Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
36 巻 , 4 号
選択された号の論文の25件中1~25を表示しています
Review
  • Akira Haji, Satoko Kimura, Yoshiaki Ohi
    2013 年 36 巻 4 号 p. 501-508
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Cough is an important defensive reflex that eliminates particles and secretions from the airways and protects the lower airways from the aspiration of foreign materials. Although the classical cough center is thought to be situated in or around the nucleus tractus solitarius (NTS) of the brainstem, our understanding of its profile is still incomplete. Accumulating evidence suggests a new concept of the central regulatory system for cough reflex. The cough pattern generator in the brainstem appears to be identical to the respiratory pattern generator and to function by reshaping of the discharge pattern of respiratory neurons. The generated cough motor task is transmitted to spinal motoneurons through the descending respiratory pathways. The cough-gating mechanism receives the peripheral tussigenic information through the relay neurons in the NTS and activates such a functionally flexible pattern generator by producing triggering signals. This review focuses on the cough-gating neurons that constitute the gating mechanism and play a crucial role in the generation of cough reflex.
Regular Articles
  • Ting Zheng, Ling Chen, A Long Sae Mi Noh, Mijung Yim
    2013 年 36 巻 4 号 p. 509-514
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Osteoclasts are responsible for bone erosion in diseases as diverse as osteoporosis, periodontitis, and rheumatoid arthritis. Antiseptic products have received recent attention as potential therapeutic and preventive drugs in human disease. The purpose of this study was to investigate the effect of the antiseptic cetylpyridinium chloride (CPC) on osteoclast formation using mouse bone marrow-derived macrophages (BMMs). CPC inhibited receptor activator of nuclear factor (NF)-κB ligand (RANKL)-induced osteoclast formation in a dose-dependent manner without causing cytotoxicity. The mRNA expression of cathepsin K, calcitonin receptor (CTR), and Prdm1 in osteoclasts was reduced by CPC. In experiments to elucidate its mechanism of action, CPC was found to suppress RANKL-induced expression of c-Fos and nuclear factor of activated T cells (NFATc1), transcription factors that are essential for osteoclast differentiation. CPC also inhibited RANKL-induced activation of extracellular signal-regulated kinase (ERK) and NF-κB and expression of cyclooxygenase (COX)-2. These results collectively suggest that CPC inhibits osteoclast differentiation by suppressing the activation of ERK and NF-κB and reducing the expression of COX-2, c-Fos, and NFATc1. CPC may therefore be a useful drug in the prevention of bone loss.
  • Meimi Zhao, Jinsheng Zhao, Guilin He, Xuefei Sun, Xueshi Huang, Liying ...
    2013 年 36 巻 4 号 p. 515-521
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Astragaloside IV (AS-IV) is one of the main active constituents of Astragalus membranaceus, which has various actions on the cardiovascular system. However, its electrophysiological mechanisms are not clear. In the present study, we investigated the effects of AS-IV on action potentials and membrane currents using the whole-cell patch clamp technique in isolated guinea-pig ventricular myocytes. AS-IV prolonged the action potential duration (APD) at all three tested concentrations. The peak effect was achieved with 1×10−6m, at which concentration AS-IV significantly prolonged the APD at 95% repolarization from 313.1±38.9 to 785.3±83.7 ms. AS-IV at 1×10−6m also enhanced the inward rectifier K+ currents (IK1) and inhibited the delayed rectifier K+ currents (IK). AS-IV (1×10−6m) strongly depressed the peak of voltage-dependent Ca2+ channel current (ICaL) from −607.3±37.5 to −321.1±38.3 pA. However, AS-IV was not found to affect the Na+ currents. Taken together, AS-IV prolonged APD of guinea-pig ventricular myocytes, which might be explained by its inhibition of IK. AS-IV also influences Ca2+ signaling through suppressing ICaL.
  • Chatchai Muanprasat, Lalida Sirianant, Sutthipong Sawasvirojwong, Sure ...
    2013 年 36 巻 4 号 p. 522-528
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/24
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    Our previous study has shown that dihydroisosteviol (DHIS), a derivative of stevioside isolated from Stevia rebaudiana (Bertoni), inhibits cystic fibrosis transmembrane conductance regulator (CFTR)-mediated transepithelial chloride secretion across monolayers of human intestinal epithelial (T84) cells and prevents cholera toxin-induced intestinal fluid secretion in mouse closed loop models. In this study, we aimed to investigate a mechanism by which DHIS inhibits CFTR activity. Apical chloride current measurements in Fisher rat thyroid cells stably transfected with wild-type human CFTR (FRT-CFTR cells) and T84 cells were used to investigate mechanism of CFTR inhibition by DHIS. In addition, effect of DHIS on AMP-activated protein kinase (AMPK) activation was investigated using Western blot analysis. Surprisingly, it was found that DHIS failed to inhibit CFTR-mediated apical chloride current in FRT-CFTR cells. In contrast, DHIS effectively inhibited CFTR-mediated apical chloride current induced by a cell permeable cAMP analog CPT-cAMP and a direct CFTR activator genistein in T84 cell monolayers. Interestingly, this inhibitory effect of DHIS on CFTR was significantly (p<0.05) reduced by pretreatment with compound C, an AMPK inhibitor. AICAR, a known AMPK activator, was able to inhibit CFTR activity in both FRT-CFTR and T84 cells. Western blot analysis showed that DHIS induced AMPK activation in T84 cells, but not in FRT-CFTR cells. Our results indicate that DHIS inhibits CFTR-mediated chloride secretion in T84 cells, in part, by activation of AMPK activity. DHIS therefore represents a novel candidate of AMPK activators.
  • Tetsuro Marunouchi, Yohei Abe, Mao Murata, Satomi Inomata, Atsushi San ...
    2013 年 36 巻 4 号 p. 529-539
    発行日: 2013/04/01
    公開日: 2013/04/01
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    The mechanisms underlying mitochondrial impairment in the failing heart are not yet clearly defined. In the present study, we examined the involvement of changes in small heat shock proteins (HSPs) such as HSPB1, HSPB5 and HSPB8 in mitochondrial dysfunction of the failing heart. Hemodynamic parameters of rats with myocardial infarction at the 2nd and 8th weeks (2W- and 8W-) after coronary artery ligation (CAL) were measured. The 8W-CAL rats, but not the 2W-CAL ones, showed the signs of the chronic heart failure concomitant with a reduced mitochondrial oxygen consumption rate. In the mitochondrial fraction prepared from the heart of the 2W-CAL animals, the contents of small HSPs and phosphorylated small HSPs were increased, suggesting that these increases contributed to the preservation of the mitochondrial energy-producing ability. In the failing heart, HSPB1 and HSPB8 contents and phosphorylated small HSP contents in the mitochondrial fraction were decreased, suggesting that a reduction in mitochondrial translocation of these small HSPs led to impaired mitochondrial energy-producing ability. To further define the submitochondrial locations of these small HSPs, we performed mitochondrial subfractionation. The contents of small HSPs in the 2W-CAL rats were increased in the mitochondrial inner-membrane fraction, whereas those of the 8W-CAL rats were reversed to those of the control animals. These findings suggest that small HSPs, at least in part, play an important role in the development of the impaired mitochondrial energy-producing ability that leads to heart failure after a myocardial infarction.
  • Koichi Aoki, Akihito Yokosuka, Yoshihiro Mimaki, Kohji Fukunaga, Tohru ...
    2013 年 36 巻 4 号 p. 540-547
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Ras, a small G-protein, physiologically directs cell proliferation and cell cycle via regulation of mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling cascade. Dysregulation of Ras/MEK/ERK signaling has been reported to cause tumorigenesis and gliomas. Nobiletin, a citrus flavonoid, has been shown to have anti-tumor cells action. However, it remains elusive whether nobiletin could affect Ras activity. In this study, we provide the first evidence that nobiletin suppresses the proliferation by inhibiting Ras activity in C6 glioma cells, a rat glioma cell line. First, Ras pull-down assay showed that nobiletin inhibits Ras activity in a concentration-dependent manner in C6 cells. Second, farnesyltransferase inhibitor I, a Ras inhibitor, and U0126, a MEK inhibitor, induced an inhibition of the cell proliferation in C6 cells, while the cell proliferation was inhibited by nobiletin as well. Third, western blotting revealed that nobiletin showed inhibitory effects on MEK and ERK phopsphorylation levels in a concentration-dependent manner. Finally, such an inhibitory effect on the level of ERK phosphorylation by nobiletin was appreciably prevented by Gö6976, a selective inhibitor of conventional protein kinase Cs (PKCs) showing Ca2+-sensitivity, while GF109203X, a general inhibitor for PKCs, and BAPTA, a cell-permeable Ca2+ chelator, to a lesser extent, suppressed a reduction of the phosphorylation. These findings suggest that the proliferation of C6 cells is Ras- and MEK/ERK signaling-dependent, and that nobiletin suppresses the cell proliferation by inhibiting Ras activity and MEK/ERK signaling cascade probably via a Ca2+-sensitive PKC-dependent mechanism. Thus, the natural compound has potential to be a therapeutic agent for glioma.
  • Guang-chun Sun, Yuan-yuan Ma
    2013 年 36 巻 4 号 p. 548-555
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/19
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    Vascular endothelial growth factor (VEGF), an angiogenic factor, was found to modulate synaptic plasticity by affecting K+ and Ca2+ channels and protect neuron from death by depressing glutamatergic transmission. However, whether VEGF also modulates neuronal activity through modulating voltage-gated Na+ channels (VGSCs), a main determinant of neuronal excitability, we observed the effects of VEGF on Na+ channel properties and function on cultured rat hippocampal neurons through whole-cell patch-clamp recording. We found that VEGF decreased the Na+ channel excitability by shifting the voltage-dependence of steady-state inactivation to more hyperpolarized direction, and increasing the time constants of recovery from inactivation without significantly affecting the activation process. The effect of VEGF on Na+ channel steady-state inactivation was inhibited by the specific VEGF Flk-1 receptor antagonist SU1498, but was not affected by protein kinase C (PKC)-activator 1-oleoyl-2-acetyl-sn-glycerol (OAG). Furthermore, the inhibition of Na+ currents by VEGF was frequency-dependent. In addition, the frequency of neuron firing evoked by current injection was reversibly depressed by VEGF. Therefore, our results suggest a potential role of VGSCs in the modulation of VEGF on neuronal excitability.
  • Takaharu Ishibashi, Mihoko Nomura-Nakamoto, Fusae Abe, Takayoshi Masuo ...
    2013 年 36 巻 4 号 p. 556-563
    発行日: 2013/04/01
    公開日: 2013/04/01
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    The contribution of aldehyde dehydrogenase type 2 (ALDH2) to bioactivation of glyceryl trinitrate (GTN) and isosorbide dinitrate (ISDN) was systematically examined in excised rabbit aorta and anesthetized whole animal with cyanamide, an ALDH2 inhibitor. In excised aortic preparation, the degree of inhibition by cyanamide in GTN-induced vasorelaxation (concentration ratio, calculated as EC50 in the presence of cyanamide/EC50 in the absence of cyanamide; 5.61) was twice that in ISDN-induced relaxation (2.78). However, the degree of inhibition by cyanamide, as assessed by the dose ratio (as described above, but calculated with doses) in anesthetized rabbits was 2.29 in GTN-induced hypotension (assessed by area under the curve (AUC) of 50 mmHg·min) and 7.68 in ISDN-induced hypotension. Thus, the inhibitor was 3 times more potent in ISDN-induced hypotension, a finding in conflict with to that obtained in excised aortic preparation. The rate of increase in plasma nitrite (NO2) concentration at certain hypotensive effect (50 mmHg·min of AUC) in the presence and absence of cyanamide (ΔNO2 ratio) was larger in ISDN-induced hypotension (15.01) than in GTN-induced hypotension (3.28). These results indicate that the bioactivation pathway(s) of GTN is ALDH2-dependent in aortic smooth muscle, while ADLH2-independent mechanism(s) largely take place in the whole body. In contrast, the activation mechanism(s) of ISDN is largely ALDH2-dependent in both aortic smooth muscle and whole body. Plasma NO2 may be derived from pathways other than the cyanamide-sensitive metabolic route.
  • Hiromi Sato, Hana Sugai, Hiroshi Kurosaki, Momoko Ishikawa, Asami Funa ...
    2013 年 36 巻 4 号 p. 564-573
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Peroxisome proliferator-activated receptor (PPAR) γ plays a major role in the regulation of lipid and carbohydrate metabolism. Pioglitazone is a PPARγ agonist that is widely used for the treatment of type 2 diabetes mellitus. However, female patients have been reported to experience stronger efficacy and adverse effects than male patients. This study evaluated the effects of sex hormones on PPARγ expression and activity in adipocytes. Mouse 3T3-L1 preadipocytes were used after being grown into matured adipocytes. The sex hormones 17β-estradiol (E2), testosterone (T), or 5α-androstan-17β-ol-3-one (dihydrotestosterone; DHT) were added to the matured adipocytes and the cells were then maintained for short (24–72 h) or long (1- or 2-weeks) periods. E2 significantly upregulated PPARγ protein expression in a concentration-dependent manner after extended exposure, whereas T and DHT did not have such an effect. When cells were co-treated with pioglitazone and E2, PPARγ protein expression significantly increased in an E2-dependent manner, whereas this expression seemed to be reduced by pioglitazone mono-treatment and co-treatment with DHT at higher concentrations. The secretion levels of adiponectin protein, a major indicator of PPARγ activity, were significantly decreased by DHT, but were not affected by E2. Finally a luciferase assay was performed using a PPAR response element-Luk reporter gene. Transcriptional activity was not changed by any of single sex hormone treatment, but was significantly downregulated by co-treatment with pioglitazone and DHT. Taken together, our results suggest that sex hormones may influence PPARγ expression and function, which may explain the observed sex-specific different effect of pioglitazone.
  • Hiroyuki Watanabe, Hiroaki Ikesue, Tomoko Tsujikawa, Kenichiro Nagata, ...
    2013 年 36 巻 4 号 p. 574-578
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/08
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    Intravenous injection of bendamustine often causes venous irritation and also deteriorates the patient’s quality of life. Thus, we evaluated the risk factors associated with venous irritation induced by bendamustine in patients with follicular lymphoma or mantle cell lymphoma. We also evaluated the effectiveness of intervention of changing the preparation procedure for bendamustine. All data were retrospectively collected from the electronic medical record system. In the initial analysis of the total 43 courses of bendamustine therapy, most patients (88%) were administered bendamustine with 250 mL of diluent according to the bendamustine package insert in Japan. The median concentration of bendamustine solution (0.56 mg/mL vs. 0.24 mg/mL) and the incidences of venous irritation (66% vs. 0%, p=0.01) were significantly different between the patients receiving bendamustine at 250 mL and 500 mL of diluent. Based on this result, we proposed changing the final volume of bendamustine dissolution from 250 to 500 mL, which is recommended in other countries. After this intervention, the incidence of venous irritation was significantly reduced from 58 to 20% (p=0.02). The incidence of venous irritation increased in a concentration-dependent manner (≤0.40 mg/mL: 6%; 0.41–0.60 mg/mL: 62%, p<0.001; >0.60 mg/mL: 75%, p<0.001). We conclude that a high concentration bendamustine solution is a risk factor for venous irritation and that 500 mL of diluent is ideal. To further reduce the incidence of venous irritation, the concentration of bendamustine solution is recommended to be 0.40 mg/mL or less.
  • Wei Yu, Qing Liu, Sujuan Zhu
    2013 年 36 巻 4 号 p. 579-584
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/04
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    Carvacrol (CAR), a naturally occurring phenolic monoterpene, has been shown to possess diverse biological activities. The present study was undertaken to evaluate the cardioprotective potential of CAR against myocardial ischemic damage in a rat model of acute myocardial infarction. CAR significantly diminished the infarct size and myocardial enzymes including creatine kinase (CK), the MB isoenzyme of creatine kinase (CK-MB), lactate dehydrogenase (LDH) and cardiac troponin T (cTnT). Reduced level of malondialdehyde (MDA), obviously elevated activities of superoxide dismutase (SOD) and non-enzymatic scavenger glutathione (GSH) as well as glutathione peroxidase (GSH-PX) were also found in CAR-treated groups. Treatment with CAR remarkably inhibited the protein expressions of caspase-3 and Bax, but increased the level of Bcl-2 protein in infarcted rats by Western blot analysis. The finding suggests that the cardioprotection of CAR associate with its anti-oxidative and anti-apoptotic properties in acute myocardial infarction of rats.
  • Hirokazu Hara, Asuka Maeda, Tetsuro Kamiya, Tetsuo Adachi
    2013 年 36 巻 4 号 p. 585-591
    発行日: 2013/04/01
    公開日: 2013/04/01
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    There is evidence that excessive zinc (Zn2+) release from presynaptic terminals following brain injuries such as ischemia and severe epileptic seizures induces neuronal cell death. Apomorphine (Apo), a dopamine receptor agonist, has been shown to have pleiotropic biological functions. In this study, we investigated whether Apo protects cultured cortical neurons from neurotoxicity provoked by excessive Zn2+ exposure. Pretreatment with Apo dose- and time-dependently ameliorated Zn2+ neurotoxicity. In addition, pretreatment with Apo prevented intracellular nicotinamide adenine dinucleotide (NAD+) and ATP depletion caused by Zn2+ exposure. Dopamine receptor antagonists did not influence Apo protection against Zn2+ neurotoxicity. Apo is shown to be autoxidized to produce oxidized products such as reactive oxygen species and quinones. N-Acetylcysteine, a thiol compound, partially reduced Apo protection. Entry of Zn2+ into neurons is thought to be a critical step of Zn2+ neurotoxicity. Interestingly, we found that pretreatment with Apo decreased elevation of intracellular Zn2+ levels after Zn2+ exposure and induced mRNA expression of the zinc transporter ZnT1, which transports intracellular Zn2+ out of cells, and metallothionein. Taken together, these results suggest that the protective effects of Apo are regulated, at least in part, by its oxidized products, and preventing intracellular accumulation of Zn2+ contributes to Apo protection against Zn2+ neurotoxicity.
  • Yoshihiko Nakatani, Taku Amano, Hiroshi Takeda
    2013 年 36 巻 4 号 p. 592-601
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/06
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    Macrophages are white blood cells within tissues that are produced by monocytes and help to protect against infection by bacteria through phagocytosis. Several studies have shown a correlation between the state of depression and abnormalities in the immune response. Corticosterone (CORT), which is often referred to as the stress hormone, is a well-known regulator of peripheral immune responses and also shows anti-inflammatory properties in the body. However, it is still unclear how CORT regulates macrophage function. In this study, we focused on the effects of CORT on the proliferation and survival of macrophage cells using the macrophage cell line RAW264.7. Under treatment with 10 µm CORT for 24 h, the proliferation of RAW264.7 cells decreased to 73.6% of that in the control. Moreover, this inhibition was blocked by treatment with mifepristone, a glucocorticoid receptor (GR) antagonist, but not by spironolactone, a mineralocorticoid receptor (MR) antagonist. In an lactate dehydrogenase (LDH) assay, CORT did not show any cytotoxic effect on RAW264.7 cells. JC-1 cell staining also showed that CORT did not influence mitochondrial dysfunction in RAW264.7 cells. In an investigation of the modulation of a signaling cascade by CORT, treatment with CORT promoted the translocation of GR, but not MR, from the cytosol to the nucleus in RAW264.7 cells. In conclusion, our findings suggest that CORT suppresses the proliferation of RAW264.7 cells by controlling the transcription of a particular gene, which is related to cell proliferation, through the formation of a CORT–GR complex.
  • Kohei Sano, Mayumi Okada, Hayato Hisada, Kenta Shimokawa, Hideo Saji, ...
    2013 年 36 巻 4 号 p. 602-608
    発行日: 2013/04/01
    公開日: 2013/04/01
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    On the basis of the findings obtained by X-ray crystallography of Ga-DOTA chelates and the drug design concept of bifunctional radiopharmaceuticals, we previously designed and synthesized a radiogallium-labeled DOTA chelate containing two metronidazole moieties, 67Ga-DOTA-MN2, for hypoxic tumor imaging. As expected, 67Ga-DOTA-MN2 exhibited high in vivo stability, although two carboxyl groups in the DOTA skeleton were conjugated with metronidazole moieties. In this study, we evaluated 67/68Ga-DOTA-MN2 as a nuclear imaging agent for hypoxic tumors. 67Ga-labeling of DOTA-MN2 with 67GaCl3 was achieved with high radiochemical yield (>85%) by 1-min of microwave irradiation (50 W). The pharmacokinetics of 67Ga-DOTA-MN2 were examined in FM3A tumor-bearing mice, and compared with those of 67Ga-DOTA-MN1 containing one metronidazole unit and 67Ga-DOTA. Upon administration, 67Ga-DOTA-MN2 exhibited higher accumulation in the implanted tumors than 67Ga-DOTA. Tumor-to-blood ratios of 67Ga-DOTA-MN2 were about two-fold higher than those of 67Ga-DOTA-MN1. Autoradiographic analysis showed the heterogeneous localization of 67Ga-DOTA-MN2 in the tumors, which corresponds to hypoxic regions suggested by well-established hypoxia marker drug, pimonidazole. Furthermore, in positron emission tomography (PET) study, the tumors of mice administered 68Ga-labeled DOTA-MN2 were clearly imaged by small-animal PET at 1 h after administration. This study demonstrates the potential usefulness of 67/68Ga-DOTA-MN2 as a nuclear imaging agent for hypoxic tumors and suggests that two functional moieties, such as metronidazole, can be conjugated to radiogallium-DOTA chelate without reducing the complex stability. The present findings provide useful information about the chemical design of radiogallium-labeled radiopharmaceuticals for PET and single photon emission computed tomography (SPECT) studies.
  • Yong-Sik Kim, Tamanna Zerin, Ho-Yeon Song
    2013 年 36 巻 4 号 p. 609-615
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Ellagic acid (EA) is a natural dietary polyphenol whose benefits in a variety of diseases shown in epidemiological and experimental studies involve anti-inflammation, anti-proliferation, anti-angiogenesis, anti-carcinogenesis and anti-oxidation properties. This study aimed to evaluate the effect of EA against paraquat (PQ)-induced oxidative stress. PQ decreased the viability of A549 cells in dose- and time-dependent manners, which was associated with the massive generation of reactive oxygen species (ROS). However, cell viability was significantly recovered by the treatment of EA, from 47.01±1.59% to 66.04±2.84%. The release of lactate dehydrogenase (LDH) was also decreased with the treatment of EA in PQ-treated A549 cells. EA induced the level of expression and activation of nuclear factor-erythroid 2-related factor (Nrf2) and its target cytoprotective and antioxidant genes, heme oxygenase-1 (HO-1) and quinone oxidoreductase 1 (NQO1). The antioxidant potential of EA might be directly correlated with the increased expression of HO-1 and NQO1, whose expression may have surmounted the oxidative stress generated by PQ. Notably, EA treatment significantly reduced the levels of biochemical markers as lipid peroxidation, reduced the intracellular ROS level, and surmounted total glutathione level in A549 cells. Data indicate that the antioxidant and cytoprotective properties of EA reduce PQ-induced cytotoxicity in human alveolar A549 cells.
  • Hikaru Satoh, Naohito Ide, Yoshiyuki Kagawa, Toshio Maeda
    2013 年 36 巻 4 号 p. 616-623
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/02
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    We have isolated insulin resistant mice (ddY-H mice) which are spontaneously induced even if fed with the standard chow pellets. Since marked accumulation of triglycerides (TG) in liver was observed, the present study investigated causes of hepatic TG accumulation in ddY-H mice fed with the standard chow pellets. In ddY-H mice, hepatic TG content increased from seven-weeks of age, and further marked accumulation of TG was observed at 20-weeks of age. Histologically, fat droplets appeared in pericentral parenchymal cells of the liver from nine-weeks of age, and the size and number of droplets were increased in hepatic lobules at 15-weeks of age, suggesting hepatic steatosis was spontaneously induced. Although secretion of TG from liver to blood in ddY-H mice was not increased, fat absorption from the digestive tract was significantly enhanced. The mRNA expressions of peroxisome proliferator-activated receptor γ (PPARγ) involved in fat accumulation and fatty acid translocase (CD36) involved in the transportation of fatty acid into the liver were markedly increased. However, gene expressions of factors involved in lipogenesis, β-oxidation of fatty acid and lipoprotein secretion were not changed. Pioglitazone (9 mg/kg), the PPARγ agonist, administered for six weeks deteriorated hepatic steatosis in ddY-H mice. Although pioglitazone did not affect gene expressions of PPARγ in the liver, CD36 and fat-specific protein 27 (fsp27), targets of PPARγ, were markedly elevated. These results suggest that, in the livers of ddY-H mice, hepatic steatosis is induced by increased incorporation of fatty acid into the liver via increased PPARγ expression.
  • Masataka Tajima, Nobutomo Ikarashi, Shintaro Igeta, Takahiro Toda, Mak ...
    2013 年 36 巻 4 号 p. 624-634
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/01
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    Changes in the expression level and activity of cytochrome P450 (CYP) in the liver are caused by various factors and affect the pharmacokinetics of drugs. The purpose of this study was to determine whether the expression of CYP3A is affected by a high-fat diet. In addition, we examined whether the type of diet given to mice could produce changes in the expression level and activity of CYP3A. Mice were fed a purified diet containing 10 kcal% lard (control group) or 60 kcal% lard (HF group) or regular mouse chow containing 13 kcal% of fat (MF group) for 4 weeks. No significant differences were observed in the hepatic CYP3A protein expression level between the HF group and the control group. The CYP3A protein expression in the MF group was significantly higher than that observed in the control group. In the MF group, the area under the curve (AUC) of intraperitoneally administered triazolam was lower. Because lithocholic acid (LCA) is known to increase hepatic CYP3A expression, the levels of Clostridium sordellii and LCA in the feces were measured. In the MF group, the levels of Clostridium sordellii and LCA were higher. It has been demonstrated that a high-fat diet does not cause any changes in hepatic CYP3A expression. In addition, the different diets caused alterations in the enteric environment, which triggered changes in CYP3A expression. Therefore, it is necessary to carefully consider the type of feed while performing animal experiments to evaluate the pharmacokinetics of drugs.
  • Kenichi Nakamura, Katsuko Komatsu, Masao Hattori, Makoto Iwashima
    2013 年 36 巻 4 号 p. 635-640
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/17
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    We previously isolated the human intestinal bacterium, strain PUE, which can cleave the C-glucosidic bond of puerarin to yield its aglycone daidzein and glucose. In this study, we partially purified puerarin C-glucosidic bond cleaving enzyme from the cell-free extract of strain PUE and demonstrated that the reaction was catalyzed by at least three proteins, Mn2+, and oxidized form of nicotinamide adenine dinucleotide (NAD+). We completely purified one of the proteins, called protein C, by chromatographic separation in three steps. The molecular mass of protein C was approximately 40 kDa and the amino acid sequence of its N-terminal region shows high homology to those of two putative proteins which belong to Gfo/Idh/MocA family oxidoreductase. Protein C catalyzed hydrogen-deuterium exchange reaction of puerarin to 2″-deuterated puerarin in D2O condition, which closely resembles those of glycoside hydrolase family 4 and 109.
  • Xin Wang, Xu Zhang, Zhiliang Xu, Zhizeng Wang, Xiaoxuan Yue, Hongyu Li
    2013 年 36 巻 4 号 p. 641-648
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/25
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    The success of arsenic trioxide (ATO) in treatment of acute promyelocytic leukemia (APL) attracts a great deal of attention to researchers to explore its activity of anti-leukemia. However, ATO has unavailable effect on chronic myeloid leukemia (CML), especially multidrug resistant (MDR)-CML, unless using high concentration. Realgar (As4S4) has been employed in Chinese traditional medicine for 1500 years. Research evidences confirmed realgar has similar effect on treating with APL as ATO, but the problem of large dose and long period in the CML/MDR-CML treatment still exist. By using a microbial leaching process with Acidithiobacillus ferrooxidans, we obtained realgar transforming solution (RTS) which showed significantly higher extent in inhibiting CML cell line K562 and MDR-CML cell line K562/ADM, and then trigger apoptosis. Both K562 and K562/ADM showed arsenic-dose-dependent effect on RTS. Interestingly, the overexpression of MDR1 mRNA and P-glucoprotein (P-gp) in K562/ADM cells were down-regulated by RTS, where there are no obvious effects on ATO and realgar and arsenic can be subsequently accumulated in K562/ADM cells efficiently. The intracellular accumulation of arsenic in K562/ADM cells treated with RTS for 4 h was 2-fold and 16-folds higher than those treated with realgar or ATO. Meanwhile, Western blot analysis of AQP9, the main transporter of arsenic, was increased by RTS treatment particularly in K562/ADM. Thus, these results suggested that the effect from a certain arsenical or a variety of arsenicals in RTS might be a promising candidate both for treating CML/MDR-CML alone and as combinations with currently used anti-CML/MDR-CML drug, although arsenical forms in RTS are undefined.
  • Masataka Tajima, Nobutomo Ikarashi, Takehiro Okaniwa, Yukari Imahori, ...
    2013 年 36 巻 4 号 p. 649-657
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/28
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    It has been recently reported that the consumption of a high-fat diet during pregnancy exerts various effects on fetuses and newborn mice. The purpose of this study was to determine the effects of a high-fat diet during pregnancy on the expression of cytochrome P450 (CYP) in the livers of offspring. Mouse dams were fed a high-fat diet during pregnancy from the time of conception. After their birth, the newborn mice were fed a normal diet until 12 weeks of age. In the livers of the infant male mice that consumed a high-fat diet, the protein expression of CYP3A and CYP2C was decreased, and the protein expression of CYP1A and CYP2E was increased at 6 and 12 weeks of age. However, almost no changes were observed in the CYP proteins at 6 and 12 weeks of age in the livers of the infant female mice that consumed a high-fat diet. The amount of pregnane X receptor (PXR) translocated into the nucleus was reduced in the livers of infant male mice that consumed a high-fat diet. However, there was neither an increase in tumor necrosis factor-α or interleukin-1β nor a decrease in lithocholic acid. These data suggested that CYP3A and CYP2C might decrease as a result of the decrease in the amount of nuclear PXR in infant male mice that consumed a high-fat diet. The results of this study suggested that the consumption of a high-fat diet by pregnant mothers may be one explanation for individual differences in pharmacokinetics.
  • Masako Onishi, Hiroshi Kamimori
    2013 年 36 巻 4 号 p. 658-663
    発行日: 2013/04/01
    公開日: 2013/04/01
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    In the present study, we developed a high-throughput and sensitive assay for interactions of amphotericin B (AmB) with two model lipid membranes, which mimicked mammal cell membrane and fungal membrane using surface plasmon resonance (SPR). The binding kinetics of AmB to the membrane could be analyzed by multiple sensorgrams obtained at different AmB concentrations, indicating that the binding properties could be clarified for an approximately 7-fold concentration range. AmB showed an approximately 18-fold higher affinity for ergosterol-containing membrane than for cholesterol-containing membrane. We also optimized the procedure for the reproducible immobilization of liposome containing the sterols and the estimation of binding kinetics of AmB to the lipid membranes, and the sensitivity of AmB to membrane interaction was 20-fold higher, compared with the reported method. The throughput of the established method for the binding kinetics characterization was calculated to be 10 compounds a day. The results demonstrate that the established SPR method could be a valuable tool for predicting selective binding to sterol-containing membranes.
  • Renata de Freitas Fischer Vieira, Daniela Teves Nardi, Nanci Nasciment ...
    2013 年 36 巻 4 号 p. 664-675
    発行日: 2013/04/01
    公開日: 2013/04/01
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    The present work aimed at evaluating the radiolysis effect upon a set of peptides, most of them involved in physiological functions. To generate reactive radical species, a Co60 source (up to 15 kGy) was used for controlled gamma irradiation of some peptide solutions including derivatives attaching the stable free radical Toac (2,2,6,6-tetramethypiperidine-1-oxyl-4-amino-4-carboxylic acid). Regardless of the peptide sequence, a nonlinear and progressive degradation of a total of nine peptides was detected. The results were interpreted in the light of the half-life dose (D1/2) parameter which represents the dose necessary for 50% peptide structure degradation. The vasoactive angiotensin II (AngII)’s analogue Ang-(1–7) showed greater stability towards gamma ray radiation than bradykinin (BK), Toac0-BK, Pro4-BK (D1/2 around 4 and 2 kGy, respectively) which decreased to about 0.5–1.0 kGy in the case of acetyl-α-melanocyte-stimulating hormone (Ac-α-MSH) and substance P (SP). In terms of peptide structural modifications, the data acquired from different analytical methods suggested a Phe to Tyr (or its ortho and/or meta isomers) transformation as a consequence of the hydroxyl moiety insertion. Noteworthy, this effect seemed to be position-dependent as only Phe located at or near the C-terminal portion seemed to display this transformation. In contrast, Met is comparatively more easily oxidized, thus allowing to conclude that gamma irradiation may induce a complex position and/or sequence-dependent effect on peptides. As previously applied for BK, some irradiated peptides were submitted to their by-products purification, indeed a complementary target of the present approach for development of uncommon analogues for further structure–function investigation.
  • Shinya Motohashi, Yasuaki Mino, Katsuhito Hori, Takafumi Naito, Seiji ...
    2013 年 36 巻 4 号 p. 676-681
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/02/01
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    The pharmacokinetics of aprepitant, a neurokinin-1 receptor antagonist, have not been fully evaluated in clinical settings. The aim of this study was to characterize the plasma pharmacokinetics of aprepitant and reveal their influence of laboratory tests and cytochrome P450 (CYP) 3A5 gene polymorphisms in cancer patients. Forty-four Japanese cancer patients receiving cisplatin-based chemotherapy for the first time following oral aprepitant (125 mg on day 1 and 80 mg on days 2 and 3) were enrolled. The patients did not have gastrointestinal disease and the clinical laboratory values were within their normal reference levels. The plasma concentrations of aprepitant 24 (day 2 predose), 72, and 120 h after the first aprepitant administration were determined using LC-MS/MS. The relationships between plasma exposure to aprepitant and body weight, clinical laboratory values, age, gender, or CYP3A5*3 were investigated. The median and interquartile ranges of the 120-h area under the plasma concentration time curve (AUC)0–120 of aprepitant were 73215 and 55518–91121 ng h/mL. The coefficient of variation value for aprepitant AUC0–120 was 53%. The AUC0–120 of aprepitant was correlated with the levels of total bilirubin and serum albumin, respectively (r=0.454, p<0.01 and r=0.287, p=0.06), but not with other non-genetic factors and CYP3A5 genetic variants in a univariate analysis. The AUC0–120 of aprepitant was significantly correlated with the level of total bilirubin (adjusted R2=0.187, p<0.01) in a multivariate analysis. In conclusion, the plasma pharmacokinetics of aprepitant varied markedly in cancer patients receiving cisplatin-based chemotherapy for the first time and were correlated with the level of total bilirubin.
Notes
  • Masaki Ishikawa, Yoko Tajima, Mayumi Murayama, Yuya Senoo, Keiko Maeka ...
    2013 年 36 巻 4 号 p. 682-685
    発行日: 2013/04/01
    公開日: 2013/04/01
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    Biomarkers will play important roles in disease diagnosis, drug development, and the proper use of drugs. Blood is considered the best biofluid for biomarker research because it is easy to access and a wealth of data are available. However, previous studies revealed that several ionic metabolites showed different levels (including presence or absence) in plasma and serum. Thus, attention should be paid to selecting the best biofluid for biomarker exploration. Many lipid molecules have biological significance and thus would be candidate biomarkers. However, no comprehensive study revealing differences in lipid metabolite levels between plasma and serum has been undertaken. Furthermore, gender differences have not been reported. To clarify the difference in the levels of lipid metabolites between human plasma and serum from both genders, we performed lipid metabolomic analysis using liquid chromatography-mass spectrometry-based systems for phospholipids (PLs), lysoPLs, sphingomyelins, ceramides and oxidative fatty acids. Our results revealed that most of the lipid metabolites were present at similar levels in plasma and serum and in males and females. However, several oxidative fatty acid metabolites showed differences. Of the metabolites related to clotting processes, three showed higher levels in serum than in plasma, and three were detected only in serum. Furthermore, four metabolites were present at different levels between males and females, and two were detected only in males. Thus, attention should be paid to the selection of plasma or serum when utilizing these lipid metabolites as biomarkers.
  • Yasuyoshi Ishiwata, Yutaka Takahashi, Masashi Nagata, Masato Yasuhara
    2013 年 36 巻 4 号 p. 686-690
    発行日: 2013/04/01
    公開日: 2013/04/01
    [早期公開] 公開日: 2013/01/25
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    Moxifloxacin, a fluoroquinolone antimicrobial agent, has been reported to cause serum glucose abnormalities such as hyper- and hypoglycemia. The purpose of the present study was to investigate the effect of moxifloxacin on serum glucose concentrations in rats. Rats were intravenously injected with moxifloxacin and samples of their arterial blood were collected periodically. Serum glucose concentrations increased with moxifloxacin at 100 mg/kg, and temporal elevations were observed in serum epinephrine and histamine concentrations. On the other hand, intravenous injection of moxifloxacin at 75 mg/kg did not affect serum glucose, epinephrine, or histamine concentrations. Serum immunoreactive insulin concentrations remained unchanged by moxifloxacin both at 75 and 100 mg/kg. In conclusion, moxifloxacin can induce histamine release, leading to an increase in serum epinephrine concentrations and hyperglycemia.
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