In recent years, considerable emphasis has been focused on identifying new chemopreventive agents which could be useful for the human population. In the present study, we examined the protective role of mangiferin during experimental lung carcinogenesis with reference to its effect on DNA-damage and the detoxification enzyme system. The activities of detoxifying enzymes such as glutathione transferase (GST), quinone reductase (QR) and uridin 5′-diphosphate-glucuronosyl transferase (UDP-GT) were found to be decreased while the lipid peroxidation level was increased in the lung cancer bearing animals. Supplementation of mangiferin (100 mg/kg b.wt) enhanced the detoxification enzymes and reduced DNA damage as determined by single cell electrophoresis. Furthermore, the DNA–protein cross links which was found to be high in lung cancer bearing animals was also modulated upon supplementation with mangiferin. Our present results explain the unique association between the anti-oxidant effect of mangiferin and ultimately the capability of mangiferin to prevent cancer.

抄録全体を表示

Saliva contains a large number of proteins that participate in the protection of oral tissue. We found, for the first time, small vesicles (30—130 nm in diameter) in human whole saliva. Vesicles from saliva were identified by electron microscopy after isolation by gel-filtration on Sepharose CL-4B. They resemble exosomes, which are vesicles with an endosome-derived limiting membrane that are secreted by a diverse range of cell types. We performed a biochemical characterization of these vesicles by amino acid sequence analysis and Western blot analysis. We found that they contain dipeptidyl peptidase IV (DPP IV), galectin-3 and immunoglobulin A, which have potential to influence immune response. The DPP IV in the vesicles was metabolically active in cleaving substance P and glucose-dependent insulinotropic polypeptide to release N-terminal dipeptides. Our results demonstrate that human whole saliva contains exosome-like vesicles; they might participate in the catabolism of bioactive peptides and play a regulatory role in local immune defense in the oral cavity.

抄録全体を表示

An alkaloid piperine isolated from the Piper Nigrum was found to inhibit lipid droplet accumulation in mouse macrophages, and especially inhibited cholesteryl ester (CE) synthesis (IC₅₀: 25 μM). The metabolism of cholesterol from lysosome to lipid droplet was inhibited with a similar IC₅₀ (18 μM), indicating that the site of inhibition is one of the steps between the lysosomes and the endoplasmic reticulum. Therefore, effects of piperine on acyl-CoA:cholesterol acyltransferase (ACAT) activity in the microsomes prepared from mouse macrophage and liver were studied, to show that the compounds inhibited the activity in both cases (IC₅₀: 9.1, 7.0 μM, respectively). Furthermore, piperine was found to inhibit both ACAT1 and ACAT2 isozymes to a similar extent (IC₅₀: 16, 18 μM, respectively) in cell-based assays using ACAT1- or ACAT2-expressing cells. Thus, it was suggested that piperine inhibited macrophage ACAT to decrease CE synthesis, leading to a reduction of lipid droplets.

抄録全体を表示

The basic disaccharide structure recognized by galectin family members is the lactosamine-like structure Galβ1-4(3)Glc(NAc). The 32-kDa galectin LEC-1 of the nematode Caenorhabditis elegans is composed of two domains, each of which is homologous to vertebrate 14-kDa-type galectins. The N-terminal lectin domain of LEC-1 recognizes blood group A saccharide (GalNAcα1-3(Fucα1-2)Galβ1-3GlcNAc), whereas this saccharide is poorly recognized by the C-terminal domain. Using a combination of site-directed mutagenesis and analysis of the sugar-binding profile by frontal affinity chromatography, we previously found that Thr⁴¹ in the N-terminal lectin domain of LEC-1 is important for its affinity for A-hexasaccharide. Thr⁴¹ is located on β-strand S3, next to the three β-strands S4—S6, where the conserved amino acids form the binding site for the basic Galβ1-4(3)Glc(NAc) structure. Here, we report that a second amino acid, Asp¹³³, in the N-terminal lectin domain of LEC-1, located on the β-strand S2 adjacent to that containing Thr⁴¹, is important for LEC-1-specific recognition of A-hexasaccharide. These results suggest that amino acid residues other than those located on the three β-strands S4—S6, contribute to the unique sugar binding specificity of individual galectins.

抄録全体を表示

Plant lignans are converted by the intestinal microflora to the mammalian lignans enterodiol and enterolactone, which are associated with beneficial health effects in humans. The mammalian lignans enterodiol and enterolactone are believed to protect against certain diseases, e.g., breast and prostate cancer as well as coronary heart disease. In this study, the effects of enterodiol and enterolactone on osteoblastic differentiation were examined. It was found that enterolactone and enterodiol showed the same effects. They have biphasic effects on cell viability, alkaline phosphotase (ALP) activity, transcriptional level of osteonectin, and collagen I, showing induction at low doses and inhibition at high doses. They increased transcriptional levels of ALP, osteopontin, and osteocalcin in a dose-dependent manner. The difference may be related to the estrogenic and antiestrogenic character and multiple signaling transduction of phytoestrogen.

抄録全体を表示

Neuroblastoma (NB), which is a malignant tumor of young children derived from neural crest cells that occurs in children, exhibits a wide range of clinical behaviors, from spontaneous regression to rapid progression. Advanced NB patients have a poor prognosis, and recently, autologous bone marrow transplantation (BMT) and autologous peripheral blood stem cell transplantation (PBSCT) have been attempted to improve the prognosis of these patients. In this study, we attempted to detect the expression of tyrosine hydroxylase (TH), neuroendocrine protein gene product (PGP) 9.5, ELAVL-4 and GD2 synthetase (GALGT), all of which are highly expressed in NBs, by the reverse transcription-polymerase chain reaction (RT-PCR) technique in order to detect minimal residual disease (MRD) in the bone marrow (BM) and peripheral blood (PB). Analysis of various tumor cell lines (Ewing's sarcoma, hepatoma, leukemias, and breast cancer cell lines in addition to NBs), and human normal samples (BM and PB cells) revealed that TH was the most specific marker for the detection of NB. On the other hand, PGP9.5 was the most sensitive marker, and was detected even when there was only one positive cell per 10⁷ negative cells. We concluded that TH is a better marker before the diagnosis of NB while PGP9.5 is a better marker to detect MRD after the diagnosis. Here, we describe our results on useful markers to detect MRD in patients with NB.

抄録全体を表示

Two anthraquinones which inhibit activity of the Src tyrosine kinase were isolated from a water extract of Hedyotis diffusa WILLD. and identified as 2-hydroxy-3-methylanthraquinone (compound 1) and 1-methoxy-2-hydroxyanthraquinone (compound 2). Both compounds showed inhibitory activity against protein tyrosine kinases v-src and pp60src and arrested the growth of SPC-1-A, Bcap37 and HepG2 cancer cells. Observation of mitochondrial membrane potential collapse and caspase-3 activation following treatment with the compounds indicates that their apoptotic induction activity may act via the mitochondrial apoptotic pathway. Compared with compound 2, compound 1 is more active as an antagonist of Src kinase, which might account for its higher potency to induce growth arrest and apoptosis. These results provide a deeper insight into the functions of these two simple anthraquinones and the anti-tumour pathway of Hedyotis diffusa WILLD.

抄録全体を表示

Buckwheat allergy is one of the most critical diseases manifested by severe and dangerous symptoms in Japan and other countries. We previously isolated the cDNA encoding protein BWp16, a member of the 2S albumin family with a conserved motif of 8 cysteine (Cys) residues. Comparison of the deduced amino acid sequences of BWp16 and related proteins in the 2S albumin family showed similarities between BWp16 and BW 8-kDa from buckwheat, Ara h 6 from peanuts and Ric c 1 from castor bean. Purified recombinant BWp16 (rBWp16) expressed in Escherichia coli was recognized by >80% of sera from patients with positive for IgE binding to buckwheat. Mutational analysis of rBWp16 revealed that 7 out of 10 mutants in the Cys residues showed weaker IgE binding to patient's serum than wild-type rBWp16 (rBWp16 WT). Mutations of Cys65 and Cys66 in rBWp16 decreased the pepsin digestibility of the protein, and an ELISA inhibition assay revealed a weaker inhibitory effect of rBWp16 C65S than that of rBWp16 WT. These results suggest that the Cys residues, especially Cys65, are involved in the allergenicity of rBWp16. Our findings provide new evidence for the role of Cys residues in 2S albumin family proteins and open the door to the production of hypoallergens and application to safe diagnostic methods and allergen-specific immunotherapy of buckwheat allergy.

抄録全体を表示

Several studies have shown that the interaction of CXC chemokine receptor 4 (CXCR4) with its ligand, stromal cell-derived factor-1α (SDF-1α) is closely involved in the directional migration of some tumors toward specific organs, which provides a new pathway against cancer metastasis. We previously developed an α-helix-defective mutant of SDF-1α, SDF-1/54 that displays obvious antagonistic activity to CXCR4. But it is necessary to ensure the targeting of SDF-1/54 to tumors in vivo since many normal tissues also express CXCR4. It is known that most tumor cells highly express epidermal growth factor receptor (EGFR). Meanwhile, decorin (DCN), a specific antagonist of EGFR, can target the tumor cells enriched in EGFR and cause a significant downregulation of EGFR. Hereby, we further generated a fusion construct of SDF-1/54 and DCN to expect to enhance the targeting of SDF-1/54 to tumors by dual blocking effects on CXCR4 and EGFR. This study focused on expression of recombinant chimera SDF-1/54-DCN in Escherichia coli, purification and bioactivity to inhibit the physiological functions mediated by CXCR4 and EGFR respectively in various tumor cell lines in vitro. Results indicated that SDF-1/54-DCN could inhibit both chemotaxis and proliferation of the tumor cells we used, which may be attributed to its blocking to CXCR4 and EGFR. These findings suggest that this strategy to link SDF-1/54 with DCN may be a promising approach to increase the targeting of SDF-1/54 to the tumors coexpressing CXCR4 and EGFR.

抄録全体を表示

A novel gene, factor for adipocyte differentiation 24 (fad24), promotes adipogenesis by controlling DNA replication early on during a stage referred to as mitotic clonal expansion (MCE). MCE is considered distinct from the proliferation of pre-confluent cells, so we investigated the role of fad24 in the process. First, the expression of fad24 was examined in pre-confluent and post-confluent 3T3-L1 preadipocytes, NIH-3T3 fibroblasts, and C2C12 myoblasts. fad24 was strongly expressed in the pre-confluent cells. The knockdown of fad24 by RNA interference impaired the ability of the pre-confluent cells to proliferate. Moreover, bromodeoxyuridine labeling and chromatin immunoprecipitation experiments indicated that the knockdown inhibited DNA synthesis by preventing the recruitment of histone acetyltransferase binding to ORC1 (HBO1), a component of the pre-replicative complex, to origins. fad24 plays positive roles in the proliferation of pre-confluent cells as well as adipogenesis.

抄録全体を表示

Purpose: L-Type amino acid transporter 1 (LAT1) is highly expressed in cancer cells to support their continuous growth and proliferation. We have examined the effect of 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH), an inhibitor of system L amino acid transporters, and the mechanism by which BCH suppresses cell growth in cancer cells. Methods: The effect of BCH and the mechanism of BCH on cell growth suppression in cancer cells were examined using amino acid transport measurement, MTT assay, DNA fragmentation analysis, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and immunoblotting. Results: BCH inhibited L-leucine transport in a concentration-dependent manner, and it inhibited cell growth in a time-dependent manner in KB human oral epidermoid carcinoma cells, Saos2 human osteogenic sarcoma cells and C6 rat glioma cells. The formation of a DNA ladder was observed, and the number of TUNEL-positive cells was increased with BCH treatment. Furthermore, the proteolytic processing of caspase-3 in KB and C6 cells and of caspase-7 in KB, Saos2 and C6 cells was increased by BCH treatment. Conclusion: These results suggest that the inhibition of LAT1 activity by BCH leads to apoptotic cancer cell death by inducing intracellular depletion of neutral amino acids necessary for cancer cell growth.

抄録全体を表示

The growth of Candida albicans in RPMI1640 medium was inhibited by sodium sulfite between pH 3—6. Under the condition of pH 7, growth of C. albicans was not inhibited by sodium sulfite. Under an acidic pH condition, sodium sulfite had a candidacidal effect and the activity was expressed within 150 min. The concentration of ATP in C. albicans was decreased by sodium sulfite. Ethanol production by C. albicans was inhibited by sodium sulfite at pH 5. These results indicated that the candidacidal effect of sodium sulfite under acidic conditions was caused by interruption of alcohol fermentation and aerobic respiration.

抄録全体を表示

The anti-metastatic activity of MD-Fraction extracted from the maitake mushroom (Grifola frondosa) was examined in an experimental murine model of lung metastasis. Intraperitoneal administration of MD-Fraction 2 d before tumor implantation significantly inhibited lung metastasis of colon-26 carcinoma and B16/BL6 melanoma cells. In this model, MD-Fraction enhanced IL-12 production from antigen presenting cells (APCs). MD-Fraction treatment activated NK cells and increased cytotoxicity against YAC-1 and colon-26 carcinoma cells. Furthermore, the depletion of NK cells with anti-asialo GM1 abolished the inhibitory effect of MD-Fraction on lung metastasis of colon-26 cells. Ex vivo, B16/BL6 cell adhesion to LPS-activated murine lung vascular endothelial cells was inhibited by MD-Fraction and anti-intercellular adhesion molecule (ICAM)-1 antibody. These results suggest that MD-Fraction inhibits tumor metastasis by activating NK cells and APCs, and by suppressing of ICAM-1 leading to the inhibition of tumor cell adhesion to vascular endothelial cells.

抄録全体を表示

The forced swimming test (FST) is suggested to produce abnormalities in the serotonergic and hypothalamic–pituitary–adrenal (HPA) axis systems. Therefore, compounds that attenuate these neurobiological alterations may have potential as antidepressants. The behavioral and biochemical effects of psoralen, a major furocoumarin isolated from Psoralea corylifolia, were investigated in the FST model of depression in male mice. Psoralen significantly reduced immobility and increased swimming without altering climbing in the mouse FST. Psoralen remarkably reversed FST-induced alterations in serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) levels in frontal cortex and hippocampus in mice. Furthermore, psoralen attenuated FST-induced elevations in serum corticotropin-releasing factor (CRF) and corticosterone concentrations to normalize the HPA axis activity. These results suggested that psoralen possessed potent antidepressant-like properties which were at least in part mediated by improving the abnormalities in the serotonergic and the HPA axis systems.

抄録全体を表示

Satratoxin H, a mycotoxin, is thought to induce apoptosis of PC12 cells through the activation of p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) in a glutathione (GSH)-sensitive manner. The present study was undertaken to further elucidate the mechanism by which satratoxin H induces cell death in PC12 cells. Satratoxin H caused apoptosis of PC12 cells within 24-h, as determined by DNA fragmentation and flow cytometric analysis. Satratoxin H increased reactive oxygen species (ROS) production and lipid peroxidation, as determined by malondialdehyde formation. These effects were attenuated by incubation of cells with GSH, suggesting that satratoxin H-induced increase in apoptosis of serum-deprived PC12 cells may be partially mediated through the generation of ROS.

抄録全体を表示

Microglia play important roles in the pathogenic cascade following cerebral ischemia, since they express growth factors, chemokines and regulatory cytokines as well as free radicals and other toxic mediators. P2X₇ receptor, a subtype of a family of P2 purinoceptors, is primarily expressed in microglia and macrophages, suggesting that it regulates immune function and inflammatory responses. However, the involvement of ATP in such microglial responses after cerebral ischemia is not yet understood. In this study, we investigated the possible involvement of ATP, especially through the P2X₇ receptors, in a rat model of focal cerebral ischemia. In immunohistochemical analysis, P2X₇ receptor-like immunoreactivity was predominantly detected in microglia, and then activated microglia accumulated in the ischemic region, in rats subjected to middle cerebral artery occlusion (MCAO) and reperfusion. Intracerebroventricular injection with P2X₇ receptor agonist 2′-3′-O-(4-benzoylbenzoyl)adenosine 5′-triphosphate (BzATP) improved behavioral dysfunction accessed by rota-rod test and ischemic neural injury induced by MCAO. In contrast, P2X₇ receptor antagonist adenosine 5′-triphosphate-2′,3′-dialdehyde (OxATP) exacerbated ischemic brain damage. These results suggest that microglia play an important role in neuroprotection against rat cerebral ischemia, which is regulated by a P2X₇ receptor-mediated ATP signal.

抄録全体を表示

Three derivatives of ouabain, digoxin and proscillaridin A containing the carboxylic group instead of the lactone moiety were synthesized and examined for cytotoxicity in human breast cancer cells. Evaluation of the cytotoxicity of these compounds employing an MTT assay and inhibition of [³H]thymidine incorporation into DNA in both MCF-7 and MDA-MB-231 breast cancer cells demonstrated that compound 3, the most active of the series, proved to be only slightly less potent than proscillaridin A. We evaluated the effects of these compounds 1—3 on change in intracellular Ca²⁺, appearance of apoptosis, inhibition of DNA topoisomerase I and II, and the activity of caspase-3 in breast cancer cells. These studies indicate that the increase in potency for 3 may be related, in part, to an activation of caspase-3, increasing free calcium concentration and topoisomerase II inhibition. All these data emphasize the potential usefulness of these derivatives of cardiac glycosides as anticancer agents.

抄録全体を表示

In the present study, we investigated the anti-inflammatory effects of a Kampo medicine Shosaikoto (TJ-9) using in vitro periodontal disease model, in which human gingival fibroblasts (HGFs) treated with lipopolysaccharide (LPS) from Porphyromonas gingivalis (PgLPS) produce IL-6, IL-8 and prostaglandin E₂ (PGE₂). Treatment with PgLPS (10 ng/ml), TJ-9 (up to 1 mg/ml) and their combinations for 24 h did not affect the viability of HGFs. Moreover, TJ-9 did not alter LPS-induced IL-6 and IL-8 productions. However, TJ-9 significantly suppressed LPS-induced PGE₂ production in a dose-dependent manner but TJ-9 alone did not affect basal PGE₂ level. Western blotting demonstrated that TJ-9 decreased cyclooxygenase-2 (COX-2) expression in a dose-dependent manner but not phospholipase A₂. Moreover, TJ-9 selectively and dose-dependently inhibited COX-2 activity. These results suggest that TJ-9 decreased PGE₂ production by inhibition of both COX-2 expression and activity and that TJ-9 may be useful to improve gingival inflammation in periodontal disease.

抄録全体を表示

Female hormone-dependent cancers and other diseases pose a serious health threat for women, and low-risk medicines against such cancers have not yet been discovered. The present study examines the effects of the traditional Chinese herbal mixture, Tokishakuyakusan (TS) and 17β-estradiol on the uterus of parous ovariectomized rats. Uterine atrophy that causes a reduction in uterine tissue and the uterine cavity area, was induced by ovariectomy, and slightly recovered by the daily oral administration of TS for two weeks (1000 mg/kg body weight). TS restored the decreased plasma estradiol concentration due to ovariectomy. However the yeast two-hybrid assay showed that TS did not bind estrogen receptors α and β and immunohistochemical staining revealed that 17β-estradiol stimulated the protein expression of estrogen receptor α, progesterone receptor, c-fos and c-jun in the uterus, whereas TS did not. These results suggest that TS might be useful for treating menopausal syndromes among women, as well as for patients when hormone replacement therapy (HRT) with estrogen is contraindicated.

抄録全体を表示

We investigated the skin metabolism of edaravone as a radical scavenger in Wistar and hairless rat skin. Approximately 1 g of abdominal skin was excised from 10-week-old Wistar and hairless rats, homogenized in 10 ml saline, and centrifuged at 10000 g for 20 min. The supernatant fluid was used for the examination of edaravone metabolism in the skin, and we also used supernatant fluid that was heated at 80 °C. Edaravone solution (0.05 ml, 2.4 μmol/ml) was added to 0.95 ml Wistar rat and hairless rat skin homogenate supernatant fluids. In Wistar rats, the residual amount of edaravone in skin homogenate supernatant fluid at 37 °C after 0, 5, 10, 20 and 30 min was 61.58±1.65, 41.84±8.52, 35.54±8.62, 19.73±5.99 and 13.89±4.40%, respectively. In hairless rats, the residual amount of edaravone in skin homogenate supernatant fluid at 37 °C after 0, 5 and 10 min was 50.19±14.17, 6.71±5.82 and 0.89±0.80%, respectively, and edaravone was not detected after 20 min. Although it was thought that metabolic enzyme activity in skin homogenate supernatant fluid was lost following heat treatment at 80 °C, the residual amount of edaravone in our skin homogenate supernatant fluid decreased with time. It is suggested that edaravone metabolism in the skin is necessary for non-enzymatic reactions.

抄録全体を表示

The antitumor activity of newly developed taxoids possessing a sugar moiety (glucose, galactose or mannose) to improve water solubility was evaluated. Galactose-bound taxoid (10-α-GAG-DT) proved superior to all other taxoids in both water solubility and antitumor activity. Therapeutic efficacy of 10-α-GAG-DT in terms of in vivo antitumor activity was found to be approximately equivalent to that of docetaxel, which is known to be superior to that of paclitaxel. The observation that the sugar moiety was gradually hydrolyzed in serum to release docetaxel indicates that 10-α-GAG-DT acts as a prodrug.

抄録全体を表示

We examined if tocilizumab, humanized anti-interleukin-6 receptor (IL-6R) antibody, can ameliorate joint swelling after the onset of arthritis in collagen-induced arthritis (CIA). CIA was induced by the immunization of bovine type II collagen in female cynomolgus monkeys. Tocilizumab (30 mg/kg) was administered weekly for 4 weeks after the onset of arthritis. Swelling of 16 proximal interphalangeal (PIP) joints of hands and feet was monitored by measuring the longitudinal and transverse axes of PIP joints and the oval area of each PIP was calculated. Serum was collected once a week after tocilizumab injection and blood chemistry, IL-6, soluble IL-6R (sIL-6R), and anti-tocilizumab antibody were measured. At the end of study, histopathological examination of joints was performed. Tocilizumab clearly reduced joint swelling in all animals without anti-tocilizumab antibody. Histopathological study showed significant decrease in the infiltration of neutrophils into inflamed joints was observed in tocilizumab-treated animals. In conclusion, tocilizumab improved established arthritis in monkey and monkey CIA model is useful for the analysis of anti-arthritic effect of tocilizumab.

抄録全体を表示

Several lines of evidence suggest that activation of spinal mitogen-activated protein kinases (MAPKs), including extracellular signal-regulated kinase (ERK) and p38 MAPK, contributes to the induction and maintenance of chronic pain. We recently reported that an intrathecal (i.t.) administration of ATP evoked tactile allodynia, which lasted more than 1 week in rats. The long-lasting allodynia was induced by activation of spinal P2X_2/3-receptors, and the induction and early phase of maintenance, but not the late phase, was mediated, at least in part, by the activation of spinal glial cells. In this study, we examined the involvement of spinal ERK and p38 MAPK in each phase of i.t. ATP-evoked long-lasting allodynia. I.t. administration of ATP (100 nmol) markedly increased phosphorylated ERK, which peaked at 1—8 h before gradually decreasing to a level that was sustained until 7 d after administration. In contrast, only a slight increase in phosphorylated p38 MAPK was observed. Consistent with the increased phosphorylation of MAPKs, the ERK kinase MEK inhibitor, U0126 (3 nmol), attenuated the induction phase (co-administration with ATP) and early maintenance phase (1-d post-ATP administration) of the i.t. ATP-evoked allodynia, but not the late maintenance phase (7-d post-ATP administration), while the p38 MAPK inhibitor, SB203580 (10 μg), had little effect. These results suggest that the induction phase and early maintenance phase, but not the late maintenance phase of long-lasting allodynia is mediated by the activation of ERK, rather than by the activation of p38 MAPK, in the spinal cord. These findings are informative for elucidating the mechanisms underlying the pathogenesis of chronic pain.

抄録全体を表示

Berberine is one of the main alkaloids of Rhizoma coptidis which has been used as a folk medicine to treat diabetes mellitus for more than 1400 years in China. To investigate the chronic effect of berberine on diabetic hyperlipidemic rats, fasted rats were intraperitoneally injected 35 mg/kg streptozotocin. Diabetic rats were admitted after 2 weeks and given a high-carbohydrate/high-fat diet to induce hyperlipidemia. The rats were divided into 7 groups at the end of week 16: normal and diabetic rats received no drug, 5 treatment groups were administered with either 75, 150, 300 mg/kg berberine, 100 mg/kg fenofibrate or 4 mg/kg rosiglitazone per day for 16 weeks, respectively. The blood glucose, hemoglobin A1c, lipid metabolic parameters and hepatic glycogen and triglyceride were measured, and histopathology and peroxisome proliferator-activated receptors (PPARs) α/δ/γ expression of liver were determined by hematoxylin eosin and immunohistochemical staining. Berberine reduced diabetic rats' body weight, liver weight and liver to body weight ratio. Berberine restored the increased blood glucose, hemoglobin A1c, total cholesterol, triglyceride, low density lipoprotein-cholesterol, apolipoprotein B and the decreased high density lipoprotein-cholesterol, apolipoprotein AI levels in diabetic rats to near the control ones. Berberine alleviated the pathological progression of liver and reverted the increased hepatic glycogen and triglyceride to near the control levels. Berberine increased PPARα/δ expression and reduced PPARγ expression in liver of diabetic rat to near the control ones. Berberine improved glucolipid metabolism both in blood and liver in diabetic rats possibly through modulating the metabolic related PPARα/δ/γ protein expression in liver.

抄録全体を表示

Overexpression of P-glycoprotein (Pgp) and multidrug resistance protein 1 (MRP1) by tumors results in multidrug resistance (MDR) to structurally unrelated anti-tumor agents. HZ08, a chiral compound, was a newly synthesized tetraisohydroquinoline derivative to reverse Pgp and MRP1 mediated MDR. In present studies, R, S-HZ08 and their racemate reversed the resistance to adriamycin and vincristine of adriamycin-selected human leukemia (K562/ADM) cells that overexpress Pgp. R, S-HZ08 and their racemate modulated adriamycin cytotoxicity when R, S-HZ08 and their racemate were removed 12 h prior to the cytotoxicity assay. In addition, R, S-HZ08 and their racemate increased intracellular accumulation of Rhodamine123 in Caco-2 cells that overexpress Pgp. Furthermore, using a DNA content analysis and an annexin V binding assay, R, S-HZ08 and their racemate effectively reversed the resistance to adriamycin-induced apoptosis in K562/ADM cells. R, S-HZ08 and their racemate also moderately reversed the resistance to adriamycin and vincristine of MCF-7/ADM cells that overexpress MRP1. However, R, S-HZ08 and their racemate hardly affected intracellular glutathione (GSH) levels and glutathione S-transferase (GST) activities in MCF-7/ADM cells. The result showed that R, S-HZ08 and their racemate possibly reverse MDR1 mediated multidrug resistance by a direct interaction with MRP1, not interaction with MRP1 via GSH. Thus, R, S-HZ08 and their racemate should be useful for treating patients with tumors that overexpress both Pgp and MRP1.

抄録全体を表示

8-Hydroxy-2′-deoxyguanosine (8-OHdG), one of the most abundant oxidative DNA adducts, is used as an indicator of oxidative DNA damage associated with aging. Among homologs of the silent information regulator (Sir), sirtuin 1 (SIRT1) is suggested as a regulator of the apoptotic response to DNA damage. Since it has been suggested that the aging process can be delayed by the attenuation of oxidative damage such as DNA damage or SIRT1 modulation, we focused on the protective effect against cellular oxidative damage of persimmon peel, a proanthocyanidin-rich food, in relation to its level of polymerization. We confirmed that 8-OHdG expression in TIG-1 human fibroblasts was increased by treatment with 300 μM H₂O₂ for 2 h. On the other hand, the nuclear SIRT1 level was decreased in H₂O₂-treated as compared with non-pretreated cells. However, pretreatments with polymers and oligomers led to a decrease in 8-OHdG and elevation in nuclear SIRT1 expression in a concentration-dependent manner. In particular, oligomers exerted a stronger effect. The present study supports the protective potential of proanthocyanidin from persimmon peel against oxidative damage under the aging process, and suggests that the polymerization of proanthocyanidin plays an important role in retarding aging in a cellular senescence model.

抄録全体を表示

The possibility of vitamin K₃ (VK₃) as an anticancer agent was assessed. VK₃ dose-dependently diminished the cell viability (measured as esterase activity) with IC₅₀ of 13.7 μM and Hill coefficient of 3.1 in Hep G2 cells. It also decreased the population of S phase and arrested cell cycle in the G₂/M phase in a dose-dependent manner. G₂/M arrest was regulated by the increment of cyclin A/cdk1 and cyclin A/cdk2 complex, and contrasting cyclin B/cdk1 complex decrease. Finally, combined application demonstrated that VK₃ significantly enhanced the cytotoxicity of etoposide, a G₂ phase-dependent anticancer agent, whereas it reduced the cytotoxic activity of irinotecan, a S phase-dependent agent. These findings suggest that VK₃ induces G₂/M arrest by inhibition of cyclin B/cdk1 complex formation, and is thus useful as an enhancer of G₂ phase-dependent drugs in hepatic cancer chemotherapy.

抄録全体を表示

FTY720 (fingolimod), a novel immunosuppressant, was found to become biologically activated by phosphorylation into FTY720-1-phosphate (FTY720-P), which is a high-affinity agonist for sphingosine-1-phosphate (sphingosine-1-P)-receptors. FTY720 has also been reported to have a strong antitumor activity. The association between the phosphorylation of FTY720 and the growth inhibition of FTY720 against cancer cells are still not completely understood. In this study, we investigated the effects of FTY720, sphingosine, and their related compounds on the proliferation of human breast cancer cell lines (MCF-7, MDA-MB-231 and Sk-Br-3) and human colon cancer cell lines (HCT-116 and SW620). Non-phosphorylated FTY720, sphingosine and an FTY720 derivative, ISP-I-55, showed significant growth inhibition against these cells, with IC₅₀ values of 5—20 μM at 48 h post-drug treatment. We confirmed that FTY720 induces the activation of a major mitogen-activated protein kinase, JNK, without the activation of p38 and down-regulation of phospho-ERK in MCF-7 breast cancer cells. In contrast, the phosphorylated derivatives, FTY720-P and sphingosine-1-P, as well as a phosphinane FTY720 derivative, cFTY720-P, did not inhibit the growth of the cells in the concentration range of 5—50 μM, whereas FTY720-P and sphingosine-1-P slightly induced the growth of MCF-7 cells. Combining FTY720 with dimethylsphingosine, a sphingosine kinase inhibitor, augmented the inhibitory effect of FTY720. These results indicate that the antiproliferative activity of FTY720 does not result from its phosphorylation, either endogenous or exogenous.

抄録全体を表示

The suitability of an ¹⁸F-labeled form of N-(piperidin-1-yl)-1-(2,4-dichlorophenyl)-5-{4′-(5-fluoropentyl)phenyl}-4-methyl-1H-pyrazole-3-carboxamide (1), a CB1 cannabinoid ligand with high binding affinity (K_i=0.91 nM) and moderate lipophilicity (log P_7.4=2.9), as a radiotracer for positron emission tomography imaging was evaluated in mice. Ligand 1 was labeled with ¹⁸F (T_1/2=109.7 min) by treatment of the corresponding tosyl derivative with [¹⁸F]fluoride ion in acetonitrile. Tissue distribution studies of the ¹⁸F-labeled form ([¹⁸F]1) in mice demonstrated low brain uptake with minimal specific binding in brain regions. Cyclosporin A ( a modulator of P-glycoprotein) treatment significantly increased both the brain uptake and the brain-to-blood ratio of [¹⁸F]1, indicating the possibility that P-glycoprotein regulates the ability of [¹⁸F]1 to cross the blood brain barrier. Radioligand [¹⁸F]1 does not have the required properties for imaging the cerebral cannabinoid CB1 receptor in vivo.

抄録全体を表示

Numerous studies have demonstrated the participation of glicolipids in signal transduction and the regulation of melanoma cell growth and apoptosis. Hoping to discover new anticancer drugs, we have synthesized ten glycolipids found in various invertebrates that do not have sialic acids. These compounds were tested for antiproliferative effects on a melanoma cell line, B16F10. A synthetic compound, Manβ(1–4)[Fucα(1–3)]Glcβ1-Cer, (glycosphingolipid 7), which was identified in the millipede Parafontaria laminata armigera, had an antiproliferative effect on the melanoma cells. This compound suppressed the activation of the focal adhesion kinase (FAK)-Akt pathway as well as the activation of extracellular signal-regulated kinase (Erk)1/2 pathway involved in cell proliferation. Expression of the cell cycle proteins, cyclin D1 and CDK4, was suppressed by glycosphingolipid 7. From these results, glycosphingolipid 7 suppressed the activation of the FAK-Akt pathway and of Erk1/2, which resulted in a decrease in the expression of cyclin D1 and CDK4. Glycosphingolipid 7 might be a candidate for an inhibitor of cell proliferation in melanomas.

抄録全体を表示

Cytochrome P450 lanosterol 14-α demethylase (CYP51), participated in keeping serum cholesterol homeostasis, is a key enzyme to synthesize cholesterol from lanosterol. Here we focused on investigating the mechanism of CYP51 in modulating serum cholesterol levels in mouse through RNA interference (RNAi). Mice fed on normal or high fat high cholesterol (HFHC) diets were individually treated with small interference RNA (siRNA) of CYP51 gene by tail vein injection. The results showed that administrated single dose of 10 μg CYP51-siRNAs for 48 h resulted in significantly depletion of CYP51 mRNA in liver of mice fed on normal diet (from 40 to 60%, p<0.05). CYP51-siRNAs exerted the inhibition in a dose dependent manner (from 26% in 5 μg to 40% in 20 μg, p<0.05) and most inhibitive effect from day 3 to day 6 (over 50%, p<0.05) after the treatment. Six days after administration of 30 μg CYP51-siRNAs (20 μg on day 0 and 10 μg on day 3), CYP51 mRNA (normal: 50%; HFHC: 70%, p<0.05) and protein levels (normal and HFHC: over 40%, p<0.05) were significantly knocked down in mice liver. Interestingly, low-density lipoprotein receptor (LDLR) expression was significantly elevated compared with controls in hepatic cells after CYP51-siRNAs (mRNA: about 2 times; protein: about 1.6 times, p<0.05). As a consequence, about 50% of sera low-density lipoprotein cholesterol (LDL-ch) were significantly reduced (p<0.05). The effect on LDLR increase and LDL-ch reduction lasted 8 d after a single 20 μg CYP51-siRNAs injection. In addition, CYP51-siRNAs could not cause any fatty liver compared with Buffer-group and did not interfere with mice ovulation. In conclusion, these data demonstrated that CYP51-siRNAs silenced CYP51 in mouse liver and down-regulated plasma LDL-ch levels. The potential mechanism of LDL-ch reduction may be related to up-regulated LDLR expression of hepatic cells. It indicated that there was a cholesterol levels link-modulation system between cholesterol synthetic pathway through CYP51 and cholesterol transport pathway through LDLR in vivo.

抄録全体を表示

A vegetarian diet rich in phytochemicals may prevent colon carcinogenesis by affecting biochemical processes in the colonic mucosa. Compounds passing the digestive system reaching the colon could potentially be detected in fecal water. We previously reported that intact fecal water samples from human volunteers significantly decreased prostaglandin production and COX-2 protein expression in colonic cells. The aim with the present study was to further study the composition of the fecal waters, using NMR spectroscopy and multivariate data analysis, and to trace the COX-2 inhibiting activity. Intact fecal water samples and fractions thereof were analyzed for their ability to inhibit prostaglandin E₂ production in the human colon cell line HT-29. The majority of the tested aqueous phases derived from intact fecal water showed ability to inhibit prostaglandin production in cells (13.8±1.34% inhibition, p=0.01). NMR analysis indicated the presence of significant quantities of amino acids and fatty acids. Major metabolites included; acetic acid, butanoic acid, propanoic acid, glutamic acid and alanine. Smaller amounts of glycine and fumaric acid, which are known to have anti-inflammatory and anti-tumorigenic properties, were also detected. This study describes for the first time NMR metabolomic analysis of fecal water from subjects on a vegetarian diet.

抄録全体を表示

Extract of the whole plant, Ajuga decumbens (KE) has long been used in China as a medication for the relief of joint pain. Previously, we proved that KE up-regulated the synthesis of collagen in false aged model rats. In this paper we examined the effects of KE on nitric oxide (NO) production, expression of inducible nitric oxide synthase (iNOS), osteoblast and osteoclast activity. We also investigated whether KE had any anti-osteoporosis or anti-arthritic activity by using ovariectmized mice and adjuvant induced arthritic rats. KE exhibited down-regulation of differentiation into osteoclast and up-regulation of mineralization in osteoblast-like MC3T3-E1 cells in a concentration-dependent manner. NO synthesized by iNOS plays important roles in inflammatory disease and imbalance between bone resorption and bone formation caused by estrogen depletion. KE inhibited expression of iNOS which caused concentration dependent inhibition of NO production. Furthermore, KE prevented brittle bones in ovariectomized mice and swelling of the left hind ankle in adjuvant induced arthritic rats. Therefore, KE improved the balance of bone resorption and bone formation, showing anti-inflammatory effects. Consequently, KE is beneficial for sufferers of bone and joint disease.

抄録全体を表示

The effects of pantothenic acid-supplementation on the adrenal secretion of corticosterone and progesterone in male rats were investigated using an in vitro cell culture system. Male rats at 21 d of age were given 0.03% pantothenic acid in their drinking water for 9 weeks. After 9 weeks of treatment, the animals were decapitated, and adrenal cells were cultured in the absence or presence of rat adrenocorticotropic hormone (ACTH; 10⁻¹⁵ to 10⁻¹⁰ M) and/or ovine prolactin (oPRL; 10⁻⁹ to 10⁻⁷ M) for 4 h. Adrenal cells in pantothenic acid-treated rats exhibited higher basal levels of corticosterone and progesterone than control rats. The response of ACTH and/or PRL on corticosterone and progesterone release was higher in the pantothenic acid-treated rats than in the control rats. In addition, PRL increased the stimulatory effect of ACTH-induced corticosterone secretion in both normal and pantothenic acid-treated rats. These results clearly demonstrated that pantothenic acid supplementation stimulates the ability of adrenal cells in male rats to secrete corticosterone and progesterone. Additionally, these results also showed that pantothenic acid supplementation induced adrenal hyperresponsiveness to ACTH stimulation, and PRL further stimulated adrenal sensitivity to ACTH.

抄録全体を表示

Atazanavir (ATV) is clinically coadministered with low-dose ritonavir (RTV), which boosts the oral bioavailability (BA) of ATV by inhibiting cytochrome P450 (CYP) 3A, and P-glycoprotein (Pgp) via the same metabolic pathway; however, it is well known that in the chronic phase, the inhibition effect of RTV on Pgp and CYP3A becomes an induction effect. In this study, we investigated the long-term efficacy and safety of RTV-boosted ATV in rats with a clinical relevant dosage of ATV and RTV, 7 mg/kg and 2 mg/kg, respectively, and drew a direct comparison with RTV-boosted ATV and the previously reported ATV pharmaceutical formulation based on a solid dispersion system (ATV-SLS SD+G). Rats received RTV-boosted ATV or ATV-SLS SD+G for 14 d in the pharmacokinetic study. In addition, after 14-d repeated administration of each formulation, cyclosporine A (CyA) was administered to rats and Western blot analysis of Pgp and CYP3A was performed to investigate the impact on pharmacokinetic interaction of each ATV formulation. After repeated administration of both formulations, there was no significant difference between ATV pharmacokinetic parameters on day 1 and 14; therefore, it was considered that the long-term efficacy of both ATV formulations was maintained. However, after treatment with RTV-boosted ATV, the C_max and AUC_0—∞ of the following CyA significantly decreased to 49% and 47% in comparison to the control, respectively, and the Pgp expression in the small intestine by Western blot analysis was approximately 2-fold higher than the control, whereas after treatment with ATV pharmaceutical formulation, neither significant alteration of CyA nor notable change in the expression of intestinal Pgp and hepatic CYP3A was observed. Therefore, it was considered that the BA of CyA after treatment with RTV-boosted ATV would decrease by the induction effect of RTV in chronic phase as described above. The results of this study revealed that the chronic use of low-dose RTV as a booster has great potential to compromise drug–drug interactions; therefore, it is recommended that the BA of protease inhibitors be improved by a pharmaceutical approach without pharmacokinetic interaction by RTV.

抄録全体を表示

Many pharmacokinetic studies of paclitaxel formulations with or without Cremophor (CrEL) have been performed on experimental animals. However, limited studies describe the different pharmacokinetic behaviors of paclitaxel in animals. The different distribution of drug in blood fractions may have great effect on its pharmacokinetic behaviors. Our present study was designed to study the characteristics of paclitaxel distribution in human, rabbit and rat blood, by measuring plasma-to-blood ratio (PBR) of paclitaxel in vitro and in vivo, and analyzing the results of equilibrium dialysis of paclitaxel with erythrocyte, plasma and hemoglobin. It was demonstrated that the paclitaxel PBR values in rat, unlike those in rabbit, are most significantly different from those in human, which may be due to distinct affinity of paclitaxel to blood fractions among different species. The effect of CrEL on increasing paclitaxel plasma concentration and in vitro & in vivo correlation in animal PBR values were observed. The findings in this study are of significance in the evaluation of the newly developed formulations of paclitaxel.

抄録全体を表示

Lipopolysaccharide (LPS) is a highly bioactive substance that can cause local as well as systemic damage to various organs of both humans and animals, even at very low doses. However, there are a few reports on drug pharmacokinetics during endotoxemia. In this study, we analyzed the pharmacokinetics of digoxin (a therapeutic agent for cardiac insufficiency) as a probe drug for a two-compartment model in a rat model of endotoxemia induced by LPS for 5 d. Digoxin was given to Wistar rats intravenously (i.v.), orally (p.o.), and intra-intestinally using an in situ closed-loop method (loop). The AUC_i.v. was significantly increased in the LPS (+) group throughout the experiment (p<0.05). There was significant decrease in V₂ (volume of distribution of tissue compartment) on Day 1—3 (p<0.05). On Day 1—2 after LPS administration, the AUC_p.o. was significantly increased in the LPS (+) group (p<0.05). The AUC_loop was significantly increased throughout the experiment (p<0.05). The elimination rate constant was unchanged. Thus LPS administration affected the absorption but not the excretion of digoxin. The findings of this study suggest that digoxin absorption increased and the volume of distribution of tissue compartment decreased after LPS administration (5 mg/kg, i.p.). It appears that digoxin pharmacokinetics recover over 3 d after LPS administration.

抄録全体を表示

Deoxyribonucleic acid (DNA) from bacteria or viruses has been reported as one of the pathogen-associated molecular patterns (PAMPs) and a substance that can induce endotoxemia-like inflammation in animals. However, there has been no report on digoxin pharmacokinetics in the inflammation induced by bacterial DNA containing unmethylated CpG motifs (CpG-DNA). In this study, we investigated the effects of CpG-DNA on digoxin pharmacokinetics. We determined the degree of lipopolysaccharide contamination in CpG-DNA solution and examined the changes in digoxin pharmacokinetics in rats after CpG-DNA administration. In addition, plasma concentrations of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and nitrite/nitrate (NOx) were determined after CpG-DNA administration (5 mg/kg, i.p.). The AUC_0—24 of digoxin increased significantly on Day 1—3 and CL/F decreased on Day 1 and Day 2 after CpG-DNA administration. On Day 7 after CpG-DNA administration, there were no significant differences in AUC_0—24 and CL/F compared with the control group (without CpG-DNA administration). However, Kel remained relatively unchanged throughout the experiment. Plasma TNF-α concentrations were significantly increased at 1 h and plasma IL-1β concentrations were significantly decreased at 6 h after administration of CpG-DNA, while plasma NOx concentrations were significantly increased at 12 h after CpG-DNA administration, compared with the control group. These findings suggest that CpG-DNA (5 mg/kg) induces a transient inflammatory condition, and that AUC_0—24 and CL/F of digoxin were altered after CpG-DNA administration. Digoxin pharmacokinetics recovered within 7 d after CpG-DNA exposure.

抄録全体を表示

The tissue distribution and disposition of carnitine, which plays an important role in the transport of long-chain fatty acids across the mitochondrial inner membrane for β-oxidation, are well controlled by carnitine transporter organic cation/carnitine transporter 2 (OCTN2). Since little information is available on regulation of the expression of the OCTN2 gene, we examined the factors that affect the expression level of rat Octn2 (rOctn2), focusing on nuclear receptor peroxisome proliferator activated receptor alpha (PPARα), which regulates expression of genes associated with β-oxidation of fatty acids. mRNA of rOctn2 was induced by the PPARα ligand fenofibrate in primary-cultured rat hepatocytes. Further, the PPARα ligand Wy14643 increased the expression of Octn2 in wild-type mice, but not in PPARα knockout mice. Analysis of the rOctn2 promoter region suggested the presence of putative cis elements of PPARα. Wistar rats treated with intraperitoneal fenofibrate administration showed increased expression of rOctn2 mRNA in liver, and uptake of [³H]carnitine by freshly isolated hepatocytes derived from those rats was also increased. In conclusion, our results indicate that the nuclear receptor PPARα directly up-regulates the expression of rOctn2 and increases the hepatic uptake of carnitine via rOctn2.

抄録全体を表示

Non-viral DNA vectors are promising gene delivery systems and a variety of non-viral DNA vectors have been developed to date. Recently, we developed a novel non-viral gene delivery system—multifunctional envelope-type nano device (MEND). The MEND system has high transfection activity, similar to that of adenovirus vector, which is a potent viral vector. However, conventional MEND is relatively large and heterogeneous (approximately 300 nm), probably because they contain relatively large- and heterogeneous-pDNA particles condensed with polycations, such as poly-L-lysine. Small particle size is important for in vivo delivery, because large particles are rapidly eliminated from systemic circulation. Moreover, heterogeneous size of drug carriers is difficult to apply to clinical applications. Here, we describe construction of small homogeneous MEND. First, we screened mono-cationic detergents (MCD(s)) to obtain optimal pDNA condensed particles. We determined that benzyldimethylhexadecylammonium chloride (BDHAC) and thonzonium bromide (TB) were optimal pDNA condensers. Next, we packaged the condensed pDNA particles into a lipid bi-layer. The resulting lipid-encapsulated pDNA particles were then equipped with octaarginine to facilitate cell-uptake (R8-MEND (MCD)). The carrier showed high transfection activity in cultured HeLa cells. Furthermore, the R8-MEND (MCD) were small and homogeneous compared with conventional MEND. These results indicate that R8-MEND (MCD) has potential as a novel non-viral delivery system for clinical application.

抄録全体を表示

To develop a piroxicam-loaded gelatin microcapsule with enhanced bioavailability, a gelatin microcapsule encapsulated ethanol and piroxicam has been formulated by using gelatin as a water-soluble polymer shell. The aqueous solubility and bioavailability of piroxicam in piroxicam-loaded microcapsule in rats were then evaluated compared to piroxicam powder. The piroxicam-loaded gelatin microcapsule spherical in shape with smooth surface showed the geometric mean diameter of about 19 μm. It had the piroxicam solubility of about 1.87 mg/ml and the amount of ethanol of about 4.37 μg/mg. Furthermore, it gave significantly higher total plasma concentrations, C_max and area under the blood concentration–time curve (AUC) of piroxicam in rats than did piroxicam powder, indicating that the drug from gelatin microcapsule could be more orally absorbed in rats. In particular, the AUC of piroxicam in gelatin microcapsule was significantly about 2 fold increased compared to piroxicam powder. This enhanced oral relative bioavailability of piroxicam in gelatin microcapsule was contributed by the marked increase in the absorption rate of piroxicam due to the improved solubility of piroxicam. Thus, the piroxicam-loaded gelatin microcapsule developed using spray-drying technique with gelatin, sodium lauryl sulfate and ethanol would be useful to deliver piroxicam in a pattern that allows fast absorption in the initial phase, leading to better absorption.

抄録全体を表示

In this study erythrocyte transport of prostaglandin E₁ (PGE₁) was investigated by employing inside-out membrane vesicles prepared from rat erythrocytes. The uptake of [³H]PGE₁ in the presence of ATP was significantly higher than that of AMP, suggesting the involvement of an ATP-dependent efflux system in PGE₁ transport across the erythrocyte membrane. Coincubation of glutathione with ATP further stimulated the uptake of [³H]PGE₁. The uptake of [³H]PGE₁ in the presence of ATP and glutathione was temperature-sensitive, and various eicosanoids including PGE₂ and PGF_2α decreased the uptake. Multidrug resistance-associated protein (MRP) 4 substrates/inhibitors including methotrexate, indomethacin, taurocholic acid and indocyanine green significantly inhibited [³H]PGE₁ uptake. Western blot analysis revealed that Mrp4 is expressed in rat erythrocyte membrane. These results suggest that the release of PGE₁ from the erythrocyte into the blood circulation may be mediated by ATP-dependent efflux pump(s) such as Mrp4.

抄録全体を表示

The aim of this study was to evaluate the plasma trough concentrations (C₀) of mycophenolic acid (MPA) and its major metabolite MPA 7-O-glucuronide (MPAG) in metal cation (MC)(−) (non-treated) and MC(+) (co-treated) patients who received tacrolimus (Tac) or cyclosporine (CyA). Fifty-nine Japanese stable kidney transplant recipients receiving immunosuppressive regimens containing mycophenolate mofetil (MMF) and a calcineurin inhibitor (CNI) were included in this study. Seven in the 25 patients receiving Tac and 8 in the 34 patients receiving CyA were treated with concomitant MCs administration. Multiple regression analysis revealed that concomitant MCs and CyA administration influenced MPA C₀. Their standardized partial regression coefficients were −0.29 and −0.41, respectively. Stratified analysis based on CNI treatment revealed that MPA C₀ decreased significantly by 56% with concomitant MCs administration in Tac-treated patients. There was no significant difference in MPA C₀ between the MC(−) and MC(+) groups in CyA-treated patients. With respect to MPAG C₀, MC(+) group tended to be lower by 26% than MC(−) group in Tac-treated patients. There was no significant difference in MPAG C₀ between the MC(−) and MC(+) groups in CyA-treated patients. Concomitant MCs administration did not affect the C₀ ratio of MPAG to MPA in either Tac- or CyA-treated patients. In conclusion, MCs co-administration decrease MPA C₀ in patients receiving Tac and may cause lower MPA exposure. There are little pharmacokinetic interactions between MMF and concomitant MCs in CyA-treated patients.

抄録全体を表示

The aim of the present study was to investigate the mechanism for the stereoselective presystemic clearance of carvedilol. We examined the oxidation and glucuronidation of carvedilol in human liver microsomes (HLM) and human intestinal microsomes (HIM). The oxidation of carvedilol in HLM and HIM was evaluated in the presence of NADPH, whereas glucuronidation was evaluated in the presence of UDP-glucuronic acid. Oxidation of S-carvedilol in HLM and HIM was greater than that of R-carvedilol. In addition, the oxidation of R-carvedilol in HLM was inhibited by quinidine, whereas that of S-carvedilol was inhibited by both quinidine and furafylline. On the other hand, R- and S-carvedilol oxidation in HIM was inhibited by ketoconazole. Glucuronidation of S-carvedilol in HLM and HIM was also higher than that of R-carvedilol. These results suggested that cytochrome P450 (CYP) 2D6 and CYP1A2 are involved in the stereoselective oxidation of carvedilol in the liver, that CYP3A4 is involved in intestinal oxidation, and that glucuronidation in the liver and intestine is at least partly responsible for stereoselective presystemic clearance.

抄録全体を表示

Nucleosome is the major autoantigen in systemic lupus erythematosus (SLE). Professional antigen-presenting cells (APCs), such as macrophages (MΦs) and dendritic cells (DCs), play the central roles in the acquisition of Ag-specific immune responses and activation of such APCs is required for the efficient Ag-presentation. Therefore, adjuvant activity of DNA in nucleosomes would cause the prominent effects on the production of anti-nucleosome antibodies. In this study, we report that elicited peritoneal MΦs from New Zealand Black/White F₁ (NZB/W) mice showed a unique cytokine production profile following stimulation with DNA. MΦs from 5-week old NZB/W mice produced a higher amount of IL-6 and about a half amount of TNF-α after stimulation with DNA complexed with cationic liposomes compared with those from control ICR mice. These results suggest that MΦs of NZB/W mice have altered responsiveness to DNA and this might elevate the antigenicity of nucleosomes to induce the production of anti-nucleosome antibodies.

抄録全体を表示

The quality of microparticle enzyme immunoassay (MEIA) for blood tacrolimus is guaranteed in samples with hematocrit (Ht) values of 25 to 45%. Because MEIA provides inaccurate blood tacrolimus concentrations in samples with Ht out of this range (i.e. <25% or >45%), correction of the calibration is required for therapeutic drug monitoring. The authors demonstrated previously that overestimated MEIA tacrolimus concentration could be corrected by modified, calibrated MEIA (cMEIA) using the original calibrator. Here, an equation was established to more easily derive a corrected tacrolimus concentration by calculation (MEIAcalc) using the Ht of each sample. The tacrolimus concentrations of 99 whole-blood samples with low Ht (<25%) were then tested by the 3 assay methods: MEIA, cMEIA, and MEIAcalc. MEIA gave a significantly higher blood concentration of tacrolimus (median 12.9 ng/ml, range 3.6—26.4 ng/ml) than did cMEIA (median 10.0 ng/ml, range 0.2—21.1 ng/ml, p<0.05). This overestimation was eliminated by using MEIAcalc. There was no difference in blood tacrolimus concentration between cMEIA and MEIAcalc (median 10.0 ng/ml, range 1.7—21.4 ng/ml). MEIAcalc can be used to correct the tacrolimus concentration in samples obtained from patients with unstable Ht values.

抄録全体を表示