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Toshiyuki Hata, Yu Shibata, Miku Okano, Asako Kodera, Misato Ueda, Hir ...
2015 Volume 38 Issue 3 Pages
358-364
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: January 14, 2015
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The ESR spectra of dicupric human serum-transferrin (serum-Tf) were measured from −20 to 37°C in the liquid state (56% glycerol at pH 7.6). Two coordination geometries (types B-1 and B-2) with different ESR parameters were present at the N-site. The contents of the coordination geometry of type B-1 at the N-site increased as the temperature increased. The equilibrium constant between the coordination geometries of types B-1 and B-2 was determined by ESR spectra. The enthalpy value from type B-2 to B-1 was +5.3 kcal/mol, as obtained from a van’t Hoff plot. The two conformational energies of the cluster models of the copper-binding site at the N-site of dicupric human serum-Tf, where the Arg124 residue was oriented in two different directions (conformations I and II), were calculated by Density Functional Theory, and the enthalpy value from conformation II to I was +2.1 kcal/mol. The enthalpy value was similar to that (+5.3 kcal/mol) obtained by the coordination geometrical change from type B-2 to B-1 in Cu(II)
2 serum-Tf. In conformations I and II, the residue of Arg124 at the N-site is located either far from or near the copper-binding site, respectively, and in both cases the coordination geometry of the cupric ions at the N-site has changed from a flattened tetrahedron to a trigonal bipyramid. This result implies that the ESR spectral change from type B-2 to B-1 is caused by the presence of two different orientations of Arg124 in the change from conformation II to I.
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Dan Wu, Ying-shu Quan, Fumio Kamiyama, Kosuke Kusamori, Hidemasa Katsu ...
2015 Volume 38 Issue 3 Pages
365-373
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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The purpose of the present study was to develop an alternative transdermal formulation containing sumatriptan succinate (SS) for the treatment of migraine. Novel self-dissolving SS-loaded microneedle arrays (MNs) were fabricated from sodium hyaluronate and their efficacy for transdermal delivery of SS was characterized. The resulting MNs maintained their skin piercing abilities for at least 30 min after being placed at a high relative humidity of 75%. Rapid release of SS from the MNs was also observed
in vitro. Optical coherence tomography images demonstrated that MNs were able to successfully pierce into rat skin without any bending or cracking, and needles were completely dissolved within 1 h. MNs significantly increased transepidermal water loss; however, skin barrier function gradually recovered to control levels within 24 h, in contrast to the skin damage observed after tape stripping treatment. These findings indicated that the micropores created by MNs quickly resealed, and that the skin damage was reversible. Furthermore, a dose-dependent plasma concentration of SS was obtained after transdermal delivery using SS-loaded MNs in rats. Absorption of SS delivered by MNs was similar to that observed after subcutaneous injection and was associated with high bioavailability (
ca. 90%), which was much higher than that produced by oral administration. These findings suggested that application of SS-loaded MNs to the skin provided an effective alternative approach to enhance the transdermal delivery of SS without serious skin damage, and would be likely to improve patient compliance.
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Keiichi Motoyama, Risako Onodera, Nao Tanaka, Kazuhisa Kameyama, Taish ...
2015 Volume 38 Issue 3 Pages
374-379
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Melanoma is a life-threatening disorder and its incidence is increasing gradually. Despite the numerous treatment approaches, conventional systemic chemotherapy has not reduced the mortality rate among melanoma patients, probably due to the induction of toxicity to normal tissues. Recently, we have developed folate-conjugated methyl-β-cyclodextrin (FA-M-β-CyD) and clarified its potential as a new antitumor agent involved in autophagic cell death. However, it remains uncertain whether FA-M-β-CyD exerts anticancer effects against melanomas. Therefore, in this study, we investigated the effects of FA-M-β-CyD on the folate receptor-α (FR-α)-expressing melanoma cell-selective cytotoxic effect. FA-M-β-CyD showed cytotoxic effects in Ihara cells, a human melanoma cell line expressing FR-α. In sharp contrast to methyl-β-cyclodextrin, FA-M-β-CyD entered Ihara cells [FR-α(+)] through FR-α-mediated endocytosis. Additionally, FA-M-β-CyD elicited the formation of autophagosomes in Ihara cells. Notably, FA-M-β-CyD suppressed melanoma growth in BALB/c nude recombinase-activating gene-2 (Rag-2)/Janus kinase 3 (Jak3) double deficient mice bearing Ihara cells. Therefore, these results suggest that FA-M-β-CyD could be utilized as a potent anticancer agent for melanoma chemotherapy by regulating autophagy.
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Hassan Rashidzadeh, Sanjeev Bhadresa, Steven Spencer Good, Marita Lars ...
2015 Volume 38 Issue 3 Pages
380-388
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Supplementary material
A conventional, rapid and high throughput method for tissue extraction and accurate and selective LC-MS/MS quantification of 2′-
C-methylguanosine triphosphate (2′-MeGTP) in mouse liver was developed and qualified. Trichloroacetic acid (TCA) was used as the tissue homogenization reagent that overcomes instability challenges of liver tissue nucleotide triphosphates due to instant ischemic degradation to mono- and diphosphate nucleotides. Degradation of 2′-MeGTP was also minimized by harvesting livers using
in situ clamp-freezing or snap-freezing techniques. The assay also included a sample clean-up procedure using weak anion exchange solid phase extraction followed by ion exchange chromatography and tandem mass spectrometry detection. The linear assay range was from 50 to 10000 pmol/mL concentration in liver homogenate (250–50000 pmol/g in liver tissue). The method was qualified over three intraday batches for accuracy, precision, selectivity and specificity. The assay was successfully applied to pharmacokinetic studies of 2′-MeGTP in liver tissue samples after single oral doses of IDX184, a nucleotide prodrug inhibitor of the viral polymerase for the treatment of hepatitis C, to mice. The study results suggested that the clamp-freezing liver collection method was marginally more effective in preventing 2′-MeGTP degradation during liver tissue collection compared to the snap-freezing method.
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Young Ho Seo, Yoo-na Jo, Yong Jin Oh, Soyeun Park
2015 Volume 38 Issue 3 Pages
389-395
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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The mechanical properties of cells are considered promising biomarkers for the early detection of cancer and the testing of drug efficacy against it. Nevertheless, generalized correlations between drug resistance and the nano-mechanical properties of cancer cells are yet to be defined due to the lack of necessary studies. In this study, we conducted atomic force microscopy (AFM)-based nano-mechanical measurements of cisplatin-sensitive (A2780) and cisplatin-resistant (A2780cis) ovarian cancer cells. The difference in the efficacy of cisplatin between A2780 and A2780cis was confirmed in the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2
H-tetrazolium (MTS) assay. We observed that the cisplatin-resistant ovarian cancer cells were more motile than cisplatin-sensitive cells based on the results of the wound closure experiment, and the AFM experiments showed that drug resistance induced nano-mechanical stiffening of the ovarian cancer cells. Increased mechanical stiffness caused by cisplatin resistance was consistent with the confocal microscopy images showing more distinct actin stress fibers in A2780cis than in A2780 cells. The down regulation of vinculin implicated the actin-driven elongation as a major motile mode for A2780cis cells. Our results consistently indicated that the acquisition of drug resistance in ovarian cancer cells induces an extensive reorganization of the actin cytoskeleton, which governs the cellular mechanical properties, motility, and possibly intracellular drug transportation.
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Shan-Gao Li, Jun Zhou, Ji-Hong Zhong, Hai-Jun Cao, Ling Zhu, Jun Liu, ...
2015 Volume 38 Issue 3 Pages
396-401
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: December 18, 2014
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Micro-RNAs (miRNAs) are involved in regulation of the incidence and development of several hepatic diseases. Thus manipulating miRNAs may be a promising therapeutic strategy against these entities. In this study hepatic stellate cells (HSCs) were transfected with hsa-miR-9 or anti-hsa-miR-9, treated with tetramethylpyrazine (TMP), or subjected to treatment with TMP and hsa-miR-9 transfection (combined treatment group). Then, real-time polymerase chain reaction (PCR) was performed to measure mRNA levels of hsa-miR-9. Expression of hsa-miR-9 was highest in the combination treatment group compared with other groups, and significantly higher than TMP-treated and hsa-miR-9-transfected groups (both
p<0.05). The anti-hsa-miR-9-transfected group expressed the lowest mRNA level of hsa-miR-9 with marked decrease
versus control (
p<0.05). Downstream factors that may be affected by miR-9 such as leptin, α-smooth muscle actin (SMA), and collagen I, as well as phosphorylation levels of Janus kinase 1 (JAK1)/signal transducer and activator of transcription 3 (STAT3) were investigated at the protein level. All these factors were regulated contrariwise to expression trends of hsa-miR-9, showing the lowest level in the combination treatment group and highest level in anti-hsa-miR-9-transfected group. These results suggest that both transfection of hsa-miR-9 and TMP can lead to upregulated endogenous expression of hsa-miR-9, inhibit activation of JAK1/STAT3 signal pathway induced by leptin, and lead to reduction of α-SMA and collagen I—thus impeding activation of HSC.
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Masashi Nagata, Yasuyoshi Ishiwata, Yutaka Takahashi, Hiromitsu Takaha ...
2015 Volume 38 Issue 3 Pages
402-410
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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The aim of the present study was to clarify the therapeutic range and adequate dose of sunitinib in Japanese renal cell carcinoma patients by means of a pharmacokinetic–pharmacodynamic analysis of sunitinib-induced thrombocytopenia. Six patients with renal cell carcinoma were enrolled in this study. After starting the sunitinib treatment, between three and seven blood samples were obtained from each patient just before the administration of sunitinib. Serum concentrations of sunitinib and its active metabolite
N-desethyl-sunitinib were fit to the 1-compartment model with first-order absorption. Changes in platelet counts were fit to the pharmacokinetic–pharmacodynamic model, in which the proliferation of platelet progenitor cells was assumed to be linearly inhibited by sunitinib and its metabolite. All patients using 50 mg as an initial dose of sunitinib developed grade 2 or 3 thrombocytopenia. The pharmacokinetic–pharmacodynamic model created successfully described the time course of sunitinib-induced thrombocytopenia and could predict changes in platelet counts after alterations to the dosage of sunitinib administered. The simulation results indicated that the total trough level of sunitinib to avoid severe thrombocytopenia should be <100 ng/mL, and also that the initial daily dose of sunitinib could be reduced to 37.5 mg or 25 mg in most Japanese patients. In addition to the pharmacokinetic-guided dosage adjustment, the careful monitoring of platelet counts is required for the safe use of sunitinib.
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Masako Yokoo, Yasushi Kubota, Yoko Tabe, Shinya Kimura
2015 Volume 38 Issue 3 Pages
411-416
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: January 08, 2015
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Supplementary material
Long-term treatment with imatinib mesylate (IM) allows patients with chronic myeloid leukemia (CML) to live a near-normal lifespan. However, the fact that tyrosine kinase inhibitors, including IM, are extremely expensive is a major cause of poor adherence, resulting in disease relapse or drug resistance. Therefore, physicians are encouraged to prescribe generic drugs to reduce the financial burden of medical expenses. In Japan, only generic drugs that have a basic chemical structure and pharmacokinetic data that are the same as those of the original drug are approved. However, it is not mandatory to demonstrate that generic drugs have adequate biological effects. This is one of the reasons why Japanese hematologists do not often use generic IM. The aim of the present study was to compare the anti-leukemic effects of Glivec™ (a commercial IM) and its generic formulation, OHK9511. The IC
50 values of OHK9511 and Glivec™ were comparable, and both induced similar levels of apoptosis in several CML cell lines. Furthermore, the overall survival of OHK9511-treated mice transplanted with BCR-ABL-positive cells was similar to that of mice treated with Glivec™. Although the experiments performed herein were basic, the results suggest that physicians should consider using generic IM.
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Yosuke Hashimoto, Taro Shimizu, Amr Selim Abu Lila, Tatsuhiro Ishida, ...
2015 Volume 38 Issue 3 Pages
417-424
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Supplementary material
PEGylation, which is the surface modification of nanocarriers with polyethylene glycol (PEG), has increased the circulation time and reduced the immunogenic responses to nanocarriers. However, many reports have demonstrated that the intravenous injection of sterically stabilized PEGylated liposome (SL) causes an accelerated blood clearance (ABC) of subsequent doses
via anti-PEG immunoglobulin M (IgM)-mediated complement activation. In the present study, the relationships between serum anti-PEG IgM concentration, the intensity of complement activation and the hepatic clearance of SL were quantitatively investigated for their role in the ABC phenomenon. Interestingly, with increasing serum anti-PEG IgM concentrations, the intensity of complement activation increased linearly, while the intensity of the hepatic clearance of SL was increased and then saturated. In addition, only 15–17% of anti-PEG IgM in blood circulation induced by SL at different doses was associated with a second dose SL. The present results indicate that it is the hepatic uptake of SL that is the limiting step in the ABC phenomenon, rather than the association of anti-PEG IgM to the SL and a subsequent complement activation.
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Soo-Jin Jeong, Hye-Sun Lim, Chang-Seob Seo, Seong-Eun Jin, Sae-Rom Yoo ...
2015 Volume 38 Issue 3 Pages
425-434
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Gyejibokryeong-hwan (GJBRH; Keishi-bukuryo-gan in Japan and Guizhi Fuling Wan in China) is a traditional herbal formula comprising five medicinal herbs and is used to treat climacteric syndrome. GJBRH has been shown to exhibit biological activity against diabetes, diabetic nephropathy, atherosclerosis, ischemia, and cancer. However, there is no scientific evidence of its activities against skin inflammation, including atopic dermatitis. We used the HaCaT human keratinocyte cell line to investigate the effects of GJBRH on skin inflammation. No significant cytotoxicity was observed in cells treated with GJBRH up to a concentration of 1000 µg/mL. Exposure to the proinflammatory cytokines tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) significantly increased HaCaT cell production of the following chemokines: macrophage-derived chemokine (MDC)/CCL22; regulated on activation, normal T-cell expressed and secreted (RANTES)/CCL5; and interleukin-8 (IL-8). In contrast, GJBRH significantly reduced the production of MDC, RANTES, and IL-8 compared with control cells simulated with TNF-α and IFN-γ. Consistently, GJBRH suppressed the mRNA expression of MDC, RANTES, and IL-8 in TNF-α and IFN-γ-treated cells. Treatment with GJBRH markedly inhibited phosphorylation of signal transducer and activator of transcription 1 (STAT1) in HaCaT cells stimulated with TNF-α and IFN-γ. Our findings indicate that GJBRH impairs TNF-α and IFN-γ-mediated inflammatory chemokine production and STAT1 phosphorylation in keratinocytes. We suggest that GJBRH may be a potent therapeutic agent for inflammatory skin disorders.
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Shota Tanaka, Mika Hosokawa, Takeshi Yonezawa, Wataru Hayashi, Kumiko ...
2015 Volume 38 Issue 3 Pages
435-440
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: December 27, 2014
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Supplementary material
Epithelial-mesenchymal transition (EMT) and changes in the expression of the microRNA-200 (miR-200) family were examined in the human colorectal cancer (CRC) cell line SW620 with acquired oxaliplatin (L-OHP) resistance. Two CRC cell lines, SW480, derived from primary CRC, and SW620, derived from lymph node metastasis, which were obtained from the same patient, were used in the present study. L-OHP-resistant SW620 cells were obtained by exposure to L-OHP for 155 d. The concentration of L-OHP was increased to 80 µM in a stepwise manner. The IC
50 value of L-OHP was increased 16-fold in L-OHP-resistant SW620 cells, which also displayed mesenchymal cell-like characteristics, such as the down-regulation of E-cadherin and up-regulation of vimentin. However, L-OHP-resistant SW480 cells were not obtained when the concentration of L-OHP was increased in a similar stepwise manner. The expression levels of members of the miR-200 family (miR-200a, miR-200b, miR-429, miR-200c, and miR-141) were significantly higher in SW480 cells than in SW620 cells. The expression levels of miR-200c and miR-141 were significantly lower in L-OHP-resistant SW620 cells than in control SW620 cells. L-OHP-resistant SW620 cells did not exhibit cross-resistance to other anti-cancer drugs used to treat CRC, such as 5-fluorouracil, irinotecan, and the active metabolite of irinotecan (SN-38). These results suggest that the down-regulated expression of miR-200c and miR-141 plays a role in selective resistance to L-OHP and EMT in CRC cells during repeated treatments with L-OHP.
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Possible Mechanism of Iguratimod–Warfarin Interaction
Satoshi Yamaori, Ken Takami, Ayaka Shiozawa, Kanako Sakuyama, Naoki Ma ...
2015 Volume 38 Issue 3 Pages
441-447
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: January 19, 2015
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Iguratimod is a novel disease-modifying antirheumatic drug. A blue letter (safety advisory) for drug interaction between iguratimod and warfarin was issued by the Ministry of Health, Labour and Welfare of Japan in May 2013. Iguratimod may affect warfarin metabolism catalyzed by CYP. However, it is not clear whether iguratimod inhibits warfarin oxidation. This study was performed to investigate the effects of iguratimod on warfarin 7-hydroxylation with human liver microsomes (HLMs) and recombinant CYP enzymes. Iguratimod concentration-dependently inhibited
R,
S-warfarin 7-hydroxylase activity of HLMs with an IC
50 value of 15.2 µM. The inhibitory effect was examined with
S-warfarin and
R-warfarin to determine which enantiomer was more potently inhibited by iguratimod. Iguratimod potently inhibited the
S-warfarin 7-hydroxylase activity of HLMs with an IC
50 value of 14.1 µM, but showed only slight inhibition of
R-warfarin 7-hydroxylation. Furthermore, iguratimod inhibited the
S-warfarin 7-hydroxylase activity of recombinant CYP2C9.1 (rCYP2C9.1) and rCYP2C9.3 in a concentration-dependent manner with IC
50 values of 10.8 and 20.1 µM, respectively. Kinetic analysis of the inhibition of
S-warfarin 7-hydroxylation by iguratimod indicated competitive-type inhibition for HLMs and rCYP2C9.1 but mixed-type inhibition for rCYP2C9.3. The
Ki values for HLMs, rCYP2C9.1, and rCYP2C9.3 were 6.74, 4.23, and 14.2 µM, respectively. Iguratimod did not exert metabolism-dependent inhibition of
S-warfarin 7-hydroxylation. These results indicated that iguratimod is a potent direct inhibitor of CYP2C9-mediated warfarin 7-hydroxylation and that its inhibitory effect on CYP2C9.1 was more sensitive than that on CYP2C9.3.
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Takeshi Chiba, Soichiro Kimura, Katsuo Takahashi, Yasunori Morimoto, T ...
2015 Volume 38 Issue 3 Pages
448-453
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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We previously reported that serotonin (5-hydroxytryptamine; 5-HT) suppresses β-casein expression, a differentiation marker in mammary epithelial cells,
via inhibition of the signal transducer and activator of transcription 5 (STAT5) phosphorylation in the human mammary epithelial cell line, MCF-12A. In this study, we investigated the expression pattern of the different 5-HT receptor subtypes in MCF-12A cells, and identified the receptors involved in 5-HT-mediated suppression of β-casein protein expression. β-Casein mRNA expression was inhibited by 30 µM 5-HT in a time-dependent manner. Treatment with 30 µM 5-HT for 72 h decreased β-casein protein levels and STAT5 phosphorylation (pSTAT5). The cells expressed four 5-HT receptors subtypes (5-HTR
1D, 2B, 3A, and 7) at the mRNA and protein level, and their expression was elevated by prolactin (PRL) treatment. Additionally, the mRNA levels of 5-HTR
1D and 5-HTR
7 were significantly higher than the other 5-HT receptors in the cells. Tryptophan hydroxylase 1 mRNA was detectable in the cells in the absence of PRL, and PRL treatment significantly increased its expression. β-Casein and pSTAT5/STAT5 levels in the cells co-treated with 5-HT and a selective 5-HTR
1D inhibitor, BRL15572, were equal to those observed in cells treated with 5-HT alone. However, in the cells co-treated with 5-HT and a selective 5-HTR
7 inhibitor, SB269970, β-casein and pSTAT5/STAT5 levels increased in a SB269970 concentration-dependent manner. In conclusion, we showed that 5-HT regulates β-casein expression
via 5-HTR
7 in MCF-12A human mammary epithelial cells.
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Motohiro Hoshino, Nobutomo Ikarashi, Ryuta Hirobe, Mami Hayashi, Hiron ...
2015 Volume 38 Issue 3 Pages
454-460
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
Advance online publication: January 14, 2015
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We have previously shown that menthol attenuates the anticoagulant effect of warfarin by increasing the expression levels of CYP3A and CYP2C in the liver. This study evaluated the effects of menthol on the pharmacokinetics of the CYP3A substrate triazolam and the CYP2C substrate phenytoin. Menthol was orally administered to mice for 7 d. Twenty-four hours after the administration of menthol, triazolam was orally administered, and the plasma concentration was measured. In addition, the CYP3A metabolic activity for triazolam and the CYP3A expression level in the liver were determined. The effects of menthol on the pharmacokinetics of phenytoin were assessed in the same manner. In the menthol-treated group, the area under the blood concentration–time curve (
AUC) of triazolam was lower and its clearance was higher compared with the control group. The CYP3A metabolic activity and CYP3A expression level in the liver were significantly increased in the menthol-treated group compared with the control group. Similarly, the
AUC of phenytoin was lower and the hepatic CYP2C expression level was higher in the menthol-treated group. Thus, menthol lowered the plasma concentrations of triazolam and phenytoin when concurrently administered. These effects may be attributed to an increased metabolic activity for these drugs due to the increased expression of CYP3A and CYP2C in the liver.
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Noha Essam Eldin, Hanan Mohamed Elnahas, Mahmoud Abd-Elghany Mahdy, Ta ...
2015 Volume 38 Issue 3 Pages
461-469
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Pemetrexed (PMX) is a newly developed multi-targeted anti-folate with promising clinical activity in many solid tumors including malignant pleural mesothelioma (MPM). However, PMX does not show sufficient anti-tumor activity
in vivo when used alone either due to inefficient delivery of adequate concentrations to tumor tissue or dose-limiting side effects. In order to overcome these problems and to achieve potent anti-tumor activity, PMX was encapsulated into a liposomal delivery system. In the present study, various formulations of liposomal PMX were prepared. The effect of formulation parameters on the encapsulation efficiency of PMX within liposomes was evaluated. In addition, the influence of drug release rate on the
in vitro cytotoxicity was investigated. Encapsulation of PMX within liposomes was remarkably increased by the incorporation of cholesterol within liposomal membranes and by increasing the total lipid concentration. Encapsulation efficiency was found to be unaffected by the type of phospholipid used or the inclusion of a cation lipid, DC-6-14. Interestingly, encapsulation of PMX within “fluid” liposomes was found to allow efficient release of PMX from liposomes resulting in a potent
in vitro cytotoxicity against MPM MSTO-211H cell line. On the other hand, entrapment of PMX within “solid” liposomes substantially hindered PMX release from liposomes, and thus PMX failed to exert any
in vitro cytotoxicity. These results suggest that encapsulation of PMX within “fluid” liposomes might represent a novel strategy to enhance the therapeutic efficacy of PMX while minimizing the side effect encountered by the non selective delivery of free PMX to various body tissues.
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Xianqin Wang, Meiling Zhang, Jianshe Ma, Yuan Zhang, Guangliang Hong, ...
2015 Volume 38 Issue 3 Pages
470-475
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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Numerous people die of paraquat (PQ) poisoning every year in the world. Although several studies regarding paraquat (PQ) poisoning have been conducted, the metabolic changes in plasma remain unknown. In this study, the metabolomics of 15 PQ poisoned patients with plasma PQ concentrations in excess of 0.1 µg/mL and 16 healthy volunteers were investigated. The plasma samples were evaluated through the use of gas chromatography-mass spectrometry (GC/MS) and analyzed by partial least-squares discriminant analysis (PLS-DA). Based on the metabolomics data, a support vector machine (SVM) discrimination model was developed. The results showed the plasma levels of urea, glucose oxime and L-phenylalanine decreased and cholesterol increased in PQ poisoned patients in comparison to healthy volunteers. The SVM discrimination model was developed, and performed with a high degree of accuracy, to distinguish PQ poisoned patients from healthy volunteers. In conclusion, metabolic pathways including the urea cycle, and amino acid, glucose, and cholesterol metabolism were impaired after PQ poisoning. An SVM discrimination model, based on metabolomics data, was established and may become a new powerful tool for the diagnosis of PQ poisoning.
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Yukiko K. Kaneko, Miki Takii, Yumiko Kojima, Hiroko Yokosawa, Tomohisa ...
2015 Volume 38 Issue 3 Pages
476-481
Published: March 01, 2015
Released on J-STAGE: March 01, 2015
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The effects of green tea catechins on glucose-stimulated insulin secretion (GSIS) were investigated in the β-cell line INS-1D. Epigallocatechin gallate (EGCG) at 10 µM or gallocatechin gallate (GCG) at 30 µM caused significant inhibitory effects on GSIS, and each of these at 100 µM almost abolished it. In contrast, epicatechin (EC) or catechin (CA) had no effect on GSIS at concentrations up to 100 µM. We thus investigated the structure–activity relationship by using epigallocatechin (EGC) and gallocatechin (GC) containing a trihydroxyl group in the B-ring, and epicatechin gallate (ECG) and catechin gallate (CG) containing the gallate moiety. EGC, GC, and ECG caused an inhibition of GSIS, although significant effects were obtained only at 100 µM. At this concentration, EGC almost abolished GSIS, whereas GC and ECG partially inhibited it. In contrast, CG did not affect GSIS at concentrations up to 100 µM. EGCG also abolished the insulin secretion induced by tolbutamide, an ATP-sensitive K
+ channel blocker, and partially inhibited that induced by 30 mM K
+. Moreover, EGCG, but not EC, inhibited the oscillation of intracellular Ca
2+ concentration induced by 11.1 mM glucose. These results suggest that some catechins at supraphysiological concentrations have inhibitory effects on GSIS, the potency of which depends on their structure; the order of potency was EGCG>GCG>EGC>GC≈ECG. The inhibitory effects seem to be mediated by the inhibition of voltage-dependent Ca
2+ channels, which is caused, at least in part, by membrane hyperpolarization resulting from the activation of K
+ channels.
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