Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
38 巻 , 8 号
選択された号の論文の25件中1~25を表示しています
Current Topics: Cell Death in Neuromuscular Diseases
  • Atsushi Sanbe
    2015 年 38 巻 8 号 p. 1093
    発行日: 2015/08/01
    公開日: 2015/08/01
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  • Tetsuro Marunouchi, Kouichi Tanonaka
    2015 年 38 巻 8 号 p. 1094-1097
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Loss of cardiac myocytes plays a critical role in the pathogenesis of cardiovascular disorders. A decrease in the number of cardiac myocytes in cardiac diseases results in sustained, irreversible contractile failure of myocardium. Therefore prevention of cardiac cell death is a potential therapeutic strategy for various heart diseases. It is well accepted that three types of phenomena such as apoptosis, necrosis, and autophagy may be involved in myocardial cell death. Apoptosis is a highly regulated process that is promoted via death receptor pathway in the plasma membrane or via mitochondrial pathway. Necrosis is induced via mitochondrial swelling, cell rupture, and subsequent inflammation. Autophagy is a cell survival mechanism that involves degradation and recycling of cytoplasmic components. As compared with the other two mechanisms, autophagy may mediate cell death under specific conditions. These three types of cell death in the myocardium are discussed in this article.
  • Satoru Kobayashi
    2015 年 38 巻 8 号 p. 1098-1103
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Autophagy is a degradation system for intracellular components. One of the roles of autophagy is the prompt removal of damaged organelles. Another unique role is to supply resources that maintain metabolism in response to the cellular nutritional state. Precise management of all the components in the autophagic system is essential for cellular health. Especially important are the selectivity of target cargos for autophagy, and the coordination of autophagy with the lysosomal catabolic process. This review outlines our current understanding of autophagy and discusses potential therapeutic perspectives. Emphasis will be given to lysosomal function as a central controller of metabolism, and to selective autophagy as a key mechanism for the efficient removal of dysfunctional organelles.
  • Hideki Hayashi, Norio Takagi
    2015 年 38 巻 8 号 p. 1104-1108
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Functions of the central nervous system (CNS) are based on a complex neural network. It is believed that the CNS has several neuroprotective mechanisms operated by neurons, glia and other types of cells against various types of neuronal damage. Since mature, differentiated neurons are not able to divide, it is important to protect neurons from damage prior to death. The neuroprotective effects of a number of pharmaceutical agents and natural products against necrosis and apoptosis of the CNS neurons have been reported, thus this review will mainly discuss several endogenous neuroprotectants and their mechanisms.
  • Kyaw Htet Aung, Shinji Tsukahara, Fumihiko Maekawa, Keiko Nohara, Kazu ...
    2015 年 38 巻 8 号 p. 1109-1112
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Environmental influences, such as chemical exposure, have long been considered potential risk factors for neurodegenerative disorders, including neuromuscular diseases. However, no definitive links between environmental chemical exposure and a pathogenic mechanism of neurodegenerative disease has yet been established. In this study, we describe that exposure to arsenic, an environmental pollutant naturally found in drinking water, induces neuronal cell death and alteration of morphology, particularly neurite outgrowth and in the cytoskeleton of neurons. Since progressive cell loss accompanied by the alteration of neuronal structures and cytoskeleton is considered the major pathologic feature of neurodegenerative disorders, arsenic-induced neurotoxicity might contribute to an etiologic mechanism of some neurodegenerative diseases. Further, we discuss the importance of in vitro assay, particularly an embryonic toxicity test, for assessing the neurotoxicity of chemicals, because most of chemicals found in our environment remain to be evaluated regarding their neurotoxicity risk for neurodegenerative diseases.
Regular Articles
  • Kumiko Ueda, Mika Hosokawa, Seigo Iwakawa
    2015 年 38 巻 8 号 p. 1113-1119
    発行日: 2015/08/01
    公開日: 2015/08/01
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    DNA hypermethylation, an epigenetic change that silences gene expression without altering nucleotide sequences, plays a critical role in the formation and progression of colorectal cancers as well as in the acquisition of drug resistance. Decitabine (DAC), a DNA methyltransferase 1 inhibitor of nucleoside analogues, has been shown to restore gene expression silenced by hypermethylation. In the present study, the mechanisms underlying both uridine and DAC uptake were examined in the human colon cancer cell line HCT116. Real-time polymerase chain reaction analysis revealed that ENT1 mRNA was the most abundant among the nucleoside transporters examined in HCT116 cells. The ENT1 protein was detected in the membrane fraction, as determined by Western blotting. The uptake of uridine or DAC was time- and concentration-dependent, but also Na+-independent. The uptake of these agents was inhibited by S-(4-nitrobenzyl)-6-thioinosine (NBMPR), an inhibitor of equilibrative nucleoside transporters (ENTs), and was also decreased in cells treated with ENT1 small interfering RNA. The uptake of both uridine and DAC was inhibited by uridine, cytidine, adenosine, or inosine, while that of DAC was also inhibited by thymidine. The expression of MAGEA1 mRNA, the DNA of which was methylated in HCT116 cells, was increased by DAC treatment, and this increment was attenuated by concomitant treatment with NBMPR. The IC50 value of DAC was also increased in the presence of NBMPR. These results suggest that DAC is mainly taken up by ENT1 and that this uptake is one of the key determinants of the activity of DAC in HCT116 cells.
  • Yuya Yoshida, Norihisa Mikami, Yuki Matsushima, Fumiya Otani, Mai Miya ...
    2015 年 38 巻 8 号 p. 1120-1125
    発行日: 2015/08/01
    公開日: 2015/08/01
    [早期公開] 公開日: 2015/05/20
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    We previously reported that combination treatment with fingolimod (FTY720) plus antigenic peptide of glucose-6-phosphate isomerase (residues 325–339) (GPI325–339) from the onset of symptoms significantly inhibited disease progression in a mouse model of GPI325–339-induced arthritis. In this study, we investigated the mechanism(s) involved. The model mice were treated from arthritis onset with FTY720 alone, GPI325–339 alone, or the combination of FTY720 plus GPI325–339. At the end of treatment, inguinal lymph nodes (LNs) were excised and examined histologically and in flow cytometry. Levels of apoptotic cells, programmed death-1-expressing CD4+forkhead box P3 nonregulatory T cells (non-Tregs), and cytotoxic T-lymphocyte antigen 4-expressing non-Tregs in inguinal LNs were markedly increased in the combination treatment group mice. Regulatory T cells (Tregs) were also increased. These results indicate that combination treatment with FTY720 plus GPI325–339 inhibits the progression of arthritis by inducing clonal deletion and anergy of pathogenic T cells and also by immune suppression via Tregs.
  • Chiharu Miyajima, Yuka Itoh, Yasumichi Inoue, Hidetoshi Hayashi
    2015 年 38 巻 8 号 p. 1126-1133
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Tribbles 1 (TRB1), a member of the Tribbles family, is a pseudokinase that is conserved among species and implicated in various human diseases including leukemia, cardiovascular diseases, and metabolic disorders. However, the role of TRB1 in the immune response is not understood. To evaluate this role, we examined regulation of TRB1 expression and the function of TRB1 in interleukin-2 (IL-2) induction in Jurkat cells, a human acute T cell leukemia cell line. We found that TRB1 was strongly induced by phorbol 12-myristate 13-acetate (PMA) and ionomycin in these cells. IL-2 expression was induced in Jurkat cells activated by PMA and ionomycin; however, knockdown of TRB1 resulted in decreased induction of IL-2. TRB1 null Jurkat cells established using the CRISPR/Cas9 system also showed reduction of IL-2 expression on PMA/ionomycin stimulation. TRB1 knockdown also markedly inhibited IL-2 promoter activation. To determine the mechanism of the stimulatory effect on IL-2 induction, we focused on histone deacetylases (HDACs), and found that HDAC1 preferentially interacts with TRB1. TRB1 suppressed the interaction of HDAC1 with nuclear factor of activated T cells 2 (NFAT2), which is a crucial transcription factor for IL-2 induction. These results indicate that TRB1 is a positive regulator of IL-2 induction in activated T cells.
  • Yueliu Sun, Xiaorong Hu, Gangying Hu, Changwu Xu, Hong Jiang
    2015 年 38 巻 8 号 p. 1134-1141
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Endothelial senescence has been proposed to be involved in endothelial dysfunction and atherogenesis. Curcumin, a natural phenol, possesses antioxidant and anti-inflammatory properties. However, the effect of curcumin on endothelial senescence is unclear. This study explores the effect of curcumin on hydrogen peroxide (H2O2)-induced endothelial premature senescence and the mechanisms involved. Human umbilical vein endothelial cells (HUVECs) were cultured, and premature senescence was induced with 100 µM H2O2. Results showed that pretreatment with curcumin significantly attenuated the H2O2-induced HUVECs’ premature senescence, which was evidenced by a decreased percentage of senescence-associated β-galactosidase positive cells, improved cell division and decreased expression of senescence-associated protein p21 (all p<0.05). Pretreatment with curcumin decreased oxidative stress and apoptosis in H2O2-treated HUVECs. Treatment of HUVECs with H2O2 also down-regulated the phosphorylation of endothelial nitric oxide synthase (eNOS), decreased the level of nitric oxide in the culture medium, and inhibited the protein expression and enzymatic activity of silent information regulator 1 (SIRT1), while pretreatment with curcumin partly reversed these effects (all p<0.05). Treatment with curcumin alone enhanced the enzymatic activity of SIRT1, but didn’t affect cellular senescence, cell growth or apoptosis compared to the Control. The inhibition of SIRT1 using SIRT1 short interfering RNA (siRNA) could decrease the expression and phosphorylation of eNOS and abrogate the protective effect of curcumin on H2O2-induced premature senescence. These findings suggest that curcumin could attenuate oxidative stress-induced HUVECs’ premature senescence via the activation of SIRT1.
  • Kexin Wang, Yu Sun, Peng Lin, Jun Song, Ruxing Zhao, Wenjuan Li, Xingu ...
    2015 年 38 巻 8 号 p. 1142-1149
    発行日: 2015/08/01
    公開日: 2015/08/01
    [早期公開] 公開日: 2015/06/02
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    β-Cell insufficiency plays an important role in the development of diabetes. Environmental factors, including lifestyle, play a critical role in β-cell dysfunction. Modern lifestyles affect the inherent circadian clock in central and peripheral organs. Recent studies have demonstrated that the normal intrinsic circadian clock in islets was essential for the viability of β cells and their insulin secretory function. Overall, however, the data are inconclusive. Our study demonstrated that the disrupted circadian rhythm of islets in streptozotocin induced type1 diabetic mice may be associated with impaired β-cell function and glucose intolerance. Liraglutide, a glucagon-like peptide-1 (GLP-1) analogue, could partially restore the normal circadian rhythm and activate the 5′ AMP-activated protein kinase (AMPK) signaling pathway. Our study provided evidence demonstrating that Liraglutide might restore β-cell function and protect against the development of diabetes in a mouse model by attenuating the disruption of the intrinsic circadian rhythm in islets and by activating AMPK signaling.
  • Yu Zhou, Yan Li, Tong Mu
    2015 年 38 巻 8 号 p. 1150-1160
    発行日: 2015/08/01
    公開日: 2015/08/01
    [早期公開] 公開日: 2015/06/03
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    High-mobility group box 1 (HMGB1) and its natural receptor, Toll-like receptor-4 (TLR4), are involved in various infectious or noninfectious diseases including hemorrhagic shock. HMGB1 neutralizing antibody (anti-HMGB1 monoclonal antibody (mAb)) treatment was shown to alleviate ischemia–reperfusion injury effectively. The aim of this study was to explore whether and by what mechanisms anti-HMGB1 mAb attenuates hemorrhagic shock and resuscitation (HS/R)-induced cardiac injury. Employing rat HS/R models, we found that anti-HMGB1 mAb treatment improved HS/R-induced cardiac function deterioration, attenuated cardiac enzyme elevation, improved ATP loss, and protected cardiac tissue. Anti-HMGB1 mAb also inhibited the production of inflammatory factors interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α). Moreover, anti-HMGB1 mAb reduced apoptotic responses by suppressing activated caspase-3 and reversing apoptotic gene expression of capase-3, Bax, and Bcl-2 in rat cardiac tissue. Moreover, anti-HMGB1 mAb decreased HS/R-induced HMGB1 and TLR4 expression elevation. We further confirmed that anti-HMGB1 mAb inhibited lipopolysaccharide-activated HGMB1 and TLR4 expression and decreased inflammatory factors IL-1β, IL-6, and TNF-α at the cellular level. It was concluded that anti-HMGB1 mAb treatment protects rats from cardiac injury induced by HS/R, and the beneficial effects may be related to its inhibitory effects on the HMGB1-TLR4 axis.
  • Eun-Yeong Kim, Keewon Yu, Kyungmi Choi, Hyung Eun Yu, Soo Jin Oh, Kiho ...
    2015 年 38 巻 8 号 p. 1161-1168
    発行日: 2015/08/01
    公開日: 2015/08/01
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    The objective of the present work was to investigate the potential for pharmacokinetic drug–drug interactions between glimepiride (GMP) and piperazine dithioctate (PDT) in rats to support the development of an orally combined product of the two drugs. An LC-MS/MS bioanalytical method was developed for simultaneous quantification of GMP and thioctic acid (TA) in rat plasma. The accuracy, precision, linearity, selectivity, and recovery were all within an acceptable range. The oral plasma exposure of the GMP solution was more than 14-times greater than that of the GMP suspension at a dose of 0.5 mg/kg, suggesting a dissolution-limited absorption of the GMP suspension. Oral co-administration of PDT (72 mg/kg) with GMP suspension (0.5 mg/kg) reduced the plasma GMP exposure by approximately 80% without a significant change in t1/2 and tmax. Oral co-administration of PDT with GMP solution had no significant effect on the plasma pharmacokinetics of GMP. PDT lowered the pH (from ca. 7 to 5.6) and the dissolved GMP concentration in the GMP suspension. It was also shown that GMP was more soluble at pH 7 than at 5.7 in an aqueous solution, and the oral plasma exposure of a GMP suspension at pH 7.0 was substantially higher than that of a suspension at pH 5.7. These results suggest that the pH-dependent solubility of GMP was likely responsible for PDT’s effect on the oral absorption of GMP. In conclusion, the current work suggests a possibility of drug–drug interaction between GMP and PDT upon oral co-administration.
  • Kentaro Ito, Yuri Saito, Kinya Ashida, Taketo Yamaji, Hiroyuki Itoh, M ...
    2015 年 38 巻 8 号 p. 1169-1174
    発行日: 2015/08/01
    公開日: 2015/08/01
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    A fluid-retention effect is required for beverages that are designed to prevent dehydration. That is, fluid absorbed from the intestines should not be excreted quickly; long-term retention is desirable. Here, we focused on the effect of milk protein on fluid retention, and propose a new effective oral rehydration method that can be used daily for preventing dehydration. We first evaluated the effects of different concentrations of milk protein on fluid retention by measuring the urinary volumes of rats fed fluid containing milk protein at concentrations of 1, 5, and 10%. We next compared the fluid-retention effect of milk protein-enriched drink (MPD) with those of distilled water (DW) and a sports drink (SD) by the same method. Third, to investigate the mechanism of fluid retention, we measured plasma insulin changes in rats after ingesting these three drinks. We found that the addition of milk protein at 5 or 10% reduced urinary volume in a dose-dependent manner. Ingestion of the MPD containing 4.6% milk protein resulted in lower urinary volumes than DW and SD. MPD also showed a higher water reabsorption rate in the kidneys and higher concentrations of plasma insulin than DW and SD. These results suggest that increasing milk protein concentration in a beverage enhances fluid retention, which may allow the possibility to develop rehydration beverages that are more effective than SDs. In addition, insulin-modifying renal water reabsorption may contribute to the fluid-retention effect of MPD.
  • Ning Liu, Junfeng Shi, Ying Xiao, Misato Yasue, Yoshinori Takei, Hayat ...
    2015 年 38 巻 8 号 p. 1175-1184
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Glomerulonephritis (GN) is a set of pathological conditions that result in the destruction of glomeruli and loss of renal function, commonly leading to the development of end-stage renal disease. Current pharmacotherapy is limited to immunosuppressive therapy. In the present study, we found a novel antinephritic effect of a tricaprylin emulsion in the anti-glomerular basement membrane (anti-GBM) GN rat model. We evaluated the treatment in vivo by comparing administration of the emulsion with administration of a casein kinase II (CK2) inhibitor in this rat model, and performed a gene ontology-based microarray analysis to reveal in silico the detailed mechanism of action. Our results showed that administration of the tricaprylin emulsion, or even tricaprylin alone, significantly ameliorated the anti-GBM antibody-induced renal dysfunction in these rats. We believe that tricaprylin is the key active antinephritic component of the emulsion and might be a promising drug for the effective treatment of nephritis. Moreover, with respect to microarray analysis, we developed a generally applicable and rapid method to compare gene expression profile data for multiple models of nephritis and clinical samples from a public domain microarray database.
  • Yu Sakurai, Takashi Matsuda, Tomoya Hada, Hideyoshi Harashima
    2015 年 38 巻 8 号 p. 1185-1191
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Plasmid DNA (pDNA) is expected to be a new class of medicine for treating currently incurable diseases. To deliver these nucleic acids, we developed a liposomal delivery system we have called a multifunctional envelope-type nano device (MEND). In this report, we demonstrate that a MEND containing a pH-sensitive cationic lipid, YSK05 (YSK-MEND), efficiently delivered pDNA via systemic injection, and that its expression was highly dependent on the encapsulation state of the pDNA. In the preparation, the pH, ionic strength, and sodium chloride (NaCl) concentration of the lipid/pDNA mixture strongly affected the encapsulation efficiency of pDNA. Additionally, the transgene expression of luciferase in the liver by the injected YSK-MEND was dependent on the encapsulation state of pDNA rather than the nature of the YSK-MEND. Confocal laser scanning microscopy findings revealed that injection of the YSK-MEND led to homogenous gene expression in the liver compared to injection via the hydrodynamic tail vein (HTV). Concerning the safety of the YSK-MEND, a transient increase in the activity of liver enzymes was observed. However, no significant adverse events were observed. Taken together, the YSK-MEND represents a potentially attractive therapy for the treatment of various hepatic diseases.
  • Hiroaki Ikesue, Hiroyuki Watanabe, Megumi Hirano, Ayako Chikamori, Kim ...
    2015 年 38 巻 8 号 p. 1192-1198
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Pemetrexed plus carboplatin therapy is widely administered to patients with non-squamous non-small cell lung cancer. Although severe neutropenia is often observed during this combination therapy, its predictive factors are unknown. Therefore, we investigated the predictive factors for severe neutropenia in 77 patients treated with this combination therapy at the Department of Respiratory Medicine, Kyushu University Hospital, between September 2009 and September 2013. All data were retrospectively collected from the electronic medical record system, and univariate and multivariate logistic regression analyses were performed to identify risk factors for grade 3 or 4 neutropenia. Among the 77 patients, 34 (44%) developed grade 3 or 4 neutropenia. Multivariate analysis revealed that lower baseline hemoglobin values (odds ratio [OR], 1.97 per 1 g/dL decrease; 95% confidence interval [CI], 1.39–2.99, p<0.01) and lower baseline neutrophil counts (OR, 1.71 per 1000/mm3 decrease; 95% CI, 1.14–2.71, p=0.01) were significantly associated with grade 3 or 4 neutropenia. During 4 courses of pemetrexed plus carboplatin therapy, the incidence of grade 3 or 4 neutropenia in patients with baseline hemoglobin values of <11.6 g/dL was significantly higher than that in patients with values of ≥11.6 g/dL [84% (16/19) vs. 31% (18/58), p<0.001]. In conclusion, patients with lower baseline neutrophil counts or lower baseline hemoglobin values, especially those with baseline hemoglobin values of <11.6 g/dL, should be monitored more carefully during pemetrexed plus carboplatin therapy.
  • Gyeong-Je Lee, In-A Cho, Kyeong-Rok Kang, Do Kyung Kim, Hong-Moon Sohn ...
    2015 年 38 巻 8 号 p. 1199-1207
    発行日: 2015/08/01
    公開日: 2015/08/01
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    The aim of this study was to examine the anabolic and anticatabolic functions of bavachin in primary rat chondrocytes. With bavachin treatment, chondrocytes survived for 21 d without cell proliferation, and the proteoglycan content and extracellular matrix increased. Short-term monolayer culture of chondrocytes showed that gene induction of both aggrecan and collagen type II, major extracellular matrix components, was significantly upregulated by bavachin. The expression and activities of cartilage-degrading enzymes such as matrix metalloproteinases and a disintegrin and metalloproteinase with thrombospondin motifs were inhibited significantly by bavachin, while tissue inhibitors of metalloprotease were significantly upregulated. Bavachin inhibits the expression of inducible nitric oxide synthase, a representative catabolic factor, and downregulated the expression of nitric oxide, cyclooxygenase-2, and prostaglandin E2 in a dose-dependent manner in chondrocytes. Our results suggest that the bavachin has anabolic and potent anticatabolic biological effects on chondrocytes, which may have considerable promise in treating articular cartilage degeneration in the future.
  • Da Hye Hong, Hongliang Li, Han Sol Kim, Hye Won Kim, Sung Eun Shin, Wo ...
    2015 年 38 巻 8 号 p. 1208-1213
    発行日: 2015/08/01
    公開日: 2015/08/01
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    We demonstrated the inhibitory effect of fluvoxamine, a selective serotonin reuptake inhibitor (SSRI), on voltage-dependent K+ (Kv) channels in freshly isolated rabbit coronary arterial smooth muscle cells using a whole-cell patch clamp technique. Fluvoxamine reduced the amplitude of Kv currents in a concentration-dependent manner with an IC50 value of 3.71±1.09 µM and a Hill coefficient of 0.62±0.14. Although fluvoxamine did not significantly affect the steady-state activation curve, it shifted the steady-state inactivation curve toward a more negative potential. Pretreatment with another SSRI, paroxetine, did not affect the basal Kv current and did not alter the inhibitory effect of fluvoxamine on Kv channels. We concluded that fluvoxamine inhibits the Kv current in a concentration-dependent manner and in a closed (inactivated) state of the Kv channels independent of serotonin reuptake inhibition.
  • Tadanobu Takahashi, Maiko Takano, Yuuki Kurebayashi, Takashi Agarikuch ...
    2015 年 38 巻 8 号 p. 1214-1219
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Human parainfluenza virus type 1 (hPIV1) does not form clear plaque by the conventional plaque formation assay because of slightly a cytopathic effects in many cell lines infected with hPIV1, thus making in virus titration, isolation and inhibitor evaluation difficult. We have succeeded in fluorescent histochemical visualization of sialidase activities of influenza A and B viruses, Newcastle disease virus and Sendai virus by using a novel fluorescent sialidase substrate, 2-(benzothiazol-2-yl)-4-bromophenyl 5-acetamido-3,5-dideoxy-α-D-glycero-D-galacto-2-nonulopyranosidonic acid (BTP3-Neu5Ac). In this study, we applied the BTP3-Neu5Ac assay for rapid detection of hPIV1 and hPIV type 3. The BTP3-Neu5Ac assay could histochemically visualize dot-blotted hPIVs on a membrane and hPIV-infected cells as local fluorescence under UV irradiation. We succeeded in distinct fluorescent visualization of hPIV1-infected cells in only 3 d using the BTP3-Neu5Ac assay. Due to there being no fixation, hPIV1 was isolated directly from fluorescent stained focus cells by the BTP3-Neu5Ac assay. Establishment of a sensitive, easy, and rapid fluorescent focus detection assay for hPIV, hPIV1 in particular will contribute greatly to progress in hPIV studies.
  • Tadanobu Takahashi, Sawako Kawagishi, Hiroki Funahashi, Nonoka Hayashi ...
    2015 年 38 巻 8 号 p. 1220-1226
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Cytidine monophosphate (CMP) N-acetylneuraminic acid (Neu5Ac) hydroxylase (CMAH) is an essential enzyme for N-glycolylneuraminic acid (Neu5Gc) synthesis. In humans, Neu5Gc cannot be synthesized because of a deletion in the CMAH gene. Since Neu5Gc research has not been actively performed in comparison with Neu5Ac research, little is known about the function of Neu5Gc. Possible reasons are that CMAH for controlling Neu5Gc synthesis is not understood well at the molecular level, that commercial Neu5Gc is expensive, and that addition of exogenous Neu5Gc to glycoconjugates is not a general method because of the difficulty in obtaining CMAH. One solution to these problems is to achieve large-scale production of CMAH with enzymatic activity. To produce and purify CMAH as simply as possible, we generated simian CMAH as a secretory protein with a histidine tag using a baculovirus protein expression system. After culture of baculovirus-infected cells in serum-free medium, secretory simian CMAH (approximately 180 µg) was highly purified from the supernatant (150 mL) of cell culture. HPLC analysis showed conversion of CMP-Neu5Ac to CMP-Neu5Gc by the secretory CMAH. We succeeded in producing secretory CMAH with enzymatic activity that is easy to purify. In addition, peptide-N-glycosidase F treatment of CMAH indicated that secretory CMAH was a glycoprotein with N-glycan. It will also contribute to research on Neu5Gc function by easy-to-use methods for controlling Neu5Gc synthesis, for exogenous addition of Neu5Gc to glycoconjugates and by application to industrial Neu5Gc synthesis.
  • Kyeong Ha Kang, Bonggi Lee, Sujin Son, Hwi Young Yun, Kyoung Mi Moon, ...
    2015 年 38 巻 8 号 p. 1227-1233
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Inhibiting tyrosinase is an important goal to prevent melanin accumulation in skin and thereby to inhibit pigmentation disorders. Therefore, tyrosinase inhibitors are an attractive target in cosmetics and treatments for pigmentation disorders. However, only a few tyrosinase inhibitors are currently available because of their toxic effects to skin or lack of selectivity and stability. Here, we newly synthesized thirteen (Z)-2-(benzo[d]thiazol-2-ylamino)-5-(substituted benzylidene)thiazol-4(5H)-one derivatives and examined their effect on melanogenesis. Of these compounds, MHY2081 had the strongest inhibitory effect on tyrosinase without cytotoxicity in B16F10 melanoma cells. Consistently, melanogenesis was notably decreased by MHY2081 treatment. As an underlying mechanism, docking simulation showed that compared to kojic acid, a well-known competitive tyrosinase inhibitor which forms a hydrogen bond and aromatic interaction with tyrosinase, MHY2081 has stronger affinity with tyrosinase by forming three hydrogen bonds and a hydrophobic interaction with residues of tyrosinase. In parallel with this, Lineweaver–Burk plot analysis showed that MHY2081 is a strong competitive inhibitor of tyrosinase. In conclusion, MHY2081 may be a novel tyrosinase inhibitor for prevention and treatment of pigmentation disorders.
Notes
  • Yang Liu, Wei Wang, Yuyao Li, Yunqi Xiao, Jian Cheng, Jia Jia
    2015 年 38 巻 8 号 p. 1234-1239
    発行日: 2015/08/01
    公開日: 2015/08/01
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    5-Lipoxygenase (5-LOX) inhibitors have been shown to be protective in several neurodegenerative disease models; however, the underlying mechanisms remain unclear. We investigated whether 5-LOX inhibitor zileuton conferred direct neuroprotection against glutamate oxidative toxicity by inhibiting ferroptosis, a newly identified iron-dependent programmed cell death. Treatment of HT22 mouse neuronal cell line with glutamate resulted in significant cell death, which was inhibited by zileuton in a dose-dependent manner. Consistently, zileuton decreased glutamate-induced production of reactive oxygen species but did not restore glutamate-induced depletion of glutathione. Moreover, the pan-caspase inhibitor Z-Val-Ala-Asp(OMe)-fluoromethyl ketone (ZVAD-fmk) neither prevented HT22 cell death induced by glutamate nor affected zileuton protection against glutamate oxidative toxicity, suggesting that zileuton did not confer neuroprotection by inhibiting caspase-dependent apoptosis. Interestingly, glutamate-induced HT22 cell death was significantly inhibited by the ferroptosis inhibitor ferrostatin-1. Moreover, zileuton protected HT22 neuronal cells from erastin-induced ferroptosis. However, we did not observe synergic protective effects of zileuton and ferrostatin-1 on glutamate-induced cell death. These results suggested that both the 5-LOX inhibitor zileuton and the ferropotosis inhibitor ferrostatin-1 acted through the same cascade to protect against glutamate oxidative toxicity. In conclusion, our results suggested that zileuton protected neurons from glutamate-induced oxidative stress at least in part by inhibiting ferroptosis.
  • Naoya Koizumi, Yoshiaki Yamagishi, Takamasa Hirai, Makiko Fujii, Hiroy ...
    2015 年 38 巻 8 号 p. 1240-1244
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Adenovirus (Ad) vectors are widely used in gene therapy and in vitro/in vivo gene transfer. However, Ad-mediated gene transfer in epithelial cells shows low efficiency, because Ad fiber cannot bind to the primary receptor, the coxsackievirus and adenovirus receptor (CAR), present in tight junctions. Caco-2 monolayer cells cultured on Transwell-chamber plates for approximately 2 weeks are widely used for drug membrane permeation studies, but Ad-mediated gene transfer is difficult in Caco-2 monolayer cells. First, we examined the efficiency of gene transfer into Caco-2 monolayer cells. Luciferase production in cultured Caco-2 cells transduced with Ad vectors was 20-fold lower on day 12 than on day 1. In contrast, the expression of CAR protein in Caco-2 cells gradually increased along with the duration of culture. For efficient gene transfer into Caco-2 monolayer cells, the binding ability of Ad vectors with CAR was found to be important. Capric acid (C10), a medium-chain fatty acid is a tight-junction modulator used as a pharmaceutical agent. We found that a novel gene transfer method using transduction with Ad vectors in the presence of C10 led more efficiently to LacZ expression in Caco-2 monolayer cells than Ad vectors alone. The results of the present study indicate that C10 could be very useful for Ad-mediated gene transfer in human colonic Caco-2 epithelial cells.
  • Eduardo Madrigal-Bujaidar, Isela Álvarez-González, Eduardo Osiris Madr ...
    2015 年 38 巻 8 号 p. 1245-1249
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Duloxetine is a widely used antidepressant worldwide. In the present report, we evaluated its capacity to induce micronucleated polychromatic erythrocytes (MNPEs) and micronucleated normochromatic erythrocytes (MNNEs) in mice. Two assays were performed, one with a single chemical administration and the other with daily chemical administration. In the first, we administered the antidepressant once to groups of 5 mice by the intragastric (i.g.) route (2, 20, and 200 mg/kg) and performed the analysis at 24, 48, and 72 h postadministration. A control group administered i.g. distilled water was included in the assay, as well as another treated with the micronuclei-inducing chemical daunorubicin (2.5 mg/kg, injected intraperitoneally (i.p.)). In this assay, we found significant damage induced by duloxetine starting from the first time evaluated, showing the highest MNPE increase at the end of the assay. We observed a saturation effect as well, suggested by a decreasing relative efficiency with respect to each tested dose. In a second assay, we administered the antidepressant i.g. every day for 5 weeks (2, 6, and 12 mg/kg), and micronuclei analysis was performed at the end of each week. In this study, we also found a significant increase in both MNPEs and MNNEs which was clear by the second week of administration. Our results suggest that short-term as well as cumulative damage is produced by duloxetine. Thus, confirmation of the observed genotoxic potential in other models seems advisable, as well as caution when prescribing this antidepressant.
  • Mohamed Abo El Hamd, Mitsuhiro Wada, Rie Ikeda, Shigeru Kawakami, Keni ...
    2015 年 38 巻 8 号 p. 1250-1253
    発行日: 2015/08/01
    公開日: 2015/08/01
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    Propofol (PRO) is a hypnotic used to induce and maintain general anesthesia. A risk of drug–drug interactions exists in cases of clinical co-administration of PRO and midazolam (MDZ) or carbamazepine (CBZ). Therefore a sensitive and rapid assay is needed to monitor these drugs. In this study, a sensitive and selective liquid chromatography-tandem mass spectrometry technique was developed for simultaneous determination of PRO, MDZ, and CBZ in plasma. Simultaneous selected reaction monitoring in the positive and negative ionization modes was used for mass detection. Analytes were isolated from plasma samples by a simple, economic, and rapid solid-phase extraction method. Chromatographic separations were achieved using a Chromolith Performance RP-18e analytical column (100×4.6 mm i.d.) with a mixture of acetonitrile–ammonium acetate buffer (10 mM, pH 3.5) (90 : 10, v/v) as the mobile phase. The method was fully validated for PRO, MDZ, and CBZ over concentrations ranging at 1–100, 2–100, and 7–1000 ng/mL, respectively, with acceptable validation parameters. Furthermore, the method was applied to monitor PRO and MDZ or CBZ following co-administration in rats.
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