Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
35 巻 , 7 号
選択された号の論文の30件中1~30を表示しています
Regular Articles
  • Mohammad Iqbal Hossain Bhuiyan, Joo Youn Kim, Tae Joung Ha, Seong Yun ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 999-1008
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    The present study investigated the neuroprotective effects of anthocyanins extracted from black soybean (cv. Cheongja 3, Glycine max (L.) MERR.) seed coat against oxygen–glucose deprivation (OGD) and glutamate-induced cell death in rat primary cortical neurons. Lactate dehydrogenase (LDH) release and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assays were employed to assess cell membrane damage and viability of primary neurons, respectively. OGD-induced cell death in 7 d in vitro primary cortical neurons was found to be OGD duration-dependent, and approximately 3.5 h of OGD resulted in ≈60% cell death. Treatment with black soybean anthocyanins dose-dependently prevented membrane damage and increased the viability of primary neurons that were exposed to OGD. Glutamate-induced neuronal cell death was dependent on the glutamate concentration at relatively low concentrations and the number of days the cells remained in culture. Interestingly, black soybean anthocyanins did not protect against glutamate-induced neuronal cell death. They did, however, inhibit the excessive generation of reactive oxygen species (ROS) and preserve mitochondrial membrane potential (MMP) in primary neurons exposed to OGD. In agreement with the neuroprotective effect of crude black soybean anthocyanins, purified cyanidin-3-glucoside (C3G), the major component of anthocyanins, also offered dose-dependent neuroprotection against OGD-induced neuronal cell death. Moreover, black soybean C3G markedly prevented excessive generation of ROS and preserved MMP in primary neurons that were exposed to OGD. Collectively, these results suggest that the neuroprotection of primary rat cortical neurons by anthocyanins that were extracted from black soybean seed coat might be mediated through oxidative stress inhibition and MMP preservation but not through glutamate-induced excitotoxicity attenuation.
  • Junko Ishizaki, Tomoko Tsuda, Yukio Suga, Satsuki Ito, Kunizo Arai, Yo ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1009-1013
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Changes of mycophenolic acid (MPA) pharmacokinetics with aging were investigated in rats. We also compared the effect of concomitant amoxicillin/clavulanate combination (CVA/AMPC) on the pharmacokinetics of MPA in 4-week-old and 12-week-old rats (the package insert of CVA/AMPC warns of possible interaction with MPA). Four-week-old rats showed a 1.4-fold higher total body clearance of MPA and a lower volume of distribution of MPA (65%), compared to the values in 12-week-old rats. However, the difference in MPA pharmacokinetics disappeared when enterohepatic circulation was eliminated by bile duct cannulation (BDC). Concomitant CVA/AMPC significantly reduced plasma MPA concentration in intact rats of both age groups, and the age-dependent difference of MPA pharmacokinetics was no longer apparent. The effect of CVA/AMPC was not seen in rats that had undergone BDC, suggesting that the drug–drug interaction can be attributed to inhibition of enterohepatic circulation by CVA/AMPC. These results indicate that the aging-related alteration of MPA pharmacokinetics is a consequence of immature enterohepatic circulation in 4-week-old rats. Higher doses of MPA may be necessary in juveniles.
  • Hiroshi Yukawa, Satoru Ishikawa, Takashi Kawanishi, Makoto Tamesada, H ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1014-1021
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Lentinula edodes mycelia (L.E.M.) is a dried powder extracted from shiitake mushrooms (Lentinula edodes). We previously demonstrated that it has immunomodulatory effects. In this paper, the direct cytotoxic effects of the polysaccharide-rich fraction of L.E.M. (L.E.M. ethanol precipitate; LEP) on HepG2 human hepatocellular carcinoma (HCC) cells were investigated. LEP directly killed the HepG2 cells efficaciously, but had only minor effects on normal rat hepatocytes and normal mouse dermal cells under the same conditions. Characteristic morphological changes associated with apoptosis such as shrinkage, rounding, and floating as well as chromatin condensation were confirmed; terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) staining was positive as determined by fluorescence microscopy analyses. The caspase-3 and -8 death receptor pathway was found largely responsible for the apoptotic death of HepG2 cells treated with LEP. In conclusion, LEP can directly induce apoptosis of HepG2 cells, and thus may have potential chemotherapeutic applications for the treatment of HCC.
  • Young-Hoon Song, Soo-Jin Jeong, Hee-Young Kwon, Bonglee Kim, Sung-Hoon ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1022-1028
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Although ursolic acid isolated from Oldenlandia diffusa (Rubiaceae) was known to have anticancer activities in prostate, breast and liver cancers, the underlying mechanism of ursolic acid in ovarian cancer cells was not investigated so far. In the present study, the apoptotic mechanism of ursolic acid was elucidated in SK-OV-3 ovarian cancer cells by 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay, cell cycle analysis and Western blotting. Ursolic acid exerted cytotoxicity against SK-OV-3 and A2780 ovarian cancer cells with IC50 of ca. 50 and 65 µM, respectively. Apoptotic bodies were observed in ursolic acid treated SK-OV-3 cells. Also, ursolic acid significantly increased ethidium homodimer stained cells and sub-G1 apoptotic portion in SK-OV-3 cells. Consistently, Western blotting revealed that ursolic acid effectively cleaved poly(ADP-ribose) polymerase (PARP), caspase-9 and -3, suppressed the expression of survival genes such as c-Myc, Bcl-xL and astrocyte elevated gene (AEG)-1, and upregulated phosphorylation of extracellular signal-regulated kinase (ERK) in SK-OV-3 cells. Interestingly, ursolic acid suppressed β-catenin degradation as well as enhanced phosphorylation of glycogen synthase kinase 3 beta (GSK 3β). Furthermore, GSK 3β inhibitor SB216763 blocked the cleavages of caspase-3 and PARP induced by ursolic acid and proteosomal inhibitor MG132 disturbed down-regulation of β-catenin, activation of caspase-3 and decreased mitochondrial membrane potential (MMP) induced by ursolic acid in SK-OV-3 cells. Overall, our findings suggest that ursolic acid induces apoptosis via activation of caspase and phosphorylation of GSK 3β in SK-OV-3 cancer cells as a potent anti-cancer agent for ovarian cancer therapy.
  • Cao Gao, Dun-Quan Xu, Chang-Jun Gao, Qian Ding, Li-Nong Yao, Zhi-Chao ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1029-1034
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Hemorrhagic shock (HS) is a common condition and leading cause of death in trauma patients universally. Severe inflammatory responses during HS finally lead to multiple-organ failure. Hydrogen sulphide (H2S) is increasingly recognized as an important signaling molecule with various protective effects. In the present study, we investigated the antiinflammatory and cardioprotective effects of an exogenous H2S donor, sodium hydrosulfide (NaHS), in an HS rat model. Male Sprague-Dawley rats were randomly divided into the sham-operated, sham-operated treated with NaHS (28 µmol/kg, intraperitoneally (i.p.)), HS, and HS treated with NaHS (28 µmol/kg, i.p.) groups. The HS groups were subjected to mimicked HS for 1 h and then treated with NaHS or left untreated. The rats were then resuscitated with Ringer lactate solution for 1 h. Myocardial enzymes and inflammatory cytokines were evaluated. Morphologic changes in cardiac tissue and ultrastructural injury were also analyzed. HS resulted in significant hemodynamic deterioration and increased myocardial enzyme and inflammatory cytokine levels. Intraperitoneal administration of NaHS significantly prevented hemodynamic deterioration and decreased the elevation of myocardial enzymes. NaHS also inhibited the nuclear factor κB inhibitor kinase (IKK)/nuclear factor κB inhibitor (IκB)/nuclear factor κB (NF-κB) signaling pathway. The results suggest that NaHS exerts cardioprotective effects against HS. The protective effects of NaHS may occur via down-regulation of the IKK/IκB/NF-κB signaling pathway.
  • Noppawan Phumala Morales, Yumiko Yamaguchi, Kimiyo Murakami, Nuttavut ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1035-1040
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Reduction of a nitroxyl radical, carbamoyl-PROXYL in association of free radical production and hepatic glutathione (GSH) was investigated in iron overloaded mice using an in vivo L-band electron spin resonance (ESR) spectrometer. Significant increases in hepatic iron, lipid peroxidation and decrease in hepatic GSH were observed in mice intraperitoneally (i.p.) administrated with ferric nitrilotriacetate (Fe(III)-NTA, a total 45 µmol/mouse over a period of 3 weeks). Free radical production in iron overloaded mice was evidenced by significantly enhanced rate constant of ESR signal decay of carbamoyl-PROXYL, which was slightly reduced by treatment with iron chelator, deferoxamine. Moreover, the rate constant of ESR signal decay was negatively correlated with hepatic GSH level (r=−0.586, p<0.001). On the other hand, hepatic GSH-depletion (>80%) in mice through daily i.p. injection and drinking water supplementation of L-buthionine-[S,R]-sulfoximine (BSO) significantly retarded ESR signal decay, while there were no changes in serum aspartate aminotransferase and liver thiobarbituric acid-reactive substances levels. In conclusion, GSH plays two distinguish roles on ESR signal decay of carbamoyl-PROXYL, as an antioxidant and as a reducing agent, dependently on its concentration. Therefore, it should be taken into account in the interpretation of free radical production in each specific experimental setting.
  • Takuro Ueki, Tatsuhiro Akaishi, Hidenobu Okumura, Kazuho Abe
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1041-1047
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Extract from fruits of Nandina domestica THUNBERG (NDE) has been used to improve cough and breathing difficulty in Japan for many years. To explore whether NDE may alleviate respiratory inflammation, we investigated its effect on expression of cyclooxygenase-2 (COX-2) and production of prostaglandin E2 (PGE2) in human pulmonary epithelial A549 cells in culture. Treatment with lipopolysaccharide (LPS; 6 µg/mL) resulted in an increase of COX-2 expression and PGE2 production in A549 cells. Both the LPS-induced COX-2 expression and PGE2 production were significantly inhibited by NDE (1–10 µg/mL) in a concentration-dependent manner. NDE did not affect COX-1 expression nor COX activity. These results suggest that NDE downregulates LPS-induced COX-2 expression and inhibits PGE2 production in pulmonary epithelial cells. Furthermore, higenamine and nantenine, two major constituents responsible for tracheal relaxing effect of NDE, did not mimic the inhibitory effect of NDE on LPS-induced COX-2 expression in A549 cells. To identify active constituent(s) of NDE responsible for the anti-inflammatory effect, NDE was introduced in a polyaromatic absorbent resin column and stepwise eluted to yield water fraction, 20% methanol fraction, 40% methanol fraction, 99.8% methanol fraction, and 99.5% acetone fraction. However, none of these five fractions alone inhibited LPS-induced COX-2 expression. On the other hand, exclusion of water fraction from NDE abolished the inhibitory effect of NDE on LPS-induced COX-2 expression. These results suggest that constituent(s) present in water fraction is required but not sufficient for the anti-inflammatory activity of NDE, which may result from interactions among multiple constituents.
  • Fumihiro Mizokami, Yusuke Murasawa, Katsunori Furuta, Zenzo Isogai
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1048-1053
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Iodoform gauze is used in clinical practice for treatment of infected wounds. However, effectiveness and action mechanism of iodoform gauze for removal of necrotic tissue are unknown. We therefore employed case control and biochemical studies in order to clarify the pharmacological activity of iodoform gauze. A clinical study demonstrated that treatment with iodoform gauze removed necrotic tissue more effectively than treatment with conventional ointments. More than 60% of iodoform gauze-treated wounds were completely debrided within 2 weeks. Consistent with the clinical observation, biochemical analyses revealed clear differences in wound fluid proteins after treatment with iodoform gauze or conventional gauze. The amount of macroaggregates of type I collagen from wounds were remarkably decreased in iodoform gauze. Moreover, iodoform gauze and iodoform itself released non-aggregative type I collagen from necrotic debris in vitro. Taken together, we conclude that iodoform gauze efficiently removes necrotic tissue by its lytic activity for collagen fibers.
  • Ji-Young Hong, Hye-Jin Boo, Jung-Il Kang, Min-Kyoung Kim, Eun-Sook Yoo ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1054-1063
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    We observed that (1S,2S,3E,7E,11E)-3,7,11,15-Cembratetraen-17,2-olide (LS-1), marine cembrenolide diterpene, inhibited growth and induced apoptosis in colon cancer cells via a reactive oxygen species (ROS) dependent mechanism. Treatment of HT-29 cells with LS-1 resulted in ROS generation, which was accompanied by disruption of mitochondrial membrane potential, cytosolic release of cytochrome c, sub-G1 peak accumulation, activation of Bid, caspase-3, -8, and -9, and cleavage of poly(ADP-ribose) polymerase (PARP) along with the suppressive expression of B cell lymphoma-2 (Bcl-2). All these effects were significantly blocked on pretreatment with the ROS inhibitor N-acetylcysteine (NAC), indicating the involvement of increased ROS in the proapoptotic activity of LS-1. Moreover, we showed that LS-1 induced the phosphorylation of c-Jun N-terminal kinase (JNK) and dephosphorylation of p38, extracellular signal-regulated kinase (ERK), Akt, Src and signal transducer and activator of transcription (STAT)3, which were effectively attenuated by NAC. In addition, the expressions of antioxidant catalase and glutathione peroxidase were abrogated by treatment using LS-1 with or without NAC. These findings reveal the novel anticancer efficacy of LS-1 mediated by the induction of apoptosis via ROS generation in human colon cancer cells.
  • Makoto Oba, Masakazu Tanaka
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1064-1068
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    A protein transfection reagent is a powerful tool for elucidating a protein function in a cell, and plays an important role in the fields of cell biology and drug discovery. Many researchers have developed protein delivery systems and several systems are commercially available. In this study, we focus on the biological functions of three commercially available protein transfection reagents, Pro-DeliverIN, Xfect, and TurboFect, especially in their internalization routes by HeLa cells. A cellular uptake study using specific endocytosis inhibitors and confocal laser scanning microscope observation revealed that each reagent was internalized into HeLa cells by different mechanism. It is our hope that the results presented here will help in the choice and use of protein transfection reagents for experiments.
  • Ji Hyun Shin, So Jung Park, Eun Sung Kim, Yoon Kyung Jo, Jungwoo Hong, ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1069-1075
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Autophagy is associated with cell survival and cell death. Autophagy is implicated in the pathophysiology of various human diseases. In order to identify autophagy regulatory molecules, we screened a chemical drug library in SH-SY5Y cells and selected Sertindole as a potent autophagy inducer. Sertindole was developed as an antipsychotic drug for Schizophrenia. Sertindole treatment highly induced the formation of autophagosomes as well as LC3 conversion. Subsequently, Sertindole-induced autophagy was efficiently suppressed by down regulation of ATG5. Sertindole also increased reactive oxygen species (ROS) production, which contributes to autophagy-associated cell death in neuroblastoma cells. ROS scavengers such as N-acetylcysteine and Trolox suppressed not only ROS generation but also autophagy activation by Sertindole. These results suggest Sertindole induces autophagy and autophagy-associated cell death by ROS production in neuroblastoma cells.
  • Tae Hyung Kim, Chan Woong Park, Hoi Yun Kim, Moon Hyuk Chi, Sung Kyung ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1076-1083
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    An intranasally active glucagon-like peptide-1 (GLP-1) formulation would have great advantages over conventional injectable therapies for the treatment of diabetic patients. The purpose of this study was to investigate the biological potentials of PEGylated exendin-4 (PEG-Ex4) analogs administered intranasally and the effects of polyethylene glycol (PEG) molecular weight (1, 2, 5 kDa) on nasal absorption. Initially, PEG-Ex4 analogs were site-specifically PEGylated to Lys27-amine, and their bioactivities and stabilities were studied in vitro. The hypoglycemic effects and pharmacokinetics of these analogs after nasal administration were evaluated in type 2 diabetic animal models. PEG-Ex4 analogs had 3.1-, 3.8-, and 5.9-fold increased stabilities in rat nasal homogenates than Ex4. However, Lys27-PEG1k-Ex4 was found to have well-preserved bioactivities (83.3% potency vs. Ex4), and other analogs were found to have much lower bioactivities than Lys27-PEG1k-Ex4. In particular, the in vivo pharmacokinetic parameters of Lys27-PEG1k-Ex4 in intranasally administered rats were significantly improved by PEGylation. Area under the curve (AUC) values of Lys27-PEG1k-Ex4 were 33.6-fold higher and circulating t1/2 values was 27.1-fold higher than Ex4. But, other analogs were not effectively absorbed via the intranasal route, because the higher molecular weight PEG (over 2 kDa) limited intranasal absorption. Finally, in vivo hypoglycemic experiment showed that Lys27-PEG2k-, Lys27-PEG5k-Ex4 had significantly lower hypoglycemic efficacies than Lys27-PEG1k-Ex4, probably because of their lower intrinsic bioactivities and intranasal absorptions. Taken together, our findings suggest that the site-specific conjugation of appropriately sized PEG (1 kDa) substitution onto peptides like Ex4 offers two advantages for delivery via the intranasal route, namely, increased stability and extended circulating half-life.
  • Bimit Mahat, Jung-woo Chae, In-hwan Baek, Gyu-yong Song, Jin-sook Song ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1084-1090
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Angelica gigas NAKAI is used to treat dysmenorrhea, amenorrhea, menopause, abdominal pain, injuries, migraine, and arthritis. The present study provided a physicochemical and toxicological characterization of compounds in A. gigas NAKAI (decursin, decursinol angelate, diketone decursin, ether decursin, epoxide decursin and oxim decursin). Diketone decursin (173.16 µg/mL) and epoxide decursin (122.12 µg/mL) exhibited >100 µg/mL kinetic solubility after applying nephelometry, suggesting a highly soluble compound. The Student’s t-test revealed significant differences in the pKa ranges of the compounds by automatic titration from capillary electrophoresis (p<0.05). Diketone decursin, epoxide decursin and oxim decursin might be formulated into an oral dosage form (log P: 0–3) by an automatic titration analysis. A parallel artificial membrane permeability assay demonstrated permeability coefficients of <10×10−6 cm/s for all of the compounds, suggesting poor permeability. Ether decursin exhibited a toxic effect after being applied to mouse (NIH 3T3, EC50: 57.9 µM) and human (HT-29, EC50: 36.1 µM; Hep-G2, EC50: 4.92 µM) cells. Additionally, epoxide and oxim decursin were toxic through acute oral toxicity (four and three deaths of Institute of Cancer Research (ICR) mice) and mutation toxicity testing by applying Salmonella typhimurium cells with and without S9. Although diketone decursin exhibited less permeability, it is potentially valuable pharmacological compound that should be investigated.
  • Reiko Ishii-Nozawa, Mitsuo Mita, Masaru Shoji, Koichi Takeuchi
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1091-1095
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    The Goto–Kakizaki (GK) rat is a non-obese and spontaneous model of mild Type 2 diabetes mellitus. In the present study, we compared the regulatory mechanisms of endogenous norepinephrine (NE) release from sympathetic nerves of caudal arteries of 12-week-old GK rats and age-matched normal Wistar rats. Electrical stimulation (ES) evoked significant NE release from caudal arteries of Wistar and GK rats. The amounts of NE released by ES were almost equal in Wistar and GK rats, although the NE content in caudal artery of GK rats was significantly lower than that of Wistar rats. We examined the effects of an α2-adrenoceptor agonist, clonidine (CLO), and an α2-adrenoceptor antagonist, yohimbine (YOH), on the release of endogenous NE evoked by ES. CLO significantly reduced NE release from caudal arteries of Wistar but not GK rats. On the other hand, YOH significantly increased NE release from both rats. Furthermore, we examined the effects of an A1-adenosine receptor agonist, 2-chloroadenosine (2CA), and an A1-adenosine receptor antagonist, 8-sulfophenyltheophylline (8SPT), on the release of endogenous NE evoked by ES. 2CA significantly reduced NE release from caudal arteries of Wistar but not GK rats. On the other hand, 8SPT did not affect NE release from both rats. These results suggest that the dysfunction of negative feedback regulation of NE release via presynaptic receptors on sympathetic nerves in GK rats may be involved in the autonomic nervous system dysfunction associated with diabetic autonomic neuropathy.
  • Kazuyuki Furuya, Noriko Yamamoto, Yuki Ohyabu, Akito Makino, Teruyuki ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1096-1104
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    We have studied non-steroidal selective androgen receptor modulators (SARMs) to develop anti-osteoporosis drugs for males and females. Many SARMs have been studied for their anabolic effects on bone or muscle with reduced virilizing effects in male animals. However, the tissue selectivities of these agents in female animals have not been fully evaluated. We evaluated the novel SARM S-101479 from tetrahydroquinoline libraries in ovariectomized (OVX) rats. S-101479 preferentially bound to the androgen receptor with nanomolar affinity among nuclear receptors. It increased the bone mineral density (BMD) of femurs and diminished the effects on the uterus and clitoral gland in OVX rats. We then compared the effect of S-101479 on bone with those of commercial anti-osteoporosis drugs such as alendronate, raloxifene, and teriparatide. Furthermore, we evaluated the effects of combination treatments with these agents in OVX rats. After 16-week treatment, all agents significantly increased BMD, but the magnitude of bone mineral content (BMC) and/or bone size (projected bone area) were different. Alendronate, raloxifene, and teriparatide maintained BMC and bone size in this experimental dose. Only S-101479 increased BMC with bone size on single treatments. In combination treatment, S-101479 significantly increased BMC and bone size compared with single treatments of other agents. S-101479, like natural androgen, may have showed periosteal bone formation of the cortical area and indicated additive effects with commercial anti-osteoporosis drugs. These results indicate that S-101479 may be a useful anti-osteoporosis drug, particularly for patients with established severe osteoporosis.
  • Soichiro Akashi, Kyo Shirai, Tatsumi Okada, Kanako Konishi, Toshifumi ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1105-1117
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Treatment of PC12 cells with fungus-derived alkaloid neoechinulin A for more than 12 h renders the cells resistant to subsequent superoxide (O2)/nitric oxide (NO) insults derived from 3-morpholinosydnonimine (SIN-1). However, the underlying mechanism(s) remains largely unclear. To elucidate the mechanism(s), we assessed the specificity of the cytoprotection afforded by neoechinulin A treatment using other cytocidal stressors and also clarified the resulting cellular alterations, focusing on the antioxidant and metabolic enzymes systems. Neoechinulin A treatment for more than 12 h endowed PC12 cells with significant resistance to transient NO toxicity, but not persistent NO toxicity, bolus H2O2 toxicity, or oxidative insult from the redox cycling quinone menadione. Cellular antioxidant system profiling revealed no substantial potentiation of the activity of any antioxidant enzyme in lysate from the neoechinulin A-treated cells excluding glutathione (GSH) content, which was significantly decreased (>50%), resulting in a proportional compromise in the thiol-reducing activity of the intact cells. In addition, no differences were observed in the activity for any nicotinamide adenine dinucleotide (phosphate) reduced form (NAD(P)H)-generating enzyme, steady-state NAD(P)H/nicotinamide adenine dinucleotide (phosphate) oxidized form (NAD(P)+) ratios, or the levels of total NAD(P)H. Nevertheless, the neoechinulin A-treated intact cells exhibited increased NAD(P)H redox turnover when driven by extracellular tetrazolium. The structurally inactive analog preechinulin failed to protect cells against NO toxicity or induce these alterations, suggesting their link with the cytoprotective mechanism. These results suggest that neoechinulin A, despite disabling the GSH defense system, confers cytoprotection against nitrosative stresses by elevating the cellular reserve capacity for NAD(P)H generation, which could offset crippling of energy-supplying systems due to nitrosative stress.
  • Gaigai Deng, Junzhi Wang, Qiaoyin Zhang, Haibo He, Fangfang Wu, Tianya ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1118-1125
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    The present study was to study the hepatoprotective effects of phloridzin (PHL) on hepatic fibrosis induced by carbon tetrachloride (CCl4) in rats, on the basis of this investigation, the possible mechanism of PHL was elucidated. Male Sprague Dawley (SD) rats were randomly divided into six groups: control, model, PHL-L, PHL-M, PHL-H and colchine. All rats except control group were intraperitoneally injected with CCl4, and control rats were injected with olive oil, twice a week for eight weeks. At the same time, the rats were orally given homologue drugs once a day, respectively. Hepatoprotective effects of PHL were evaluated by liver weight indexes, biochemical values, total antioxidant capacity and total-superoxide dismutase, histopathological observations, hepatic fibrosis, and the hepatic fibrosis relative gene and protein expressions. PHL significantly improved hepatic function; remarkably decreased serum hyaluronic acid (HA), transforming growth factor-β1 (TGF-β1), aspartate aminotransferase (AST), alanine aminotransferase (ALT) and liver tissues hydroxyproline, malondialdehyde (MDA) levels, increased glutathione peroxidase (GSH-Px), total-antioxygen capacity (T-AOC) and total-superoxide dismutase (T-SOD) contents of liver tissues; Real-time polymerase chain reaction (PCR) and immunohisto-chemical results showed PHL might markedly reverse the up-regulated mRNA and protein expressions of the α-smooth muscle actin (SMA), TGF-β1 and tissue inhibitor of metalloproteinase-1 (TIMP1), up-regulate the matrix metalloproteinase-1 (MMP1) mRNA and protein expressions. Histopathological observations provided supportive evidence for biochemical analyses and the hepatic fibrosis relative gene and protein expressions, and with the dose of PHL increasing, the aforesaid improvement became more and more strong. The studies demonstrated that PHL exerted beneficially hepatoprotective effects on hepatic fibrosis induced by CCl4, mainly enhancing antioxidant capacity of liver organizations, reduce the level of lipid peroxidation induced by CCl4, and protect hepatocyte membranes from damage, and alleviate hepatic fibrosis.
  • Tetsuro Kamiya, Misato Izumi, Hirokazu Hara, Tetsuo Adachi
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1126-1131
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Propolis is a natural product made by honeybees and contains various compounds, including flavonoids, amino acids and fatty acids. These compounds are considered to have antiviral, antibacterial and antioxidative properties. On the other hand, cadmium (Cd), an industrial and environmental pollutant, preferentially accumulates in the kidney and induces kidney injury. We previously reported that exposure to CdCl2 induced cell death though intracellular reactive oxygen species (ROS) generation in kidney tubule epithelial COS7 cells. Here, we investigated whether propolis extracts suppress CdCl2-induced cytotoxicity. Predictably, pretreatment with propolis extracts significantly suppressed CdCl2-induced cytotoxicity and intracellular ROS generation. Propolis extracts not only showed superoxide dismutase and antioxidative activities, but also increased the expression of heme oxygenase-1 (HO-1), an antioxidative enzyme. Moreover, we determined the involvement of hypoxia inducible factor-1α in propolis extract-derived HO-1 induction. We demonstrate for the first time the utility of propolis for Cd-related COS7 cytotoxicity, and these novel findings are considered to contribute to the control of ROS-derived disorders.
  • Yuki Tazawa, Kazunori Matsumura, Yoh Takekuma, Mitsuru Sugawara
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1132-1136
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    In allogeneic bone marrow transplantation (allo-BMT) in patients with leukemia, the combination of VP-16 and cyclophosphamide (CY) is commonly used for the conditioning regimen. In the present study, we demonstrated schedule-dependent cytotoxicity of VP-16 and CY in K-562 cells. K-562 cells were pretreated with low concentrations (2.5 and 5 µg/mL) of 4-hydroperoxycyclophosphamide (40487S), which is a preactivated analog of CY. It was confirmed that these concentrations did not influence cell viability. Cells subsequently exposed to 0.5–100 µg/mL of VP-16 showed reduced the viability compared to that of control cells not treated with 40487S. In contrast, there was no change in the viability of K-562 cells pretreated with low concentrations (0.5 and 1 µg/mL) of VP-16. It was confirmed that these concentrations did not influence cell viability. Viability of subsequently exposed to 1–20 µg/mL was not different from that of control cells not treated with VP-16. VP-16 caused cell cycle arrest at G2/M phase. On the other hand, 40487S arrested the cell cycle at S phase. Thymidine-synchronized cells, VP-16 showed cell cycle specificity for cell killing from early-S to mid-S phase. On the other hand, 40487S showed cell cycle-independent cytotoxicity. Exposure of cells to VP-16 after 40487S induced a greater cytotoxic effect on K-562 cells. The findings may lead to improvements in clinical combination chemotherapy.
  • Waesarat Soontornchaiboon, Seong Soo Joo, Sang Moo Kim
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1137-1144
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Violaxanthin is a major carotenoid of microalgae Chlorella ellipsoidea and is also found in dark-green leafy vegetables, such as spinach. In this study, the anti-inflammatory effect of violaxanthin isolated from C. ellipsoidea was examined using lipopolysaccharide (LPS)-stimulated RAW 264.7 mouse macrophage cells. In addition, the anti-inflammatory activity and mechanism of action of purified violaxanthin was assessed using various assays, such as quantitative real-time polymerase chain reaction (PCR), Western blotting, and electrophoretic-mobility shift assay (EMSA). The results of this combined analysis revealed that violaxanthin significantly inhibited nitric oxide (NO) and the prostaglandin E2 (PGE2). Interestingly, violaxanthin effectively inhibited LPS-mediated nuclear factor-κB (NF-κB) p65 subunit translocation into the nucleus, suggesting that the violaxanthin anti-inflammatory activity may be based on inhibition of the NF-κB pathways. In conclusion, violaxanthin of C. ellipsoidea holds promise for use as a potential anti-inflammatory agent for either therapeutic or functional adjuvant purposes.
  • Tatsuji Enoki, Takanari Tominaga, Fuyuko Takashima, Hiromu Ohnogi, Hir ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1145-1149
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    We have previously reported that agaro-oligosaccharides (AGOs) suppressed the elevated levels of nitric oxide (NO), prostaglandin E2 (PGE2), and pro-inflammatory cytokines in activated monocytes/macrophages, via heme oxygenase-1 induction. In this report, we initially demonstrated that AGOs intake inhibited NO production in activated peritoneal macrophages. Then, we tested for the ability of AGOs to prevent tumor promotion on the two-stage mouse skin carcinogenesis model. As a result, AGOs feeding led to delayed tumor appearance and decreased tumor number. It is known that PGE2 is one of key players in carcinogenesis. Thus, we confirmed that PGE2 production was suppressed by AGOs intake in TPA-induced ear edema model. We also demonstrated that cyclooxygenase-2 and microsomal PGE synthase-1, rate-limiting enzymes in PGE2 production, were down-regulated by AGOs in human monocytes. Consequently, AGOs are expected to prevent tumor promotion by inhibiting PGE2 elevation in chronic inflammation site.
  • Ying Liu, Qi-Feng Shi, Min Qi, Shin-Ichi Tashiro, Satoshi Onodera, Tak ...
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1150-1158
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    The aim of this study was to elucidate the molecular mechanisms mediating hepatocyte growth factor (HGF)-induced protection against oridonin-induced apoptosis in A549 cells. Oridonin induced decrease in Bcl-2/Bax ratio and activation of caspase-3, while these processes were reversed by HGF, suggesting that HGF played an anti-apoptotic role in oridonin-induced A549 cell death. HGF-induced protective effect was partially attributed to the activation of nuclear factor (NF)-κB and cyclooxygenase 2 (COX-2), since the protective effect was abolished by inhibition of NF-κB or interruption of COX-2. Then the activated COX-2 could prevent cells from initiating the apoptotic response by promoting prostaglandin E2 (PGE2) release. Activation of NF-κB-COX-2 by HGF-treatment triggered the increase in Bcl-2/Bax ratio, inhibition of procaspase-3 cleavage, promotion of Ca2+-independent intracellular phospholipase A2 (iPLA2) expression and augmentation of PGE2 release, leading to antagonizing oridonin-induced cell death in A549 cells. HGF-induced cell survival in response to oridonin administration was associated with the activation of c-Met-NF-κB-COX-2 and c-Met-Bcl-2-caspase-3 signaling pathways. iPLA2, downstream effector of caspase-3, also participated in these processes.
  • Masayuki Takakuwa, Jun Iwamoto
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1159-1165
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Intramuscularly administered elcatonin (ECT) reduces pain via the central nervous system. A prospective study was performed to determine whether ECT has a beneficial effect on back pain and function in postmenopausal women with osteoporosis during bisphosphonate therapy. Sixty-one postmenopausal osteoporotic women with back pain (mean age: 73.7 years, range: 54–96 years) were divided into two groups: the control group (n=30) and the ECT (intramuscular, 20 units a week) group (n=31). All patients received treatment with risedronate (17.5 mg weekly). The duration of the study was 8 weeks. Urinary levels of cross-linked N-terminal telopeptides of type I collagen (NTX), visual analogue scale (VAS) for back pain at rest and movement, and Roland–Morris Disability Questionnaire (RDQ) score for function were assessed. Urinary NTX levels, VAS at rest and movement, and RDQ score markedly decreased during 8 weeks of treatment in both ECT and control groups. A significant reduction in VAS at movement, but not in VAS at rest and RDQ score, was noted in the ECT group than in the control group. This effect was observed from 2 weeks after the start of therapy. These results suggested that ECT in combination with risedronate was more effective than risedronate alone for reducing back pain in postmenopausal women with osteoporosis.
  • Kouichi Minato, Hidenori Nagao, Tetsuya Imazu, Kenjo Takahashi
    原稿種別: Regular Article
    2012 年 35 巻 7 号 p. 1166-1170
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Little is known about the kinetics and metabolism of thyroid hormones in the hypothyroid state. In order to optimize hormone replacement therapy, it is important to understand variations in the kinetics and metabolism of thyroid hormones. To investigate these factors, we monitored serum thyroxine (T4) and triiodothyronine (T3) levels in iodine-deficient diet (ID) rats using online solid-phase extraction liquid chromatography-mass spectrometry/mass spectrometry (Online SPE LC-MS/MS). Furthermore, we evaluated supply and turnover rates of T4 in ID rats using a stable isotope-labeled T4 ([13C9]T4). Although serum T4 levels gradually declined after beginning ID treatment, T3 levels were unchanged throughout the experimental period. After intravenous administration of [13C9]T4 to ID rats, [13C9]T4 levels were monitored. We previously reported that significant differences of supply and turnover rates for T4 were observed in surgically thyroidectomized (Tx) rats. Surprisingly, there were no differences of supply and turnover rates for T4 between ID rats and intact rats. In conclusion, there were significant differences of supply and turnover rates for T4 between the hypothyroid states of ID and Tx rats. In ID rats, T3 might be preferentially biosynthesized in the thyroid, and ID treatment might not affect T4 kinetics. Our method, online SPE LC-MS/MS monitoring using a stable isotope tracer, has the potential to be used as a diagnostic tool to investigate the pathogenesis of thyroid disease and is valuable for optimizing the dosage in thyroid hormone replacement therapy.
Notes
  • Feihua Wu, Hui Wang, Juan Li, Jingyu Liang, Shiping Ma
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1171-1177
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Recent data have indicated that inflammation plays an important role in the development of insulin resistance. The present study aims at examining the activity of homoplantaginin, a flavonoid from a traditional Chinese medicine Salvia plebeia R. BR., on palmitic acid (PA)-induced insulin sensitivity and the underlying mechanisms of its anti-infammatory properties in the endothelial cells. Pre-treatment of homoplantaginin on human umbilical vein endothelial cells (HUVECs) significantly inhibited PA induced tumour necrosis factor-α (TNF-α) and interleukin-6 (IL-6) mRNA expression, and inhibitory κB kinase beta (IKKβ) and nuclear factor-κB (NF-κB) p65 phosphorylation. To the PA-impaired insulin-dependent tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1) and decrease in nitric oxide (NO) production, pretreatment of homoplantaginin could effectively reverse the effects of PA. Additionally, homoplantaginin significantly modulated the Ser/Thr phosphorylation of IRS-1, improved phosphorylation of Akt and endothelial nitric oxide synthase (eNOS), and increased NO production in the presence of insulin. Taken together, our results demonstrated that homoplantaginin ameliorates endothelial insulin resistance by inhibiting inflammation and modulating cell signalling via the IKKβ/IRS-1/pAkt/peNOS pathway, suggesting it may be used for the prevention and treatment of endothelial dysfunction associated with insulin resistance.
  • Masashi Tsukamoto, Tomoyuki Okuda, Hirokazu Okamoto, Yuriko Higuchi, S ...
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1178-1181
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    The development of efficient and selective therapeutic gene delivery methods is a potential medical treatment for intractable diseases. Dry powder inhalers (DPIs) can efficiently deliver drugs locally to the lung. Many reports discuss preparation methods for DPIs. Spray-freeze drying is a method by which highly porous particulates can be prepared. However, altered physical properties after preparation may result in changes in gene expression. In this study, bovine serum albumin (BSA) was added as a lyoprotectant, and 1,2-dioleoyl-3-trimethylammonium propane/cholesterol liposomes/pCMV-Luc DPIs (lipoplex DPIs) were prepared by spray-freeze drying. The mean particle sizes of the lipoplex DPIs prepared without BSA increased by approximately 6.7-fold compared with that of the lipoplexes solution. In contrast, the mean particle sizes of the lipoplex/BSA DPIs increased only slightly. Gene expression was evaluated after the intratracheal administration of the lipoplexes solution, with maximum gene expression observed at 12 h after the administration. In contrast, maximum gene expression of the lipoplex/BSA DPIs occurred at 6 h after administration. The gene expression associated with the lipoplex DPIs was significantly lower compared with that of the lipoplex/BSA DPIs at 6 (p<0.01), 12 (p<0.01), and 24 h (p<0.05). These variances may be due to the difference in mean particle size between the DPI formulations. The results suggest that BSA is a useful lyoprotectant for dry powder formulation preparations of DNA using the spray-freeze drying method, given that the preparation results in minimal variation of physical properties and gene expression.
  • Yoshiaki Umemoto, Shigeru Kawakami, Yuki Otani, Yuriko Higuchi, Fumiyo ...
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1182-1186
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    PhiC31 integrase has the potential to achieve long-term transgene expression because of site-specific and unidirectional recombination. In this study, plasmid DNA (pDNA) encoding Gaussia luciferase (Gluc) cDNA was constructed and the optimal condition for long-term gene expression using phiC31 integrase in mouse liver after hydrodynamic injection was investigated. Gluc is secreted and thus allows quantification of its expression level in blood and easy determination of the expression time-course. Mice were co-transfected with 25 µg donor pDNA (pORF-Gluc/attB) and different amounts of helper pDNA (pCMV-int; from 1 to 50 µg). Serum Gluc expression was evaluated during 120 d. The most highly sustained gene expression was obtained when 10 µg of helper pDNA was co-transfected in ICR and C57BL/6 mice. However, the Gluc expression in C57BL/6 mice was slightly lower and less durable than that in ICR mice. Little hepatic damage caused by phiC31 integrase was observed during 120 d in ICR and C57BL/6 mice.
  • Nobuyasu Yamaguchi, Akiko Sakotani, Tomoaki Ichijo, Takehiko Kenzaka, ...
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1187-1190
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Asian dust (called ‘Kosa’ in Japan) is comprised of a large number of soil particles originating from the arid regions and deserts of China and Mongolia and dispersed long-range to Japan. A major public concern about Asian dust is its impact on human health. We collected Asian dust particles over the Japan Sea at an altitude of 900 m to directly estimate their effects on health. We examined the properties of the collected particles on wet surfaces. Through size distribution measurements and scanning electron microscopy with energy dispersive X-ray (SEM-EDX) analysis, we demonstrated that small dust particles (less than 1 µm) form aggregations with water-soluble salts such as calcium and sodium and they are transported to Japan as aggregates. These aggregates probably break down into small particles on nasal mucous membranes and may cause adverse respiratory health effects.
  • Waleed Osman, Satoshi Endo, Kentaro Oh-hashi, Yoshiaki Kitamura, Yukio ...
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1191-1196
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    Asymmetrical diadenosine 5′,5″-P1,P4-tetraphosphate hydrolase (EC 3.6.1.17) from human malaria parasite Plasmodium falciparum was expressed in Escherichia coli, purified to homogeneity, and characterized for the first time as a biological target for chemotherapeutic agents against malaria. Plasmodium falciparum Ap4A (PfAp4A) hydrolase not only catalyzes diadenosine 5′,5″-P1,P4-tetraphosphate (Ap4A) to ATP and AMP, but also diadenosine 5′,5″-P1,P5-pentaphosphate (Ap5A) to ATP and ADP. Marked enzyme heat stability corresponding to the highest level of activity was observed at 60°C. The recombinant enzyme showed maximal activity in the presence of 5 mM Mg2+ ions. Kinetic analysis revealed the values of Km and Kcat as 0.6 µM and 2.5 min−1, respectively. Comparative protein modeling indicated an additional space in the substrate binding site of the parasitic enzyme compared with that of humans. Mutagenic analysis of the amino acid residue (Pro133) forming the additional space revealed a 5-fold increase in the wild-type Km value when replaced by a smaller (Ala) residue. Furthermore, catalytic activity was markedly affected by introducing a larger residue (Phe), thus creating the potential to develop a specific inhibitor of PfAp4A hydrolase.
  • Pongpun Siripong, Jantana Yahuafai, Suratsawadee Piyaviriyakul, Kwanja ...
    原稿種別: Note
    2012 年 35 巻 7 号 p. 1197-1200
    発行日: 2012/07/01
    公開日: 2012/07/01
    ジャーナル フリー
    We previously observed that rhinacanthins, which are the main naphthoquinone esters isolated from the roots of a Thai medicinal plant, Rhinacanthus nasutus KURZ. (family Acanthaceae), suppress the growth of Meth-A sarcoma in the tumor-bearing mice and that rhinacanthin-N has the strongest antitumor activity among these naphthoquinone esters tested. In the present study, we investigated the effect of rhinacanthin-N on pulmonary metastasis induced by B16F10 melanoma cells in mice. C57BL/6 male mice were injected intravenously with B16F10 melanoma cells, and liposomal rhinacanthin-N was administered intraperitoneally from day 1 to 7 after tumor implantation. Liposomes were used to formulate an injectable form of the hydrophobic agent. Treatment of the mice with 5 or 10 mg/kg/d of liposomal rhinacanthin-N significantly inhibited the pulmonary metastatic colonization of the melanoma cells. Based on these data, our findings demonstrate that rhinacanthin-N possesses antimetastatic efficacy, which may make it a lead compound for the development of a new anticancer drug for use in cancer chemotherapy.
feedback
Top