Biomedical subjects

H Ohtani

Publications and source records attributed to H Ohtani.

At least 73 records · Page 4Linked to original sources

PubMed2000

Human placental transport of vinblastine, vincristine, digoxin and progesterone: contribution of P-glycoprotein.

To elucidate the role of P-glycoprotein in human placenta, we examined its expression in placenta, and the transcellular transport and uptake of P-glycoprotein substrates in cultured human placental choriocarcinoma epithelial cells (BeWo cells). The uptake of [(3)H]vinblastine and [(3)H]vincristine into BeWo cells was increased in the presence of a metabolic inhibitor, sodium azide. The basolateral-to-apical transcellular transport of [(3)H]vinblastine, [(3)H]vincristine and [(3)H]digoxin was greater than the apical-to-basolateral transcellular transport. In the presence of cyclosporin A, the basolateral-to-apical transcellular transport of [(3)H]vinblastine, [(3)H]vincristine and [(3)H]digoxin was significantly increased, and the apical-to-basolateral transcellular transport was decreased. The uptake of [(3)H]vinblastine, [(3)H]vincristine and [(3)H]digoxin into BeWo cells was significantly enhanced in the presence of several inhibitors, such as verapamil or mouse monoclonal antibody anti-P-glycoprotein MX-MDR (MRK16) as well as cyclosporin A. Although progesterone significantly enhanced the uptake of [(3)H]vinblastine, [(3)H]vincristine and [(3)H]digoxin into BeWo cells, the uptake of [(3)H]progesterone was not affected by these inhibitors. Immunoblot analysis revealed that P-glycoprotein with a molecular weight of 172 kDa was expressed in BeWo cells and isolated trophoblast cells. Furthermore, P-glycoprotein was detected in human placental brush-border membrane vesicles, but not in human placental basolateral membrane vesicles. In conclusion, these data suggest that P-glycoprotein is expressed on the brush-border membrane (maternal side) of human placental trophoblast cells. P-Glycoprotein is considered to regulate the transfer of several substances including vinblastine, vincristine and digoxin from mother to fetus, and to protect the fetus from toxic substances.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed2000

Inhibition of P-glycoprotein by orange juice components, polymethoxyflavones in adriamycin-resistant human myelogenous leukemia (K562/ADM) cells.

We investigated the effects of the ethyl acetate extract of grapefruit juice (GFJ), that of orange juice (OJ) and their components on the uptake of [(3)H]vincristine into adriamycin-resistant human myelogenous leukemia cells. Its uptake was increased by the extracts of GFJ and OJ up to 7- and 3-fold, respectively, as well as verapamil and cyclosporin A. OJ components, i.e. 3,3',4',5,6,7,8-heptamethoxyflavone, nobiletin and tangeretin, also increased the uptake of [(3)H]vincristine in a concentration-dependent manner. Although GFJ components, dihydroxybergamottin and bergamottin, significantly increased the uptake, their potencies were considerably weaker than those of OJ components. These data suggest that OJ components inhibit P-gp-mediated efflux of [(3)H]vincristine, resulting in the intracellular accumulation of chemotherapeutic drugs. These components may become candidates of multi-drug resistance reversing agents in cancer chemotherapy.

ATP Binding Cassette Transporter, Subfamily B↗

PubMed2000

Molecular identification of a rat novel organic anion transporter moat1, which transports prostaglandin D(2), leukotriene C(4), and taurocholate.

We have isolated a rat novel multispecific organic anion transporter, moat1. The isolated clones were originated by alternative splicing of the moat1 mRNA. The nucleotide sequences predict a protein of 682 amino acids with moderate sequence similarity to LST-1, the oatp family, and the prostaglandin transporter. Northern blot analysis of rat moat1 identified a predominant transcript of 4.4 kilonucleotides in all tissues. Northern blot and in situ hybridization analyses of rat brain further indicated that moat1 mRNA is widely distributed in neuronal cells of the central nervous system, especially in the hippocampus and cerebellum. moat1 transports prostaglandin D(2) (K(m); 35.5 nM), leukotriene C(4) (K(m); 3.2 microM) and taurocholate (K(m); 17.6 microM) in a sodium-independent manner. moat1 also transports prostaglandin E(1), E(2), thromboxane B(2), and iloprost but not dehydroepiandrosterone sulfate and digoxin, of which the substrate specificity is similar, but definitively different from those of any other organic anion transporters.

Alternative Splicing↗

PubMed2000

Synchronous multiple primary gastrointestinal cancer exhibits frequent microsatellite instability.

Colorectal (CRC) and gastric cancers (GC), the most common gastrointestinal malignancies, have been known to develop occasionally in a same patient. Previous studies have focused on the etiology of patients with multiple primary gastric and colorectal cancer (MPGCC); however, the carcinogenic process of MPGCC remains unclear. In this study, we have examined the genetic alterations in MPGCC in order to clarify the carcinogenic pathway. Twenty patients with sporadic MPGCC were examined for microsatellite instability (MSI) and frameshift mutations of target genes such as TGFbetaRII, BAX and IGFIIR. In 10 (50%) of 20 patients with MPGCC, MSI was present at least at 1 lesion of GC or CRC. Four (50%) of 8 cases with synchronous MPGCC displayed MSI in both GC and CRC, while only 1 (8%) of 12 cases of metachronous MPGCC exhibited MSI in both organs. Carcinogenic process of MPGCC was fairly associated with the MSI pathway, particularly in cases of synchronous MPGCC. MSI was found in 5 (25%) of 20 GCs and in 10 (50%) of 20 CRCs. MSI was involved more closely in CRC than in GC among MPGCC. Although most frameshift mutations at target genes were found in the MSI-positive MPGCC, infrequent mutations were observed in the genes. Frameshift mutation was found in only 1 of 5 cases of MSI-positive GC at TGFbetaRII. Only 2 of 10 cases of CRC with MSI showed mutation at TGFbetaRII, and 1 case also showed mutation at BAX and IGFIIR. Our findings suggest that TGFbetaRII, BAX and IGFIIR are not the main target genes for carcinogenesis in MSI-positive MPGCC.

Aged↗

PubMed2000

Effect of bioflavonoids on vincristine transport across blood-brain barrier.

Several grapefruit juice bioflavonoids, including quercetin, are reported to stimulate P-glycoprotein-mediated drug efflux from cultured tumor cells. To see whether these bioflavonoids alter the permeation of vincristine across the blood-brain barrier, we conducted experiments with cultured mouse brain capillary endothelial cells (MBEC4 cells) in vitro and ddY mice in vivo. The steady-state uptake of [3H]vincristine by MBEC4 cells was decreased by 10 microM quercetin, but increased by 50 microM quercetin. Similarly, the in vivo brain-to-plasma concentration ratio of [3H]vincristine in ddY mice was decreased by coadministration of 0.1 mg/kg quercetin, but increased by 1.0 mg/kg quercetin. Kaempferol had a similar biphasic effect on the in vitro uptake of [3H]vincristine. Other aglycones tested (chrysin, flavon, hesperetin, naringenin) increased [3H]vincristine uptake in the 10-50 microM range, and glycosides (hesperidin, naringin, rutin) were without effect. We then addressed the mechanism of the concentration-dependent biphasic action of quercetin. Verapamil, a P-glycoprotein inhibitor, inhibited the efflux of [3H]vincristine from MBEC4 cells, while 10 microM quercetin significantly stimulated it. The uptake of [3H]vincristine by MBEC4 cells was increased by inhibitors of protein kinase C, but decreased by phorbol 12-myristate-13-acetate (PMA), as well as by 10 microM quercetin. The phosphorylation level of P-glycoprotein was increased in the presence of 5 microM quercetin or 100 nM PMA, but decreased by the protein kinase C inhibitor H7 (1-(5-isoquinolinesulfonyl)-2-methylpiperazine, 30 microM). We conclude that low concentrations of quercetin indirectly activate the transport of [3H]vincristine by enhancing the phosphorylation (and hence activity) of P-glycoprotein, whereas high concentrations of quercetin inhibit P-glycoprotein. Our results indicate that patients taking drugs which are P-glycoprotein substrates may need to restrict their intake of bioflavonoid-containing foods and beverages, such as grapefruit juice.

3-O-Methylglucose↗

PubMed2000

Fluorescence resonance energy transfer from pyrene to perylene labels for nucleic acid hybridization assays under homogeneous solution conditions.

We characterized the fluorescence resonance energy transfer (FRET) from pyrene (donor) to perylene (acceptor) for nucleic acid assays under homogeneous solution conditions. We used the hybridization between a target 32 mer and its complementary two sequential 16 mer deoxyribonucleotides whose neighboring terminals were each respectively labeled with a pyrene and a perylene residue. A transfer efficiency of approximately 100% was attained upon the hybridization when observing perylene fluorescence at 459 nm with 347-nm excitation of a pyrene absorption peak. The Förster distance between two dye residues was 22.3 A (the orientation factor of 2/3). We could change the distance between the residues by inserting various numbers of nucleotides into the center of the target, thus creating a gap between the dye residues on a hybrid. Assuming that the number of inserted nucleo-tides is proportional to the distance between the dye residues, the energy transfer efficiency versus number of inserted nucleotides strictly obeyed the Förster theory. The mean inter-nucleotide distance of the single-stranded portion was estimated to be 2.1 A. Comparison between the fluorescent properties of a pyrene-perylene pair with those of a widely used fluorescein-rhodamine pair showed that the pyrene-perylene FRET is suitable for hybridization assays.

DNA, Bacterial↗

PubMed2000

Clinicopathologic significance of urokinase receptor- and MMP-9-positive stromal cells in human colorectal cancer: functional multiplicity of matrix degradation on hematogenous metastasis.

Our previous clinicopathologic study revealed an inverse association of liver metastasis of colorectal cancer and stromal expression of matrix metalloproteinase-9 (MMP-9) or urokinase receptor (uPAR). This suggests that host cells, particularly macrophages, expressing matrix-degrading enzymes/factors could be protective for the host against hematogenous metastasis. However, our previous study was unable to differentiate whether our results were causes or effects of widely spread cancer. To solve this point, we designed the present study on colorectal cancers that developed hematogenous metastasis after operation, ie., metachronous hematogenous metastasis. These cancers, being solely micrometastasized at the time of operation, allowed us to eliminate possible systemic effects by widely spread cancer. Sixty-two primary tumors with metachronous metastasis showed a decreased number of MMP-9+ stromal cells and CD68+ macrophages along the invasive margin with unchanged uPAR+ stromal area as compared with those in 72 control cases, which were free from tumor metastasis or recurrence for more than 5 years. Therefore, we judged the decrease of MMP-9+ host cells or macrophages in the primary site is irrelevant of effects of widely spread metastasis but probably related to causes of metastasis. Our data also characterized the metachronous metastasis group by uPAR expression in fibroblasts. The number of uPAR+ cancer cells, although small in number, were also larger in the metachronous metastasis group. Our data revealed that macrophages, a major source of uPAR and one of the sources of MMP-9, could be inhibitory to hematogenous metastasis, while uPAR+ fibroblasts and cancer cells, in turn, facilitate hematogenous metastasis. This suggests the functional multiplicity of matrix degradation processes in cancer tissue.

Bone Neoplasms↗

PubMed2000

Alteration of the sex determining system resulting from structural change of the sex chromosomes in the frog Rana rugosa.

Rana rugosa in Japan is divided into four geographical races on the basis of the karyotype of the sex chromosomes: one in which heteromorphic sex chromosomes occur in the female sex (ZW/ZZ-system), another in which they are present in males (XX/XY-system), and the remaining two in which no heteromorphism is seen in either sex. The last two inherit the XX/XY sex determining system. Y and Z chromosomes in the former two are of the same karyotype as the no. 7 chromosomes seen in one of the latter two, whereas X and W are caused by two inversions that occurred in the original Xs (no. 7). In this study, we first attempted to detect the structural difference between the resulting X and W by examining their chiasma formation. The chiasma distribution between X and W was closely similar to that between two Xs, suggesting that the W and X are identical in structure. Regarding the change from XX/XY- to ZW/ZZ-system, the simplest explanation is that the putative female-determining gene(s) on the W grew functionally stronger by inversions. Next, we examined the sex of triploids having two Xs and one Z. The data showed that the triploids with two original Xs and a Z were all male, whereas most of those with two resulting Xs and a Z developed into females as expected. We speculated that the female-determining gene(s) on the resulting X grew mildly stronger functionally by position effect, whereas those on the W grew much stronger for some other reason (e.g., duplication). J. Exp. Zool. 286:313-319, 2000.

Animals↗

PubMed2000

Determination of erythromycin, clarithromycin, roxithromycin, and azithromycin in plasma by high-performance liquid chromatography with amperometric detection.

In this study, a high-performance liquid chromatographic method was developed for the quantitative determination of erythromycin (EM), roxithromycin (RXM), and azithromycin (AZM) in rat plasma with amperometric detection under a standardized common condition using clarithromycin (CAM) as an internal standard. This method was also proved to be applicable for the determination of CAM by employing RXM as an internal standard. Each drug was extracted from 150 microl of plasma sample spiked with internal standard under an alkaline condition with tert.-butyl methyl ether. The detector cell potential for the oxidation of the drugs was set at +950 mV. The linearity of the calibration curves were preserved over the concentration ranges of 0.1-10 microg/ml for EM and RXM, and 0.03-3.0 microg/ml for CAM and AZM. Coefficients of variation and relative error were less than 9% and +/-7%, respectively. The analytical method presented here was proved to be useful for the investigation of the pharmacokinetic characteristics of EM, CAM, RXM, and AZM in rats.

Animals↗

PubMed2000

In situ expression of fibrogenic growth factors and their receptors in biliary atresia: comparison between early and late stages.

Progressive fibrosis, despite successful surgical treatment, is one of the serious complications of biliary atresia. To understand the mechanism of this fibrosis, the in situ expression of fibrogenic growth factors (TGF-beta and PDGF) and their corresponding receptors was studied by immunohistochemistry using frozen sections. The results were compared between the early (n=12) and late (n=6) stages. The early stage was characterized by abundant expression of all ligands and receptors, together with type I procollagen (PC-I). The major cellular sources were activated fibroblasts/myofibroblasts distributed mostly in the portal tracts. Macrophages also expressed all the ligands and the receptors, but to a lesser degree. Bile duct cells strongly expressed TGF-beta RI and RII and PDGF AA and BB, but focally expressed TGF-beta. All of these decreased in the late stage of biliary atresia. These results suggest that TGF-beta and PDGF play important roles in the fibrogenesis of biliary atresia, especially in its early stage, acting either by autocrine or paracrine mechanisms involving activated fibroblasts/myofibroblasts, bile duct cells, and macrophages.

Adult↗

PubMed2000

Enantioselective determination of bromoisovalerylurea by liquid chromatography on chiral stationary phase in reversed- or normal-phase partition mode.

Bromoisovalerylurea (bromvalerylurea) is a sedative-hypnotic given orally as a racemate. Enantiomers of this drug could be separated by high-performance liquid chromatography on the three chiral stationary phases (a vancomycin-bonded, beta-cyclodextrin derivative-bonded, or urea derivative-bonded phase). Biological fluids of human subjects who had ingested toxic or therapeutic doses of the racemate were chromatographed after liquid-liquid extraction. The (+)-enantiomer concentration was almost equal to the (-)-enantiomer concentration in the serum of one overdosed patient. In all the other subjects, the (+)-enantiomer was less than the (-)-enantiomer in their sera and saliva. The data suggest that the drug is absorbed non-stereoselectively from the gastrointestinal tract and eliminated from the blood stereoselectively.

Adult↗

PubMed2000

Quantitative prediction of catalepsy induced by amoxapine, cinnarizine and cyclophosphamide in mice.

Parkinsonism can be a side effect of antipsychotic drugs, and has recently been reported with peripherally acting drugs such as calcium channel blockers, antiarrhythmic agents and so on. In this study, we examined the quantitative prediction of drug-induced catalepsy by amoxapine, cinnarizine and cyclophosphamide, which have been reported to induce parkinsonism. Dose-dependent catalepsy was induced by these drugs in mice. In vivo dopamine D(1), D(2) and muscarinic acetylcholine (mACh) receptor occupancies by these drugs in the striatum were also examined. The in vitro binding affinities (K(i) values) of amoxapine and cinnarizine to dopamine D(1), D(2) and mACh receptors in rat striatal synaptic membrane were 200 and 2900 nM, 58.4 and 76.4 nM and 379 and 290 nM, respectively. Cyclophosphamide did not bind to these receptors at concentrations up to 100 microM. Twenty drugs, including those mentioned above, showed a significant correlation between the observed intensity of catalepsy and the values predicted with a pharmacodynamic model (Haraguchi K, Ito K, Kotaki H, Sawada Y, Iga T. Prediction of drug-induced catalepsy based on dopamine D(1), D(2), and muscarinic acetylcholine receptor occupancies. Drug Metab Disp 1997; 25: 675-684) based on in vivo occupancy of dopamine D(1), D(2) and mACh receptors. We conclude that occupancy of dopamine D(1) and D(2) receptors contributes to catalepsy induction by amoxapine and cinnarizine.

Amoxapine↗

PubMed2000

A novel bacteriorhodopsin-like protein from Haloarcula japonica strain TR-1: gene cloning, sequencing, and transcript analysis.

The gene encoding a novel bacteriorhodopsin (bR)-like protein from Haloarcula japonica strain TR-1 was cloned and sequenced. The nucleotide sequence of the gene contained an open reading frame that corresponded to a protein of 250 amino acids. The deduced amino acid sequence of Ha. japonica bR-like protein exhibited the highest homology to those of cruxrhodopsins (cRs) produced by members of the genus Haloarcula, suggesting that the bR-like protein of Ha. japonica belonged to the cR subfamily. The hydropathy analysis of Ha. japonica bR-like protein (cR) revealed that the Ha. japonica cR had a transmembrane heptahelical structure similar to that of bR. Furthermore, transcription of the cR gene in Ha. japonica was confirmed by the reverse transcription-polymerase chain reaction method.

Amino Acid Sequence↗

PubMed2000

Immunological activation of dermal Langerhans cells in contact with lymphocytes in a model of human inflamed skin.

Langerhans cells play an important role in the skin's immune system. Little is known, however, about the antigen-presenting capacity of Langerhans cells in the context of skin inflammation. By immunohistochemistry we investigated the phenotypic characteristics of epidermal and dermal Langerhans cells and their spatial relationship with infiltrating lymphocytes. We studied skin flaps autotransplanted to the oral cavity to fill a defect after maxillofacial cancer surgery. In 15 of 21 cases sampled for the present study, the skin flaps were severely inflamed by Candida albicans infection. In contrast to the normal skin, such inflamed skin showed a marked increase in CD1a(+) dermal Langerhans cells. Double immunohistochemistry revealed that dermal Langerhans cells abundantly expressed B7-2 (CD86), a representative costimulatory molecule, and CD83, a marker of mature dendritic cells. Furthermore, these dermal Langerhans cells were in close contact with CD4(+)/CD45RO(+) lymphocytes. This cell-to-cell contact was further visualized by immunoelectron microscopy. Langerhans cells were also observed within lymphatic vessels that were identified by the expression of vascular endothelial growth factor receptor-3. Ki-67 labeling indices were 4.2% in CD4(+) T cells and 0.8% in CD8(+) T cells within the dermis. Factor XIIIa(+) dermal dendrocytes were distributed outside the clusters of lymphocytes and were not in contact with them. Our observations indicate that dermal Langerhans cells in the inflamed skin are activated to express common phenotypes to mature dendritic cells so that they could stimulate neighboring memory CD4(+) T cells.

Antigens, CD↗

PubMed2000

Induction of apoptosis in mouse brain capillary endothelial cells by cyclosporin A and tacrolimus.

Although cyclosporin A and tacrolimus are used clinically as potent immunosuppressants, there have been reports of neurotoxicity and encephalopathy. A possible mechanism is that these drugs damage the blood-brain barrier (BBB), inducing dysfunction and increased permeability, and are then able to enter the brain. We studied the cytotoxicity of cyclosporin A and tacrolimus, focused on apoptosis induction, using an immortalized cell line established from BALB/c mouse cerebral microvessel endothelial cells (MBEC4). We found that these two drugs induced cell shrinkage, chromatin condensation and DNA fragmentation, which are characteristics of apoptosis. Our data suggest that the induction of apoptosis on the brain capillary endothelial cells may be at least partly involved in the occurrence of immunosuppressant-induced encephalopathy.

Animals↗

PubMed2000

Disappearance of nodular mesangial lesions in a patient with light chain nephropathy after long-term chemotherapy.

A 64-year-old man developed multiple myeloma (kappa light chain type), nephrotic syndrome, and renal insufficiency in 1993. A renal biopsy showed typical histological findings of light chain nephropathy: nodular glomerulosclerosis with deposition of kappa light chains in the mesangial area and subendothelial space of the glomerular capillary walls. Long-term intermittent MEVP chemotherapy (melphalan, 4 mg/d for 4 days; cyclophosphamide, 100 mg/d for 4 days; vincristine, 1 mg/d; prednisolone, 40 mg/d for 4 days) diminished proteinuria and improved renal function. In April 1999, a follow-up biopsy showed remarkable diminution of nodular lesions and disappearance of kappa light chain deposits. Although the prognosis of light chain nephropathy has been considered poor, long-term successful chemotherapy can clear light chain deposits and restore renal function.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed2000

Extensive intraglomerular thrombi of monoclonal IgM-kappa in a patient with malignant lymphoma.

We describe an 80-year-old man who developed malignant lymphoma (ML) complicated by extensive intraglomerular thrombi of immunoglobulin M (IgM)-kappa monoclonal immunoglobulin. The clinical picture was characterized by nephrotic syndrome and systemic lymphadenopathy. Laboratory examination showed mild anemia and a small amount of monoclonal IgM-kappa in the blood. The histopathologic findings and surface immunoglobulin analysis of the lymph node biopsy specimen were consistent with CD5-positive diffuse large B-cell (type, IgM-kappa) lymphoma. The subsequent renal biopsy showed a massive deposition of amorphous material in the glomerular capillary lumens, subendothelial areas, and mesangium. Nodular glomerulosclerosis was not found. An immunofluorescent study showed that the deposits consisted of IgM-kappa monoclonal immunoglobulin. Ultrastructurally, the deposits were composed of granular electron-dense material. Chemotherapy was effective for both the ML and nephrotic syndrome, and the patient's urine analysis results returned to normal. The histopathologic manifestations of this case are rare, and the pathogenesis of these glomerular lesions was obviously associated with ML.

Aged↗

PubMed2000

Propiverine-induced Parkinsonism: a case report and a pharmacokinetic/pharmacodynamic study in mice.

PURPOSE: We present a case report of propiverine-induced Parkinsonism. We previously reported the induction of catalepsy by amiodarone, aprindine and procaine, which possess a diethylaminomethyl moiety and demonstrated selective blockade of dopamine D2 receptors by these drugs in mice. We hypothesized that drugs possessing a diethylaminomethyl structure may generally induce Parkinsonism and/or catalepsy. METHODS: Thus, we performed a study to examine whether oxybutynin, pentoxyverine and etafenone, as well as propiverine, induce catalepsy in mice. RESULTS: The intensity of drug-induced catalepsy was in the order: haloperidol > etafenone > pentoxyverine > propiverine > oxybutynin. In vivo occupancy of dopamine D1, D2 and mACh receptors in the striatum was also examined. The in vitro binding affinities to the D1, D2 and mACh receptors in the striatum synaptic membrane were within the ranges of 2.4-140 microM, 380-4,200 nM, and 1.2-2,800 nM, respectively. CONCLUSIONS: These results support the idea that any drug possessing a diethylaminomethyl moiety may contribute to the induction of catalepsy, possibly by occupying dopamine receptors.

Animals↗