Tatsumi, Hiroyuki M ...
    Type: Regular Articles
    Subject area: Pharmacology
    2009 Volume 32 Issue 9 Pages 1576-1583
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    We investigated the involvement of glutamic acid in neural development by injecting phencyclidine (PCP) into neonatal ICR mice. Neonatal mice were injected with PCP at 10 mg/kg or saline on postnatal days 7, 9 and 11, and their behavioral, anatomical and neurochemical changes were analyzed in adulthood. PCP-treated mice exhibited an increase in PCP-induced hyperactivity and impairments of spatial working memory and social interaction behavior. The impairment of social interaction behavior was significantly reversed by administration of clozapine, D-cycloserine, flumazenil, or SHC50911, a γ-aminobutyrate B (GABAB) receptor antagonist. A decrease in the number of parvalbumin-positive cells and spine density in the frontal cortex, nucleus accumbens and hippocampus were evident in the brains of PCP-treated mice. Measurement of brain monoamine and their metabolite contents in adulthood indicated brain area-dependent and neurotransmitter-specific changes in monoamine metabolism. These findings suggest that neonatal treatment with PCP in mice leads to enhanced sensitivity to PCP and impairment of spatial working memory and social interaction behaviors in adulthood, which may be associated with reduced spine density and GABAergic interneurons and changes in monoamine metabolism. Furthermore, pharmacologic experiments suggest the potential applicability of neonatally PCP-treated mice as a useful animal model for new antipsychotic drug screening.
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Medicinal Chemistry
Regular Articles
  • Yasuyuki Sadzuka, Makoto Matsuura, Takashi Sonobe
    Type: Regular Articles
    Subject area: Medicinal Chemistry
    2009 Volume 32 Issue 9 Pages 1584-1587
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Taurine is contained in seafood and has been studied extensively on life-style related diseases. Theanine increased the effects of the doxorubicin (DOX) as an antitumor agent in some tumors and enhanced the DOX level in tumor cells. It is expected that the advanced effect of food uptake in cancer chemotherapy may be effective from the viewpoint of quality of life (QOL) improvement, although this approach has not been investigated in detail. In this study, the effect of taurine as a functional amino acid was examined. Taurine did not change the DOX influx into M5076 cells, whereas it significantly inhibited DOX efflux, which maintained the DOX level in tumor cells. Furthermore, experiments with taurine decreased tumor weight by 40%, compared to the DOX-alone group and significantly increased its antitumor effect. Moreover, as taurine did not increase DOX concentration in normal tissue, it is suggested that it increased the antitumor effect without enhancing DOX-induced adverse effects. DOX efflux is inhibited by β-alanine as a taurine transporter inhibitor, therefore, enhancement of the DOX level by taurine was suggested to act via taurine transport. Namely, it was clarified that taurine was useful as a modulator to enhance the therapeutic index of cancer patients and improve QOL.
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Notes
Pharmacognosy
Notes
  • Ai Inoko, Nobuko Kakiuchi, Michiyo Yoshimitsu, Shaoqing Cai, Masayuki ...
    Type: Notes
    Subject area: Pharmacognosy
    2009 Volume 32 Issue 9 Pages 1621-1623
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Wild Ephedra plants growing near the Tibetan border of Yunnan and Sichuan Provinces and north-central Sichuan were surveyed and their DNA and ephedrine alkaloids content were analyzed. By analysis of internal transcribed spacer 1 (ITS) 1 DNA, E. likiangensis was found to be the dominant species in these regions, which clustered into 2 major groups in the phylogenic tree. Most Ephedra plants in these regions of ordinal size contained ephedrine and pseudoephedrine of more than 0.7%, the requirement for Japanese Pharmacopoeia 15th edition, suggesting that they have potential for crude drug production of Ephedra herbs.
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  • Lan-Lan Fan, Shu Zhu, Hu-Biao Chen, Dong-Hui Yang, Shao-Qing Cai, Kats ...
    Type: Notes
    Subject area: Pharmacognosy
    2009 Volume 32 Issue 9 Pages 1624-1627
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Stemona sessilifolia, S. japonica and S. tuberosa are the three genuine sources of Stemonae Radix specified in the Chinese Pharmacopoeia (CP) for antitussive and insecticidal remedy. Significant variations in alkaloids composition and content, as well as different degree of antitussive activity were found among them. In order to accurately identify the genuine sources of Stemonae Radix in the genetic level, two polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methods were developed based on the sequence differences in chloroplast DNA trnL-trnF and petB-petD regions of the species recorded in CP, as well as S. parviflora and a counterfeit of Stemonae Radix, Asparagus cochinchinensis. By using the restriction enzymes MwoI, AciI and XmnI which were able to recognize specific sequence sites in the trnL-trnF region, and BclI, HincII and BslI which can recognize those in the petB-petD region to digest the corresponding PCR products, the specific digestion pattern enabled the discrimination of the botanical sources of Stemonae Radix effectively and efficiently.
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Biopharmacy
Regular Articles
  • Hiromi Fujimoto, Maiko Higuchi, Hiroshi Watanabe, Yasuhiro Koh, Arun K ...
    Type: Regular Articles
    Subject area: Biopharmacy
    2009 Volume 32 Issue 9 Pages 1588-1593
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Darunavir (DRV) is a nonpeptidic protease inhibitor (PI) approved for the treatment of human immunodeficiency virus (HIV) infection. DRV displays potent activity against HIV strains resistant to other available PIs. Coadministration with ritonavir (RTV) improves the oral bioavailability of DRV. Inhibition of cytochrome P450 by RTV has been proposed as a mechanism for enhanced DRV bioavailability. However, interaction of these drugs with intestinal transporters has not been elucidated. This study was performed to explore the involvement of P-glycoprotein in transcellular DRV transport in monolayers of human intestinal Caco-2 and in ABCB1 multidrug resistance 1, (MDR1) gene-transfected renal LLC-PK1 (L-MDR1) cell lines. Transepithelial transport of DRV in Caco-2 cell monolayers was 2-fold greater in the basal-to-apical direction compared to that in the opposite direction. RTV had a significant inhibitory effect on the efflux transport of DRV in Caco-2 cells. The apical-to-basal DRV transport was enhanced by P-glycoprotein inhibitors, cyclosporin A and verapamil, as well as multidrug resistance-related protein (MRP/ABCC) inhibitors, probenecid and MK571. Using the L-MDR1 cell line, basal-to-apical DRV transport was much greater than in the opposite direction. Furthermore, cyclosporin A markedly inhibited the basal-to-apical DRV transport. RTV significantly increased the apical-to-basal transport of DRV in L-MDR1 cells, but reduced transport in the opposite direction. DRV inhibited P-glycoprotein-mediated efflux of calcein-acetoxymethyl ester in L-MDR1 cells with the inhibitory potency of 121 μM. These findings suggest that DRV is a substrate of P-glycoprotein and MRP, most likely MRP2. RTV appeared to inhibit P-glycoprotein, thereby enhancing the absorptive transport of DRV.
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  • Noriaki Nagai, Takatoshi Murao, Yoshimasa Ito, Norio Okamoto, Masahiro ...
    Type: Regular Articles
    Subject area: Biopharmacy
    2009 Volume 32 Issue 9 Pages 1594-1599
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    The protein sericin is the main constituent of silk. We investigated the effects of sericin on corneal wound healing in Otsuka Long-Evans Tokushima Fatty (OLETF) rats, a model for human type 2 diabetes. Corneal wounds were prepared by removal of the corneal epithelium, and documented using a TRC-50X equipped with a digital camera. Sericin solutions were instilled into the eyes of rats five times a day following corneal abrasion. Plasma glucose and triglycerides were determined using an Accutrend GCT. Cholesterol and insulin were measured using a Cholesterol E-Test Kit and ELISA Insulin Kit, respectively. The plasma levels of glucose, triglycerides, cholesterol and insulin in 38-week-old OLETF rats were significantly higher than in Long-Evans Tokushima Otsuka (LETO) rats used as normal controls, and the rate of corneal wound healing in OLETF rats was slower than in LETO rats. The corneal wounds of rats instilled with saline showed almost complete healing by 72 h after corneal epithelial abrasion. On the other hand, the corneal healing rate of OLETF rats instilled with 10% sericin solution was significantly higher than that of LETO rats instilled with saline, and the wounds showed almost complete healing at 48 h after abrasion. The corneal healing rate increased with increasing sericin concentration. The present study demonstrates that the corneal wound healing rate in OLETF rat is slower than in LETO rats, and the instillation of sericin solution has a potent effect in promoting wound healing and wound-size reduction in LETO and OLETF rats.
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  • Mehrdad Hamidi, Najmeh Zarei, Mohammad-Ali Shahbazi
    Type: Regular Articles
    Subject area: Biopharmacy
    2009 Volume 32 Issue 9 Pages 1600-1603
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    An available, simple, sensitive, and rapid method has been developed for determination of the 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitor, lovastatin in human plasma. The analytical procedure involves a one-step liquid–liquid extraction method using atorvastatin as internal standard. Chromatographic separation was carried out on a reversed phase C18 column using a mixture of 0.05 M phosphate buffer (pH 7) and acetonitrile (44.5 : 55.5, v/v) as mobile phase with UV detection set at 238 nm. The total run time of analysis was 6 min with the retention time of lovastatin being 4.3 min. A complete set of analytical method validation tests were carried out on the method. Accordingly, the method was linear in the wide range of 1—100 ng/ml. The limit of detection (LOD) and limit of quantification (LOQ) for lovastatin were 0.5 and 1 ng/ml, respectively. The method was shown to be precise with average within-run and between-run variations of 10.45±6.88 and 8.68±5.13%, respectively. The average within-run and between-run accuracy of the method throughout its linear range was 113.33±3.98 and 105.72±5.07%, respectively. The mean relative recovery of lovastatin from human plasma by the developed method was 88.61±7.00%. The applicability of the method in real pharmacokinetic situations was evaluated successfully during a bioequivalence study in 14 fasting healthy male volunteers.
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Notes
  • Kazumi Nakamura, Yukari Kuramoto, Hidefumi Mukai, Shigeru Kawakami, Yu ...
    Type: Notes
    Subject area: Biopharmacy
    2009 Volume 32 Issue 9 Pages 1628-1631
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Targeted gene delivery to macrophages is important for the treatments of various immune diseases. Since macrophages express mannose receptors, development of efficient mannosylated non-viral carriers is an effective approach to macrophages-selective in vivo gene transfection. In this study, a pH-sensitive mannosylated cholesterol derivative, Man-His-C4-Chol, which possesses histidine (His) residues, containing lipoplexes (Man-His-lipoplexes) was characterized for transfection both in vitro and in vivo. In primary cultured macrophages, both Man-His-lipoplexes and mannosylated (Man)-lipoplexes showed significantly higher cellular uptake than bare-lipoplexes and there was no significant difference between Man-His-lipoplexes and Man-lipoplexes at 37 °C but the cellular uptake of these three lipoplexes was reduced at 4 °C. Similarly, the transfection efficacy of Man-His-lipoplexes showed significantly higher gene expression than bare-lipoplexes and Man-lipoplexes. After intraperitoneal administration to mice, Man-His-lipoplexes showed higher gene expression in peritoneally exuded cells (PECs) which contained macrophages than Man-lipoplexes and bare-lipoplexes at 3, 6, and 24 h. In addition, Man-His-lipoplexes showed higher gene expression than Gal-His-lipoplexes in PECs, suggesting that Man-His-lipoplexes are taken up by macrophages via mannose receptor-mediated endocytosis. These results suggest that Man-His-lipoplexes have superior transfection activity to Man-lipoplexes in macrophages.
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Miscellaneous
Notes
  • Kei Shimizu, Mayumi Ikegami-Kawai, Tomoko Takahashi
    Type: Notes
    Subject area: Miscellaneous
    2009 Volume 32 Issue 9 Pages 1632-1635
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    Oxidative stress is implicated in the pathogenesis of diabetes mellitus. We investigated whether or not oxidized protein hydrolase (OPH), which preferentially degrades oxidized and glycated proteins, has a preventive or delaying role in diabetes mellitus. Using streptozotocin (STZ)-induced diabetic Wistar and Goto-Kakizaki (GK) rats as models of types 1 and 2 diabetes, respectively, we traced the changes in serum and urinary OPH activity with the pathological progress. Serum OPH activity increased nearly in parallel with increases in the blood glucose level in the two rat models, clarifying first in this study that serum OPH activity increases from a very early stage of diabetes. These findings suggest that increased serum OPH has at least a delaying action against the progression of diabetes through the removal of damaged proteins. Urinary OPH activity increased only in STZ-induced diabetic rats with microalbuminuria, but not in GK rats still without marked renal disorder, indicating the leakage of circulating OPH; the increase in urinary OPH activity may be a useful marker for diabetic nephropathy. OPH may provide a new therapeutic strategy against diabetes and its complications.
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  • Tatsuya Higashi, Yukiko Nagura, Kazutake Shimada, Toshimasa Toyo'oka
    Type: Notes
    Subject area: Miscellaneous
    2009 Volume 32 Issue 9 Pages 1636-1638
    Published: September 01, 2009
    Released: September 01, 2009
    JOURNALS FREE ACCESS
    It is well known that fluoxetine (Fluox), a selective serotonin reuptake inhibitor, increases the brain content of allopregnanolone (AP), a potent neuroactive steroid that positively modulates the action of γ-aminobutyric acid (GABA) at the GABA type A receptors, but the influence of Fluox on the brain and serum levels of a neuroactive androgen, 5α-androstane-3α,17β-diol (3α,5α-Adiol), is poorly understood. In this study, we examined the Fluox-evoked changes in the 3α,5α-Adiol levels and compared the level changes of 3α,5α-Adiol with those of AP. The brain and serum 3α,5α-Adiol and AP levels were determined using previously developed LC-MS/MS. The ratio of the brain 3α,5α-Adiol to the serum 3α,5α-Adiol concentrations (B/S value) was significantly elevated in the rats intraperitoneally administered Fluox (10 mg/kg). Although the magnitude of the Fluox-evoked level change in 3α,5α-Adiol was much lower than that in AP, this study demonstrated that the 3α,5α-Adiol content is also influenced by Fluox. The most probable cause for the increase in the B/S value by the Fluox treatment is the activation of the 3α-hydroxysteroid dehydrogenase enzyme followed by the promotion of the de novo biosynthesis of 3α,5α-Adiol in the brain.
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