Biological and Pharmaceutical Bulletin
Online ISSN : 1347-5215
Print ISSN : 0918-6158
ISSN-L : 0918-6158
Volume 35 , Issue 8
Showing 1-30 articles out of 30 articles from the selected issue
Reviews
  • Takashi Hirooka, Toshiyuki Kaji
    Type: Review
    2012 Volume 35 Issue 8 Pages 1201-1205
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Methylmercury is an environmental neurotoxin that induces severe neurological damage in the brain of humans and animals. The main pathological characteristic of methylmercury neurotoxicity is the location of the damage; lesions are localized around the deep sulci and fissures in the cerebral cortex, such as the calcarine fissure, and the granule cell layer of the cerebellum. Since the localization of the damage is suggested to be a result of secondary damage occurring due to edematous change in the white cortex, the toxicity of methylmercury to cells that compose the microvessels―endothelial cells and pericytes―may be important for understanding the neurotoxicity of methylmercury. We investigated the toxicity of methylmercury to human brain microvascular endothelial cells and pericytes using a cell culture system. It was revealed that the toxicity of methylmercury to microvascular cells depends on the cell type and density. It is suggested that vascular tissue is one of the targets of methylmercury toxicity and that this may contribute to the progression of edematous change in the brain. Methylmercury may also be involved in the progression of cardiovascular diseases.
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  • Hiroshi Fukasawa, Madoka Nakagomi, Naoko Yamagata, Hiroshi Katsuki, Ko ...
    Type: Review
    2012 Volume 35 Issue 8 Pages 1206-1212
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Tamibarotene (Am80), a synthetic retinoid approved in Japan for treatment of acute promyelocytic leukemia (APL), is a retinoic acid receptor (RAR) agonist with high specificity for RARα and RARβ over RARγ. Temporarily and spatially specific expression of RARs suggests their pivotal roles in the adult brain. Am80 is considered to be a promising candidate drug for treatment of Alzheimer’s disease (AD) because of its transcriptional controls of multiple target genes involved in etiology and pathology of AD. In APP23 AD model mice, administration of Am80 decreased the deposition of insoluble amyloid-β(42). In senescence-accelerated mice (SAMP8), Am80 ameliorated the decrease of cortical acetylcholine, as well as reducing anxiety in behavioral tests and improving the sleep deficit. Am80 also effected a significant improvement of memory in the rat scopolamine-induced memory deficit model. Like other retinoids, Am80 also has an immunomodulatory effect and reduces secretion of proinflammatory cytokines and chemokines by astrocytes and microglia surrounding amyloid-β plaques. In a rat experimental autoimmune encephalomyelitis model, Am80 reduced inflammatory cytokines and showed significant efficacy. Retinoids also promote differentiation of neural stem cells, and Am80 improved the recovery of spinal cord-injured rats. Am80 may also improve vascular factors involved in onset and/or progression of AD. Am80 has been in clinical use for treatment of APL in Japan since 2005, and has been reported to have fewer side effects than other retinoids. We have recently started a clinical study to evaluate the efficacy and safety of Am80 for the treatment of Alzheimer’s disease.
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Communication to the Editor
  • Shinichirou Nakagawa, Yusuke Matsuoka, Hideaki Ichihara, Hitoji Yoshid ...
    Type: Communication to the Editor
    2012 Volume 35 Issue 8 Pages 1213-1215
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    It is well known that trastuzumab (TTZ) is molecular target drug for breast cancer overexpressing human epidermal growth factor receptor 2 (HER2). Novel immunotherapy by human peripheral blood mononuclear cells (PBMCs) activated with TTZ were examined. Proliferation of lymphocytes after adding of TTZ was obtained. Furthermore, lymphocytes activated with TTZ inhibited growth of breast cancer cells in vitro. It is noteworthy that remarkably high cellular cytotoxicity in lymphocytes activated with TTZ compared with that of CD3- and lymphokine (interleukin (IL)2)-activated killer (CD3-LAK) cells commonly used in immunotherapy were revealed.
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Regular Articles
  • Jingtao Wu, Guiwen Yang, Wenxing Zhu, Wujun Wen, Fumiao Zhang, Jinduo ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1216-1221
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    This study examined the effects of platycodin D (PD), a triterpene saponin from the the root of Platycodon grandiflorum A.DC on human umbilical vein endothelial cells (HUVECs) in vitro, which were pre-treated with PD (0.01, 0.15, 0.25 mg/mL), respectively, and treated with 50 mg/L oxidized low-density lipoprotein (oxLDL). The levels of nitric oxide (NO) and malonaldehyde (MAD) in the culture medium, vascular cell adhesion molecule-1 (VCAM-1) and intercellular cell adhesion molecule-1 (ICAM-1) mRNA expression in endothelium cells and the adhesion of monocytes to endothelial cells were measured. The results showed that PD increased NO concentration and decreased MDA level induced by oxLDL in the medium of endothelial cells. Moreover, PD significantly inhibited the oxLDL-induced increase in monocyte adhesion to endothelial cells as well as decreasing mRNA expression levels of VCAM-1 and ICAM-1 on these cells. Based on these results, it is suggested that PD is a promising anti-atherosclerotic activity, which is at least in part the result of its increasing NO concentration, reducing the oxLDL-induced cell adhesion molecule expression in endothelial cells and the endothelial adhesion to monocytes.
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  • Min-Soo Kim, Wan-Gi Kim, Hwan-Suck Chung, Byoung-Woo Park, Kyoo-Seok A ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1222-1229
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Atopic dermatitis (AD) is characterized as a multi-factorial inflammatory skin disease that has been increasing worldwide. Previously, we demonstrated that FPG, which is Platycodon grandiflorum (PG) fermented by Lactobacillus plantarum (LP), increases the level of interferon (IFN)-gamma in mouse splenocytes in vitro. In this study, we investigated the effects of FPG in an animal model of AD, with a particular emphasis on its effects on T helper (Th)1 and Th2 immune responses. To assess the potential use of FPG for the inhibition of AD, we established a model of AD-like skin lesions in NC/Nga mice. Immunoglobulin isotypes (Igs) and Th1/Th2 cytokines in the sera and spleens of AD-like mice were examined. In addition, histological examination was also performed. AD symptoms in skin lesions improved following oral administration of FPG. IgE secretion was significantly down-regulated, and this was accompanied by decreased levels of interleukin (IL)-4 and IgG1 and increased serum levels of IL-12p40 and IgG2a in FPG-treated animals. In splenocytes, the production of the Th1 cytokines IL-12p40 and IFN-gamma was up-regulated, while the levels of the Th2 cytokines IL-4 and 5 were down-regulated by FPG treatment. These results suggest that FPG inhibits the development of AD-like skin lesions in NC/Nga mice by suppressing the Th2 cell response and increasing the Th1 cell responses. Our results indicate that FPG is safe and effective for the prevention of AD-like skin lesions.
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  • Mei-ling Yu, Cui-ling Zhang, Dong-dong Yuan, Xu-hui Tong, Liang Tao
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1230-1237
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Panax Notoginseng Saponins (PNS) have been well known to have anti-tumor activity and enhance cytotoxicity of some cancer chemotherapy agents, but the mechanisms underlying these effects are still unknown. This study investigates the effect of PNS on cytotoxicity of cisplatin and the relationship between this effect and the modulation of gap junctions (GJ) function by PNS in a transfected cell line. The cytotoxicity of cisplatin (0.25–1 µg/mL) was increased in the presence of GJ. Inhibition of gap junction by either GJ blocker or interception of Connexin (Cx) expression decreased the cytotoxicity of cisplatin. Increasing GJ function enhanced cytotoxicity of cisplatin, only in the cells with functional GJ. PNS (50–200 µg/mL) significantly enhanced cisplatin cytotoxicity, but this effect required functional gap junctions between the cells. Exposure of the cells to PNS (50–200 µg/mL) for 4 h leads to a significant enhance in dye coupling of GJ in a dose-dependent manner. These results suggest that PNS increases the cytotoxicity of cisplatin through enhancement of GJ activity.
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  • Seiichi Yoshida, Hirohisa Takano, Masataka Nishikawa, He Miao, Takamic ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1238-1243
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Urban particulate matter (UPM) has been shown to have an aggravating effect on Th2-associated immune systems in adult mice. However, the effects of fetal exposure to UPM on immune response in offspring have not been elucidated. In the present study, we administered UPM (200 µg/animal) by intratracheal injection to pregnant dams on days 7 and 14 of gestation. Subsequently, 9- and 24-week-old male offspring were intratracheally injected with ovalbumin (OVA) (four times at 2-week intervals) to create a mouse model of bronchial asthma. We then evaluated the progression of allergic manifestations in the offspring through histological findings, the number of inflammatory cells in bronchoalveolar lavage fluid (BALF), and protein concentration of cytokines and chemokines in BALF 5, 10, 15, and 30 weeks after birth. Histological examination showed that fetal exposure to UPM alone caused slight eosinophil and lymphocyte infiltration in the submucosa of the airway and bronchial epithelium and significant increases in the number of macrophages. Moreover, postnatal intratracheal administration of OVA to offspring exposed to UPM in utero caused significant increases in the numbers of macrophages, eosinophils, and lymphocytes and in the concentrations of their relevant cytokines and chemokines, showing that fetal exposure to UPM aggravated the chemically sensitized immune system of male offspring.
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  • Yoritaka Onzawa, Yasuhiro Kimura, Kengo Uzuhashi, Megumi Shirasuna, Ta ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1244-1248
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    It has been well known that 3-O-methyldopa (3-OMD) is a metabolite of L-3,4-dihydroxyphenylalanine (L-DOPA) formed by catechol O-methyltransferase (COMT), and 3-OMD blood level often reaches higher than physiological level in Parkinson’s disease (PD) patients receiving long term L-DOPA therapy. However, the physiological role of 3-OMD has not been well understood. Therefore, in order to clarify the effects of 3-OMD on physiological function, we examined the behavioral alteration in rats based on locomotor activity, and measured dopamine (DA) and its metabolites levels in rats at the same time after 3-OMD subchronic administration. The study results showed that repeated administrations of 3-OMD increased its blood and the striatum tissue levels in those rats, and decreased locomotor activity in a dose dependent manner. Although 3-OMD subchronic administration showed no significant change in DA level in the striatum, DA metabolite levels, such as 3,4-dihydroxyphenylacetic acid (DOPAC), 3-methoxytyramine (3-MT), and homovanillic acid (HVA) were significantly decreased. After 3-OMD washout period (7 d), locomotor activity and DA turnover in those rats returned to normal levels. Furthermore, locomotor activity and DA turnover decreased by 3-OMD administration were recovered to normal level by acute L-DOPA administration. These results suggested that 3-OMD affect to locomotor activity via DA neuron system. In conclusion, 3-OMD itself may have a disadvantage in PD patients receiving L-DOPA therapy.
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  • Hong Xin, Ming Gu, Wen Wei Wang, Shi Ying Huang, Fang Ping Li, Hui Cai ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1249-1256
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Leonurine (Leo) is a special alkaloid principle of Herba leonuri that has recently been suggested to improve cardiovascular functions. To date, there is no direct ionic evidence of Leo on regulating calcium channels in the heart. In the present study, we examined the effects of Leo on action potentials and membrane currents recorded from isolated rat ventricular myocytes with the whole-cell patch clamp technique. Leo 100 µM shortened the action potential duration in a dose-dependent manner. Leo up to 200 µM had no significant effect on the Na+ current (INa) and K+ current (IK). However, Leo depressed the L-type Ca2+ current (ICa,L). In the presence of 20 and 100 µM Leo, the current density was decreased and the voltage at half maximal inactivation V0.5 shift to more negative potential. The recovery time constant was also delayed. In addition, the transcription and protein expression levels of L-type calcium channel (Cav1.2) in primary cultured neonatal myocytes from Sprague-Dawley rats were reduced by Leo treatment in a dose-dependent fashion as assessed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assays. We conclude that Leo inhibits L-type calcium channels in cardiomyocytes.
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  • Hongbo Chai, Masaharu Hazawa, Yoichiro Hosokawa, Jun Igarashi, Hiroaki ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1257-1263
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    The cytotoxicity of novel acridine-based N-acyl-homoserine lactone (AHL) analogs was investigated on the human oral squamous carcinoma cell line SAS. One analog induced G2/M phase arrest at 5.3–10.6 µM and induced polyploidy at a higher dose (21.2 µM). Importantly, treatment of SAS cells with a combination of the AHL analog and the Jun N-terminal kinase (JNK) inhibitor, SP600125, prevented mitosis and induced polyploidy. The AHL analog synergized with X-irradiation to inhibit clonogenic survival of SAS cells; however, its radiosensitizing effects were relative to not X-irradiation-induced apoptosis but mitotic failure following enhanced expression of Aurora A and B. These results suggest that the active AHL analog showed growth-suppressive and radiosensitizing effects, which involve polyploidy followed by G2/M accumulation and atypical cell death in the SAS cell line.
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  • Miyuki Shirosaki, Yoko Goto, Saori Hirooka, Hideki Masuda, Tomoyuki Ko ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1264-1268
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Peach leaf extract has anti-hyperglycemic effects on the postprandial blood glucose level in glucose-loaded mice. In our previous study, the mechanism of action was considered to be the inhibition of glucose absorption in the small intestine. To elucidate the active principle in peach leaf, purification of the active compound and a structure determination were performed. With the use of bioassay-guided fractionation using glucose-loaded mice, the acetylated kaempferol glycoside multiflorin A (MFA), a potent inhibitor of glucose absorption from the intestine, was isolated from the MeOH extract of leaf of the edible peach Prunus persica. The structure was identified by HPLC using thiazolizine derivatives and by an analysis of its spectral data. The inhibitory effect of MFA against glucose absorption was demonstrated in the dose dependent manner in mice. However, as the deacetylated analog of MFA, multiflorin B did not show the activity at the in vivo, the activity of MFA was suggested to depend on the acetyl group on the sugar moiety. This is the first report of anti-hyperglycemic activity of MFA in peach leaf extract. MFA may be useful in functional foods or medicines for preventing the postprandial absorption of glucose in hyperglycemia.
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  • Kyohei Takahashi, Ryosuke Tatsunami, Keisuke Sato, Yoshiko Tampo
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1269-1274
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    4-Hydroxy-2-nonenal (HNE), an aldehyde produced by lipid peroxidation, induces cytotoxicity and oxidative stress. Glutathione (GSH) protects against the cytotoxicity of HNE. However, the protective mechanism of GSH has not been fully examined. We examined the protective role played by the relationship between GSH and multidrug resistance associated protein 1 (MRP1) against the HNE-induced oxidative stress in bovine aortic endothelial cells (BAECs). HNE induced the loss of viability of BAECs. Exogenous GSH, which is membrane-impermeable, prevented the loss of viability induced by HNE by inhibiting HNE uptake in BAECs, probably due to the formation of the HNE-SG complex in the extracellular space. We demonstrated that HNE induced the expression of MRP1 protein, which can transport the HNE-SG complex. The induction of MRP1 protein expression by HNE disappeared in BAECs pretreated with L-buthionine sulfoximine, a GSH-depleting agent. This result suggests that HNE, together with intracellular GSH, contributes to the regulation of MRP1 protein expression. Moreover, we found that MK571, an MRP1 inhibitor, promoted the HNE-induced oxidative stress and cell death. Taken together, these findings suggest that MRP1, together with GSH, plays a protective role against the HNE-induced oxidative stress in BAECs.
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  • Daisuke Kiyonaga, Noriko Tagawa, Yuko Yamaguchi, Midori Wakabayashi, T ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1275-1280
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Excess glucocorticoids promote visceral obesity and insulin resistance. The main regulator of intracellular glucocorticoid levels are 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts inactive glucocorticoid into bioactive glucocorticoid such as cortisol in humans and corticosterone in rodents; therefore, the inhibition of 11β-HSD1 has considerable therapeutic potential for metabolic diseases including obesity and diabetes. Benzofuran is a key structure in many biologically active compounds such as benzbromarone, malibatol A and (+)-liphagal. The aim of this study was to investigate the inhibitory effect of benzofuran derivatives on 11β-HSD1 in mesenteric adipose tissue from rodents. 11β-HSD1 activity was determined by incubation of rat mesenteric adipose tissue microsomes in the presence of reduced nicotinamide adenine dinucleotide phosphate (NADPH) with and without benzofuran derivatives (Compounds 114). The corticosterone produced was measured by HPLC. More than 40% of 11β-HSD1 inhibition was observed in Compounds 1, 5, 7 and 8. Moreover, Compounds 7 and 8 inhibited the 11β-HSD1 activity in adipose microsomes dose- and time-dependently, as well as in 3T3-L1 adipocytes. Compounds 7 and 8 did not inhibit 11β-HSD type 2 (11β-HSD2), whereas Compounds 1 and 5 inhibited 11β-HSD2 by 18.7% and 56.3%, respectively. Further, a kinetic study revealed that Compounds 7 and 8 acted as non-competitive inhibitors of 11β-HSD1. Ki (nmol/h/mg protein) values of Compounds 7 and 8 were 17.5 and 24.0, respectively, with IC50M) of 10.2 and 25.6, respectively. These data indicate that Compounds 7 and 8 are convincing candidates for seed compounds as specific inhibitors of 11β-HSD1 and have the potential to be developed as anti-obesity drugs.
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  • Kwang-Seok Oh, Byung Koo Oh, Cheon Ho Park, Jihye Mun, Suk Hyun Won, B ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1281-1286
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Baicalein is a flavonoid (5,6,7-trihydroxy-2-phenyl-4H-1-benzopyran4-one) and an active principle in Scutellaria baicalensis. The present study was performed to investigate the mechanisms underlying the anti-fibrotic effects of baicalein with a focus on Rho kinase (ROCK) inhibition. The effect of baicalein on ROCK activity was analyzed using an immobilized metal affinity for phosphochemicals (IMAP)-based time-resolved fluorescence resonance energy transfer (TR-FRET) assay. The underlying mechanisms of baicalein were examined using angiotensin II-stimulated H9c2 cells. Rho kinase (ROCK1 and ROCK2) studies using IMAP-TR-FRET showed that baicalein possesses potent ROCK inhibitory activity with IC50 values of 6.55 and 2.82 µM, respectively. Pretreatment with baicalein (for 2 h) concentration-dependently decreased the angiotensin II-induced phosphorylation of myosin phosphatase (MYPT) and myosin light chain (MLC). Furthermore, baicalein also concentration-dependently suppressed actin stress fiber formation in angiotensin II-stimulated H9c2 cells. These results suggest that baicalein potently inhibits ROCK and that by so doing it modulates actin stress fiber formation. These anti-fibrotic effects of baicalein explain, at least in part, its pharmacology and mode of action.
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  • Hyo Geun Kim, Mi Sun Ju, Sang Keun Ha, Hyangsook Lee, Hyejung Lee, Sun ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1287-1294
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Acacetin (5,7-dihydroxy-4′-methoxyflavone), a constituent of flavone naturally present in plants, has anti-cancer and anti-inflammatory activities. Neuroinflammation is thought to be one of the major pathological mechanisms responsible for Parkinson’s disease (PD), and has been a primary target in the development of treatment for PD. In the present study, we evaluated the neuroprotective effect of acacetin in PD induced by 1-methyl-4-phenylpyridine (MPP+)/or 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and examined the related pathways in vitro and in vivo. In primary mesencephalic culture, acacetin protected dopaminergic (DA) cells and inhibited production of inflammatory factors such as nitric oxide, prostaglandin E2, and tumor necrosis factor-α against MPP+-induced toxicity in a dose-dependent manner. Then, we confirmed the effect of acacetin (10 mg/kg/d for 3 d, per os (p.o.)) in a mouse model of PD induced by MPTP (30 mg/kg/d for 5 d, intraperitoneally (i.p.)). In the behavioral test (pole test), the acacetin-treated mice showed decreased time of turning and locomotor activity, which were longer in MPTP-only treated mice. In addition, the acacetin-treated group inhibited degeneration of DA neurons and depletion of dopamine level induced by MPTP toxicity in the substantia nigra and striatum of the brain. Moreover, the acacetin-treated group inhibited microglia activation, accompanied by production of inducible nitric oxide synthases and cyclooxygenase-2. These results suggest that acacetin can protect DA neurons against the neurotoxicity involved in PD via its anti-inflammatory action.
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  • Pei Yu, Longfei Xia, Jing Zhao, Gaoxiao Zhang, Zaijun Zhang, Ming Lang ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1295-1299
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    We have synthesized new 10-hydroxycamptothecin (HCPT) analogs and evaluated their anticancer activity in cell culture and in experimental animal tumor model. Although the new analogs were less potent against L1210 leukemia cells in vitro, some of them were more efficacious against L1210 leukemia in vivo compared to the parent HCPT.
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  • Aiko Nishio, Naoki Kitamura, Saki Tanaka, Shunsuke Miyazaki, Tsuyoshi ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1300-1305
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Gymnopilin is one of the substances produced by the hallucinogenic mushroom, Gymnopilus junonius. In this study, we examined effects of gymnopilins purified from wild fruiting bodies of G. junonius on contractile activity of aorta preparations and blood pressure in rats. Gymnopilins at lower concentrations than 5 mg/mL did not evoke contraction of helical strips of the thoracic aorta. In contrast, gymnopilins (5 mg/mL) applied to the aorta strips pre-contracted by norepinephrine (100 nM) caused relaxation. This relaxing action did not depend on the activity of the endothelium cells. The relaxing effect of 5-mg/mL gymnopilins was observed in aorta strips contracted by angiotensin II (10 nM) and the high K+ solution (60 mM). Moreover, the adenylyl cyclase inhibitor, SQ-22536, significantly inhibited the relaxing effect of gymnopilins at 1 mg/mL on the norepinephrine-contracted strips. These results suggested that gymnopilins acted directly on smooth muscle cells of the aorta and activated the cAMP-dependent cascade to cause the vasodilation. Paradoxically, gymnopilins injection into the jugular vein transiently increased blood pressure without affecting the heart rate. This result suggests that gymnopilins increase cardiac output and/or tension of the artery through the excitation of the vasomotor nerve that overcame the direct relaxing effect on the vascular smooth muscle.
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  • Tomoya Araki, Yusuke Kono, Ken-ichi Ogawara, Takaichi Watanabe, Tsutom ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1306-1313
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    To develop potent paclitaxel (PTX) formulations for cancer chemotherapy, we formulated PTX into polymeric nanoparticles composed of polyethylene glycol (PEG) and polylactic acid (PLA) block copolymer (PN-PTX). First, the physicochemical properties of PN-PTX prepared were assessed; the mean particle size was around 80 nm and the zeta potential was found to be almost neutral. Next, the in vitro PTX release property was assessed by a dialysis method. Although rapid release of PTX was observed just after dosing, around 70% of PTX was stably incorporated in polymeric nanoparticles for a long time in the presence of serum. Then, the in vivo pharmacokinetics of PN-PTX after intravenous administration was investigated in Colon-26 (C26) tumor-bearing mice. Both polymeric nanoparticles and PTX incorporated exhibited a long blood circulating property, leading to enhanced permeability and retention (EPR) effect-driven, time-dependent tumor disposition of PTX. Tumor distribution increased gradually for 24 h, and tissue uptake clearance of polymeric nanoparticles in the liver and spleen was lower than that of PEG liposomes. Since these results indicated that the in vivo disposition characteristics of PN-PTX were very favorable, we then evaluated the anti-tumor effect of PN-PTX in C26 tumor-bearing mice. However, PN-PTX did not exhibit any significant anti-tumor effect, presumably due to the poor PTX release from polymeric nanoparticles. From these results, it is considered that the favorable pharmacokinetic properties of nanoparticles and the drug incorporated do not always lead to its potent in vivo pharmacological activity, suggesting the importance of PTX release properties within tumor tissues.
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  • Takumi Tsuji, Yuya Yoshida, Rumi Iwatsuki, Mariko Inoue, Tetsuro Fujit ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1314-1319
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    The therapeutic efficacy of the novel immunomodulator FTY720 (Fingolimod), alone and in combination with betamethasone ointment, was examined in the NC/Nga mouse model of spontaneous steroid-resistant dermatitis. Male NC/Nga mice which had developed severe dermatitis were divided into six groups: 1) a biweekly betamethasone group (betamethasone ointment, twice a week), 2) a daily betamethasone group (betamethasone ointment, six times a week), 3) an FTY720 group (FTY720, orally, three times a week), 4) a biweekly combination group (oral FTY720 plus betamethasone ointment, twice a week), 5) a daily combination group (oral FTY720 plus betamethasone ointment, six times a week) and 6) a placebo group (vehicle alone). The therapeutic efficacy was evaluated in terms of severity of dermatitis, epidermal hypertrophy, accumulation and degranulation of mast cells and infiltrated CD3+ T cells into the dermis after 4 weeks of treatment. Biweekly and daily betamethasone treatments had little effect, confirming that the dermatitis was steroid-resistant. In the FTY720 and biweekly combination groups, the dermatitis showed no marked improvement. In the daily combination group, the dermatitis was significantly (p<0.05, Mann–Whitney U-test) improved as compared with the FTY720 group, biweekly and daily betamethasone groups and placebo group. Further, epidermal hypertrophy and accumulation of mast cells were suppressed. Therefore, combination therapy with FTY720 and daily betamethasone ointment is a promising candidate for treatment of steroid-resistant atopic dermatitis.
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  • Hidesuke Fukazawa, Tetsuro Suzuki, Takaji Wakita, Yuko Murakami
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1320-1327
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    We describe a cell-based, microplate colorimetric screen for anti-hepatitis C virus (HCV) drugs that exploits the HCV-JFH1 viral culture system. Antiviral activity was assessed by measuring protection against the HCV-JFH1-induced cytopathic effect (CPE) in Huh7.5.1 cells using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) viability assay. The use of serum-free medium substantially sensitized Huh7.5.1 cells to HCV-induced CPE, causing sufficient cell death to perform colorimetric assays for anti-HCV activity in 96-well plates. As a proof of concept, we carried out a pilot screen of an inhibitor library and identified cyclosporin A and tamoxifen, two compounds with reported anti-HCV activity. Using the assay, we discovered the anti-HCV properties of the plant flavonoids epigallocatechin gallate (EGCG) and 7,8-benzoflavone (α-naphthoflavone). Other gallate-type catechins and flavones also displayed anti-HCV activity, but 5,6-benzoflavone (β-naphthoflavone), flavanone, and non-gallate catechins were inactive. EGCG apparently acted mainly on HCV entry, although it may also block other steps. In contrast, 7,8-benzoflavone was presumed to inhibit later stages of the HCV life cycle. This assay is simple, reliable and cost-effective; does not require any specially engineered cell lines or viruses; and should be useful in the identification of compounds with anti-HCV activity.
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  • Xican Li, Gang Wei, Xiaozhen Wang, Dong-Hui Liu, Ru-Dong Deng, Hui Li, ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1328-1335
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Molecules that enhance chondrogenic differentiation in mesenchymal stem cells (MSCs) were identified and isolated using an in vitro Gli reporter gene assay in MSCs incorporating a Sonic Hedgehog (Shh) target. Atractylenolide III, which promoted Gli1-mediated transcriptional activity, was isolated from an ethyl acetate extract of the Rhizoma, Atractylodis macrocephalae. After dehydration, atractylenolide III was transformed to atractylenolide I. Both atractylenolides were confirmed by MS, UV, IR, 1H- and 13C-NMR spectra. Atractylenolide III (which contains –OH at the 8-position) and atractylenolide I (which lacks –OH at the 8-position) were found to effectively promote the activity of the Gli promoter. While the hydroxyl group of atractylenolide III was not essential for the effect of atractylenolide, its effect was dependent on Shh signaling. Phenotypic cellular analysis indicated that atractylenolides induced MSCs to differentiate into chondrocytes, as shown by increased expression of specific chondrogenic markers including collagen II, aggrecan and the cartilage related transcription factor, Sox9. Atractylenolides significantly increased the expression of Shh and its target gene Gli-1, indicating that Shh signaling was activated by atractylenolides. Moreover, inhibition of Shh signaling reduced the effect of atractylenolides on the chondrogenic phenotype. The discovery that atractylenolides induce chondrocytes from MSCs is promising for bony disease therapy.
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  • Taro Shimizu, Masako Ichihara, Yasuo Yoshioka, Tatsuhiro Ishida, Shins ...
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1336-1342
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    A single intravenous administration of polyethylene glycol-coated (PEGylated) bovine serum albumin (BSA) and ovalbumin (OVA) elicited an anti-PEG immunoglobulin M (IgM) response, similar to that from PEGylated liposomes, although the administration did not elicit specific neutralizing antibodies to BSA and OVA. A cross-reactivity was observed between anti-PEG IgMs elicited by PEG-BSA and PEGylated liposomes. The anti-PEG IgM level induced by PEGylated proteins (BSA and OVA) reached the maximum at day 5 following intravenous injection. This production pattern was consistent with that induced by PEGylated liposomes. Splenectomy suppressed the anti-PEG IgM response against PEG-BSA and PEGylated liposomes. These observations relating PEG-BSA and PEGylated liposomes indicate that PEGylated proteins might promote the immune responses against PEG with a mechanism similar to that of PEGylated liposomes. In addition, a single intravenous administration of PEGylated adenovirus (PEG-Ad) also elicited an anti-PEG IgM response in a PEG-modification ratio dependent manner. To the best of our knowledge, this is the first report showing that an intravenous administration can elicit an anti-PEG IgM response against PEGylated substances. It appears that anti-PEG IgMs can be produced by the systemic administration of a PEGylated substance and may limit the efficacy of PEGylated substances such as proteins, Ad vector and nanoparticles, due to a cross-reactivity seen in some patients. The immunogenicity of PEGylated substances is usually tested against those very substances, rather than against covalently attached PEG. Our study suggests that the PEG immunogenicity of PEGylated therapeutic agents and particles merits further investigation.
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  • Daisuke Horita, Hiroaki Todo, Kenji Sugibayashi
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1343-1348
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    It has been demonstrated that ethanol (EtOH) can enhance skin permeation of drugs when simultaneously applied with drugs. However, only a few studies have reported on the pretreatment effect of EtOH on skin permeations. In this study, the pretreatment effects of EtOH on skin permeation of drugs were investigated by measuring changes in skin permeation and electrical skin resistance. Permeabilities of deuterium oxide (D2O), isosorbide mononitrate (ISMN), isosorbide dinitrate (ISDN), calcein sodium (CA-Na), and fluorescein isothiocyanate-dextran 4 kDa (FD-4, 3.3–4.4 kDa) were evaluated through Yucatan micropig skin pretreated with different concentrations of EtOH solution. From the results, almost constant skin permeabilities of D2O and ISDN were observed independent of EtOH concentration. Skin permeabilities of ISMN, CA, and FD-4 increased with low concentrations of EtOH, but decreased with high concentrations of EtOH. At 99.5% EtOH pretreatment, skin permeabilities of hydrophilic compounds (ISMN, CA, and FD-4) decreased to non-detectable levels. In addition, low molecular ion transports were almost constant at any EtOH concentration. Since molecular (ion) sizes of ISMN, CA, and FD-4 are larger than Na+, Cl, and D2O, permeation pathway sizes for hydrophilic compounds in the skin barrier may be remarkably decreased by pretreatment with high concentrations of EtOH. However, the permeability coefficient of ISDN was not influenced by any EtOH concentration, since ISDN is a lipophilic, low-molecular compound that permeated through the lipophilic stratum corneum pathway. The present results show useful information for repeatedly and topically applied formulations containing EtOH, and also contribute to the effective use of alcohol formulations.
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  • Koji Mizukoshi, Katsuo Matsumoto, Ryoji Hirose, Tetsuro Fujita
    Type: Regular Article
    2012 Volume 35 Issue 8 Pages 1349-1353
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    We investigated the structure–activity relationship between various ISP-I (myriocin, thermozymocidin) analogous which has sphingosine-like structure and serine palmitoyltransferase (SPT) in Chinese hamster ovary (CHO) cells utilizing sphingolipid production as a marker. Our data suggest that the double bond and/or ketone group within the alkyl chain as well as the alkyl chain are necessary for ISP-I to inhibit SPT. In addition, a serine structure is necessary for SPT inhibitory activity, which confirms previous findings.
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Notes
  • Marie Takekawa, Tadahide Furuno, Naohide Hirashima, Mamoru Nakanishi
    Type: Note
    2012 Volume 35 Issue 8 Pages 1354-1360
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    The ability of mitochondria to take up Ca2+ has important functional implications for modulation of cellular Ca2+ signaling. Mitochondrial Ca2+ uptake is stimulated by an increase in cytosolic Ca2+ concentration ([Ca2+]c). Here, we found that the increase in mitochondrial Ca2+ concentration ([Ca2+]m) occurs in two steps in a single antigen-activated mast cell in the presence of extracellular Ca2+ (1.0 mM). The two-step elevation of [Ca2+]m was also observed after adding thapsigargin, an inhibitor of sarcoplasmic/endoplasmic reticulum Ca2+-ATPase. The proportion of mitochondria showing the two-step Ca2+ elevation dropped off in direct accord with decrease in extracellular Ca2+ concentration. The second step of the [Ca2+]m increase was suppressed significantly in the absence of extracellular Ca2+ and in knockdown cells of stromal interaction molecule 1 (STIM1), an essential molecule on endoplasmic reticulum (ER) membrane for store-operated Ca2+ entry, in the presence of extracellular Ca2+ (1.0 mM), while the first elevation was not affected in either case. The results indicate that mitochondria take up cytosolic Ca2+ in two steps; first and second uptakes are derived from the Ca2+ release from ER and the Ca2+ influx through store-operated Ca2+ channels, respectively. Additionally, rotenone and antimycin A, which are inhibitors of mitochondrial electron transport complex I and III, respectively, diminished mitochondrial Ca2+ uptake and significantly suppressed degranulation stimulated with antigen. The mitochondrial Ca2+ uptake may modulate mast cell function by regulating the [Ca2+]c.
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  • You Jin Lee, Jinhee Kim, Jin-Mu Yi, Se-Mi Oh, No Soo Kim, Haejin Kim, ...
    Type: Note
    2012 Volume 35 Issue 8 Pages 1361-1366
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Activity-guided fractionation of an 80% EtOH extract from the aerial parts of Saururus chinensis led to isolation of three anti-proliferative neolignans (13) along with four flavonoids (47) and four aristolactams (811). Their chemical structures were identified by analysis of spectroscopic data. All compounds 111 were evaluated for their activities against 28 human cancer cell lines using an in vitro cell proliferation assay. Compounds 13 showed potent anti-proliferative activities against cervical (C33a, IC50=0.01 µM for 1; 0.28 µM for 2; 2.80 µM for 3) and lung (NCI-H460, IC50=0.05 µM for 1; 1.37 µM for 2; 6.46 µM for 3) cancer cells without any remarkable cytotoxic effects on human normal lung cells as a control. Taken together, these data demonstrated the identification of anti-proliferative neolignans which are active components of S. chinensis.
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  • Mizuki Iwama, Yoshitaka Kondo, Kentaro Shimokado, Naoki Maruyama, Akih ...
    Type: Note
    2012 Volume 35 Issue 8 Pages 1367-1370
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Here we quantified the uric acid (UA) levels in 11 tissues and plasma of C57BL/6 male mice to track its turnover during 3 to 30 months of aging. UA levels in the adrenal glands, heart, and spleen increased with aging until 30 months of age. Similarly, UA levels in the liver, kidneys, pancreas, and testes increased until the mice were 24 months old. UA levels also rose in the lungs and skeletal muscles from 3 to 6 months and 6 to 12 months, respectively, and then remained at almost the same levels until 30 months of age. In the skin, UA decreased from 3 to 6 months and then stayed nearly constant until 30 months of age. Moreover, the small intestines and plasma had quite stable UA levels during aging. Thus, our assessment of 11 tissue types from mice showed that the UA levels increased in most tissues during aging.
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  • Tamaki Cho, Jun-ichi Nagao, Rieko Imayoshi, Jun Kodama, Takahiko Morot ...
    Type: Note
    2012 Volume 35 Issue 8 Pages 1371-1373
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Elevation in the temperature induces heat stress to both host cells and the invading pathogen. This study aimed to determine whether continuous mild heat stress (increased temperature without causing significant damage to host cells) can increase susceptibility of biofilm formation of the opportunistic fungal pathogen Candida albicans to low concentrations of three typical antifungal agents. In this way the side effects associated with higher concentrations of the antifungal agents on host cells would be reduced. Fluconazole and micafungin at concentrations ranging from 0.0625 to 2 µg/mL and amphotericin B at concentrations ranging from 0.0625 to 1 µg/mL inhibited less than 20% of cells in biofilm formation. Biofilm formation at 39 or 41°C compared to 37°C resulted in increased susceptibility to the three agents, but especially micafungin. These data suggest that mild heat stress (39°C) would be valuable for increasing the effectiveness of low concentrations of antifungal agents against C. albicans biofilm formation. Thus, the concept of continuous mild heat stress at the site of insertion of medical devices or catheters combined with antifungal agents could be beneficial.
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  • Ye Jin, Hyun Ok Yang, Jong Keun Son, Hyeun Wook Chang
    Type: Note
    2012 Volume 35 Issue 8 Pages 1374-1378
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Pinusolide, an herbal medicine isolated from Biota orientalis L. (B. orientalis), inhibited 5-lipoxygenase (5-LO)-dependent leukotriene C4 (LTC4) generation in immunoglobulin E (IgE)/Ag-induced bone marrow-derived mast cells (BMMCs) in a concentration-dependent manner. To clarify the action mechanism of pinusolide on the inhibition of LTC4 generation, we examined the effect of pinusolide on phosphorylation of cytosolic phospholipase A2 (cPLA2), as well as translocation phospho-cPLA2 and 5-LO to nucleus. Inhibition of LTC4 generation by pinusolide was accompanied by a decrease in cPLA2 phosphorylation which occurred via a decrease in intracellular Ca2+ influx and blocking the c-Jun N-terminal kinase (JNK) pathways. However, pinusolide had no effect on extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein (MAP) kinas phosphorylation. Taken together, the present results suggest that potent inhibition of 5-LO dependent LTC4 generation by pinusolide requires both suppression of calcium influx and JNK phosphorylation.
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  • Thi Kim Van Le, Jin Ju Jeong, Dong-Hyun Kim
    Type: Note
    2012 Volume 35 Issue 8 Pages 1379-1383
    Published: August 01, 2012
    Released: August 01, 2012
    JOURNALS FREE ACCESS
    Phosphatidylinositol 3-kinase (PI3K)/Akt inhibitors were isolated from the rhizome of Polygala tenuifolia WILLD (PT, Polygalaceae), which has been used in traditional Chinese medicine for inflammation, dementia, amnesia, neurasthenia and cancer, by activity-guided fractionation. For the assay of PI3K/Akt pathway, cytoprotective Tat-transduced CHME5 cells, which are the cytoprotective phenotype against lypopolysaccharide (LPS)/cycloheximide (CHX), were used. We isolated 4 anti-cytoprotective compounds, clionasterol (1), ethyl cholestan-22-enol (2), 3-O-β-D-glucosyl ethyl cholestan-22-enol (3), and 3-O-β-D-glucopyranosyl clionasterol (4) from EtOAc fraction of PT against Tat-transduced CHME5 cells. Of them, (1) and (2) most potently abolished cytoprotective effect of Tat-transduced CHME5 cells. These constituents (1) and (2) inhibited the activation of 3-phosphoinositide-dependent kinase 1 (PDK1) and its downstream molecules, Akt/glycogen synthase kinase (GSK)3β, in PI3K/Akt cell survival signaling pathway, but did not suppress the activation of PI3K. Based on these finding, (1) and (2) may abolish the cytoprotective phenotype of Tat-transduced CHME5 cells by inhibiting PDK1 phosphorylation in PI3K/Akt pathway.
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