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Biomedical subjects

Y Okada

Publications and source records attributed to Y Okada.

At least 199 records · Page 11Linked to original sources
PubMed2000

Hydrophobic extracts of a Chinese herb (CMX-13) exhibit potent immunosuppressive properties and prevent acute rejection in a highly histoincompatible model of rat lung transplantation.

BACKGROUND: The potential of higher plants as sources for new immunosuppressive medications is well recognized. In our experiments we investigated the immunosuppressive effect of a highly refined and potent extract of a Chinese herbal preparation, CMX-13, on inhibiting acute allograft rejection (AR) in a highly histoincompatible rat lung transplant model, BN-->LEW, and on lymphocyte activation and cytokine gene expression in vitro. METHODS: Left lung transplants: the control group (group 1) received only dimethylsulfoxide (DMSO) which is the solvent for CMX-13. Group 2 received intramuscular cyclosporin A (CsA, 25 mg/kg) on day 2 posttransplant. Group 3 and 4 received i.p. CMX-13 (0.5 mg/day, low dose and 5 mg/day, high dose, respectively) on day 1, 2, and 3 posttransplant. All animals were killed on day 6 posttransplant. Several pathological categories of inflammation were examined. In vitro experiments: rat spleen cells were incubated with Con A or irradiated stimulator cells with/without serial dilutions of CMX-13 or CsA. Cell proliferation was measured by 3H-thymidine incorporation. mRNA expression of interleukin-2 and interferon-gamma was examined by reverse transcriptase-polymerase chain reaction. RESULTS: The severity of AR in animals receiving high dose CMX-13 was significantly reduced (stage II, P<0.05) compared with controls (stage IV). Significant differences were also seen when more specific parameters of inflammation were examined (necrosis, 0 vs. 1.7+/-1.0, P<0.05; interalveolar hemorrhage, 0 vs. 3.0+/-0.9, P<0.05). The responses seen in the animals treated with high dose CMX-13 were similar to those in the CsA group. CMX-13 inhibited T cell proliferative responses induced by Con A and alloantigen stimulation in a dose-dependent manner that were similar to CsA. Interleukin-2, and interferon-gamma mRNA expression in Con A-stimulated spleen cells was not inhibited by CMX-13 although CsA showed significant inhibition. CONCLUSIONS: 1) CMX-13 significantly reduces the stage of AR and parameters of inflammation in a highly histoincompatible rat lung transplant model. 2) CMX-13 has equal potency to CsA in the inhibition of Con A and alloantigen stimulated rat spleen cell proliferation. 3) CMX-13 showed no inhibitory effects on IL-2 and gamma-IFN mRNA expression, suggesting that its mechanism of action is different from CsA. 4) CMX-13 or derivatives may have potential utility as an immunosuppressive agent(s) in modulation of AR and management of other inflammatory and immunological disorders.

Acute Disease↗
PubMed2000

Inhibitory effect of various thrombin inhibitors on shear-induced platelet function and dynamic coagulation.

We assessed the effects of active site-directed, fibrinogen recognition exosite (FRE)-directed and bifunctional thrombin inhibitors, on shear-induced platelet reactivity (adhesion/aggregation) and dynamic coagulation (coagulation of flowing blood). An in vitro test for shear-induced haemostatic plug formation and dynamic coagulation (haemostatometry) was employed using non-anticoagulated rat blood. The active site-directed inhibitors (argatroban, P891, P899) caused inhibition of platelet reactivity and coagulation at 1-, 100- and 100-microM concentrations, respectively. Bifunctional inhibitors (P553, P1053) exerted inhibitory effects at 0.1 microM. A dimeric bifunctional inhibitor P824 caused significant inhibition at 1 microM. The FRE-directed inhibitor (P960) inhibited shear-induced platelet reactivity at 10 microM but the dynamic coagulation at 1 microM. Combination of active site-directed argatroban and FRE-directed P960 did not show any synergistic effect. The most potent inhibition was observed in monomeric bifunctional inhibitors. The inhibitory effects were compared with the K(i) values against human thrombin and with the IC(50) values against fibrin clot formation. The minimum effective concentrations on shear-induced platelet reactivity and dynamic coagulation were comparable with the IC(50) values, but not with the K(i) values.

Animals↗
PubMed2000

Development of plasmin and plasma kallikrein selective inhibitors and their effect on M1 (melanoma) and HT29 cell lines.

trans-4-Aminomethylcyclohexanecarbonyl-Tyr(O-Pic)-octylamide (YO-2) inhibited plasmin (PL) selectively, while trans-4-aminomethylcyclohexanecarbonyl-Phe-4-carboxymethylanili de (YO-1) inhibited plasma kallikrein (PK). YO-2 induced apoptosis of M1 (melanoma) cell line and HT29 colon carcinoma cells during 24 h through activation of caspase-3, while YO-1 did not affect either cell line even during 48 h.

Antineoplastic Agents↗
PubMed2000

Expression of emmprin (CD147), a cell surface inducer of matrix metalloproteinases, in normal human brain and gliomas.

EMMPRIN (extracellular matrix metalloproteinase inducer), also called CD147, basigin or M6 in the human, is a member of the immunoglobulin superfamily that is present on the surface of tumor cells and stimulates adjacent fibroblasts to produce matrix metalloproteinases (MMPs). In our study, we investigated expression of EMMPRIN in human normal brain and gliomas, since mouse basigin and chicken HT7, the species homologues of human EMMPRIN, are associated with neuronal interactions and normal blood-brain barrier function, respectively. EMMPRIN expression was detected in all samples of non-neoplastic brain and glioma tissues examined. However, expression levels of EMMPRIN mRNA and protein were significantly higher in gliomas than in non-neoplastic brain. Moreover, levels of mRNA expression and immunohistochemical staining correlated with tumor progression in gliomas: They were highest in the most malignant form of glioma, glioblastoma multiforme, followed by anaplastic astrocytoma and then low-grade astrocytoma. Also, immunolocalization revealed quite different distributions in non-neoplastic brain and glioma: EMMPRIN was demonstrated only in vascular endothelium in non-neoplastic regions of the brain, whereas it was present in tumor cells but not in proliferating blood vessels in malignant gliomas. These data indicate that an MMP inducer molecule EMMPRIN is differently expressed in human normal brain and gliomas and could be associated with astrocytoma progression. Possible mechanisms whereby glioma cell EMMPRIN could influence tumor progression will be discussed.

Antigens, CD↗
PubMed2000

Synthesis of stereoisomeric analogues of endomorphin-2, H-Tyr-Pro-Phe-Phe-NH(2), and examination of their opioid receptor binding activities and solution conformation.

All sixteen stereoisomeric analogues of endomorphin-2 (H-Tyr-Pro-Phe-Phe-NH(2)) were synthesized by Fmoc-strategy using solid phase methods. Although synthetic endomorphin-2 exhibited similar mu- and delta-opioid receptor-binding activity to the natural compound, endomorphin-2 analogues containing d-amino acid isomers exhibited lower interaction with mu-receptors depending on the particular combination. The data clearly indicated that the three dimensional structure of endomorphin-2 with the natural l-configuration was the most suitable for binding within the mu receptor, but specific residues are important for activity. Circular dichroism studies verified that changes in chirality of amino acids in the endomorphin-2 sequence resulted in structural conformation. These alterations significantly reduced the specificity for mu-receptor-binding sites.

Animals↗
PubMed2000

Effect of combined leukotriene D(4) and thromboxane A(2) receptor antagonist on mediator-controlled resistance in guinea pigs.

The effects of YM158 (3-[(4-tert-butylthiazol-2-yl)methoxy]-5'-[3-(4-chlorobenzenesu lfonyl )propyl]-2'-(1H-tetrazol-5-ylmethoxy)benzanilide monosodium salt monohydrate), a new dual antagonist for leukotriene D(4) and thromboxane A(2) receptors, on antigen-induced increases in airway resistance were investigated in mediator-controlled novel asthmatic models using actively sensitized guinea pigs. While the predominant mediator was thromboxane A(2), complete inhibition of cyclooxygenase induced mediation by cysteinyl-leukotrienes. About 1-mg/kg indomethacin induced a state where both mediators participated equally. YM158 inhibited increases in resistance whether only one or both mediators were involved. When leukotriene D(4) and thromboxane A(2) equally participated, ED(50) values for 4-oxo-8-[4-(4-phenylbutoxy)benzoylamino]-2-(tetrazol-5-yl)-4 H-1-benzo pyran hemihydrate (pranlukast; 3.9 mg/kg) and 7-(3,5,6-trimethyl-1, 4-benzoquinon-2-yl)-7-phenylheptanoic acid (seratrodast; 2.1 mg/kg) were similar to that for YM158 (8.3 mg/kg), although those effects on the corresponding mediator-induced reaction were 10 times stronger than those of YM158. Additionally, the maximum inhibition of YM158 was stronger than those of either single receptor antagonist. In conclusion, YM158 has a potentially greater efficacy in wider types of experimental asthmatic models than single receptor antagonists.

Administration, Oral↗
PubMed2000

Glioma cell extracellular matrix metalloproteinase inducer (EMMPRIN) (CD147) stimulates production of membrane-type matrix metalloproteinases and activated gelatinase A in co-cultures with brain-derived fibroblasts.

Extracellular matrix metalloproteinase inducer (EMMPRIN) also called CD147, basigin or M6 in the human is a member of the immunoglobulin superfamily that is enriched on the surface of tumor cells and stimulates adjacent stromal cells to produce several matrix metalloproteinases (MMPs). In this study, we have demonstrated that coculturing of EMMPRIN-expressing human glioblastoma multiforme cells (U251) with brain-derived human fibroblasts not only stimulates production, but also activation of pro-gelatinase A (proMMP-2), an enzyme that is enriched in malignant gliomas and most likely crucial to tumor progression. Production of membrane types 1 and 2-MMPs (MT1-MMP and MT2-MMP), which are activators of proMMP-2, was also stimulated in these cocultures. Stimulation of MMP-2, MT1-MMP and MT2-MMP production was inhibited by anti-EMMPRIN monoclonal antibody in a dose-dependent manner. Thus, we have shown, for the first time, that EMMPRIN causes increased expression of MT1-MMP and MT2-MMP, as well as increased production and activation of MMP-2.

Antibodies, Monoclonal↗
PubMed2000

KIF5C, a novel neuronal kinesin enriched in motor neurons.

Kinesin superfamily proteins (KIFs) are the molecular motors conveying cargos along microtubules. KIF5s, the heavy chains of conventional kinesin (KHC), are originally identified members of KIFs, and neuronal KIF5A and ubiquitous KIF5B have been identified so far. In the present work, we cloned a novel member of KIF5, KIF5C, and generated specific antibodies against three KIF5s to investigate their distribution and functions. KIF5A showed pan-neuronal distribution in the nervous system. KIF5B showed a glial cell distribution pattern in general; however, interestingly, its expression was strongly upregulated in axon-elongating neurons, such as olfactory primary neurons and mossy fibers. KIF5C was also a neuronal KIF5 like KIF5A but was highly expressed in lower motor neurons in 2-week-old or older mice, suggesting its important roles in the maintenance of motor neurons rather than in their formation, such as axonal elongation. Because a large part of KIF5s in adult motor neurons were expected to be KIF5C, we generated mice lacking the kif5C gene to investigate the functions of KIF5C in neurons in living animals. The mutant mice showed smaller brain size but were viable and did not show gross changes in the nervous system. Closer examinations revealed the relative loss of motor neurons to sensory neurons. Because three KIF5s showed high similarity in the amino acid sequence, could rescue the KIF5B mutant cells, and could form heterodimers, we think that there are functional redundancy among the three KIF5s and that KIF5A and KIF5B prevented the KIF5C null mice from the severe phenotype.

Amino Acid Sequence↗
PubMed2000

The influence of lorazepam on somatosensory-evoked fast frequency (600 Hz) activity in MEG.

The generators of high frequency bursts (600-Hz activity) detected at the parietal scalp over the primary somatosensory cortex after electrical stimulation of peripheral nerves are not yet known. We investigated the influence of benzodiazepine on the somatosensory-evoked 600-Hz activity by means of neuromagnetic measurements and source analysis. After oral administration of lorazepam, the latency of the 600-Hz burst activity was increased; specifically later peaks were delayed more than earlier peaks. In contrast, the latency of the concurrent primary cortical low frequency response (N20m) was not significantly changed. The source strengths of both N20m and 600-Hz bursts were significantly increased. Our results provide evidence for two components of the 600-Hz activity with a different generator structure.

Adult↗
PubMed2000

CalDAG-GEFIII activation of Ras, R-ras, and Rap1.

We characterized a novel guanine nucleotide exchange factor (GEF) for Ras family G proteins that is highly homologous to CalDAG-GEFI, a GEF for Rap1 and R-Ras, and to RasGRP/CalDAG-GEFII, a GEF for Ras and R-Ras. This novel GEF, referred to as CalDAG-GEFIII, increased the GTP/GDP ratio of Ha-Ras, R-Ras, and Rap1 in 293T cells. CalDAG-GEFIII promoted the guanine nucleotide exchange of Ha-Ras, R-Ras, and Rap1 in vitro also, indicating that CalDAG-GEFIII exhibited the widest substrate specificity among the known GEFs for Ras family G proteins. Expression of CalDAG-GEFIII was detected in the glial cells of the brain and the glomerular mesangial cells of the kidney by in situ hybridization. CalDAG-GEFIII activated ERK/MAPK most efficiently, followed by CalDAG-GEFII and CalDAG-GEFI in 293T cells. JNK activation was most prominent in cells expressing CalDAG-GEFII, followed by CalDAG-GEFIII and CalDAG-GEFI. Expression of CalDAG-GEFIII induced neuronal differentiation of PC12 cells and anchorage-independent growth of Rat1A cells less efficiently than did CalDAG-GEFII. Thus, co-activation of Rap1 by CalDAG-GEFIII apparently attenuated Ras-MAPK-dependent neuronal differentiation and cellular transformation. Altogether, CalDAG-GEFIII activated a broad range of Ras family G proteins and exhibited a biological activity different from that of either CalDAG-GEFI or CalDAG-GEFII.

Animals↗
PubMed2000

Normotonic cell shrinkage because of disordered volume regulation is an early prerequisite to apoptosis.

A major hallmark of apoptosis is normotonic shrinkage of cells. Here, we studied the relation between apoptotic cell shrinkage and apoptotic cell death. Induction of the apoptotic volume decrease (AVD) under normotonic conditions was found to be coupled to facilitation of the regulatory volume decrease (RVD), which is known to be attained by parallel operation of Cl(-) and K(+) channels, under hypotonic conditions. Both the AVD induction and the RVD facilitation were found to precede cytochrome c release, caspase-3 activation, DNA laddering, and ultrastructural alterations in three cell types after apoptotic insults with two distinct apoptosis inducers. Also, the AVD was not prevented by a broad-spectrum caspase inhibitor. When the AVD induction and the RVD facilitation were prevented by blocking volume-regulatory Cl(-) or K(+) channels, these cells did not show succeeding apoptotic biochemical and morphological events and were rescued from death. Thus, it is concluded that the AVD, which is caused by disordered cell volume regulation, is an early prerequisite to apoptotic events leading to cell death.

Animals↗
PubMed2000

Recognition of formamidopyrimidine by Escherichia coli and mammalian thymine glycol glycosylases. Distinctive paired base effects and biological and mechanistic implications.

The activity of prokaryotic and mammalian thymine glycol (Tg) glycosylases including Escherichia coli endonuclease III (Endo III) and endonuclease VIII (Endo VIII) and mouse Endo III homologue (mNth1) for formamidopyrimidine (Fapy) has been investigated using defined oligonucleotide substrates. 2, 6-Diamino-4-hydroxy-5-N-methylformamidopyrimidine, a methylated Fapy derived from guanine, was site specifically incorporated in the oligonucleotide. The substrates containing Fapy:N pairs (N = A, G, C, T) as well as a Tg:A pair, a physiological substrate of Endo III, Endo VIII, and mNth1, were treated by the enzymes and nicked products were quantified by gel electrophoresis. The activity of Endo III and Endo VIII for Fapy varied markedly depending on the paired base, being the highest with G (activity relative to Tg = 0. 55 (Endo III) and 0.41 (Endo VIII)) and the lowest with C (0.05 (Endo III) and 0.06 (Endo VIII)). In contrast, mNth1 recognized all Fapy pairs equally well and the activity was comparable to Tg. The results obtained in the nicking assay were further substantiated by the analysis of the Schiff base intermediate using NaBH(4) trapping assays. These results indicate that Escherichia coli and mammalian Tg glycosylases have a potential activity to recognize Fapy. However, as demonstrated for Fapy:C pairs, their distinctive activities implicate unequal participation in the repair of Fapy lesions in cells.

Animals↗
PubMed2000

Assignment of functional amino acids around the active site of human DNA topoisomerase IIalpha.

An expression library for active site mutants of human topoisomerase IIalpha (TOP2alpha) was constructed by replacing the sequence encoding residues 793-808 with a randomized oligonucleotide cassette. This plasmid library was transformed into a temperature-sensitive yeast strain (top2-1), and viable transformants were selected at the restrictive temperature. Among the active TOP2alpha mutants, no substitution was allowed at Tyr(805), the 5' anchor of the cleaved DNA, and only conservative substitutions were allowed at Leu(794), Asp(797), Ala(801), and Arg(804). Thus, these 5 residues are critical for human TOP2alpha activity, and the remaining mutagenized residues are less critical for function. Using the x-ray crystal structure of yeast TOP2 as a structural model, it can be deduced that these 5 functionally important residues lie in a plane. One of the possible functions of this plane may be that it interacts with the DNA substrate upon catalysis. The side chains of Ser(803) and Lys(798), which confer drug resistance, lie adjacent to this plane.

Amino Acid Sequence↗
PubMed2000

ADAMTS-1 cleaves a cartilage proteoglycan, aggrecan.

A disintegrin-like and metalloproteinase with thrombospondin type I motifs-1 (ADAMTS-1) is an extracellular matrix-anchored metalloproteinase. In this study we have demonstrated that ADAMTS-1 is able to cleave a major cartilage proteoglycan, aggrecan. N-terminal sequencing analysis of the cleavage product revealed that ADAMTS-1 cleaves the Glu(1871)-Leu(1872) bond within the chondroitin sulfate attachment domain of aggrecan. In addition, deletional analysis demonstrated that the C-terminal spacer region of ADAMTS-1 is necessary to degrade aggrecan. These results suggest that ADAMTS-1 may be involved in the turnover of aggrecan in vivo.

ADAM Proteins↗
PubMed2000

Self-assembled peptide fibers from valylvaline bola-amphiphiles by a parallel beta-sheet network.

A series of dipeptide-based bola-amphiphiles, bis(N-alpha-amide-L-valyl-L-valine) 1, n-alkane dicarboxylate (n=4-12), have been synthesized. The bola-amphiphiles with n=4 and 6 self-assembled to form crystalline solids in water, whereas those with n=7-12 produced peptide fibers. FT-IR spectroscopy and X-ray diffraction patterns revealed that the peptide fibers have parallel-type beta-sheet networks between the valylvaline units. FT-IR deconvolution study of carboxyl regions indicated that these crystalline solids and peptide fibers are stabilized by interlayer bifurcated and intralayer lateral hydrogen-bond networks between the end carboxylic acid groups, respectively.

Crystallization↗
PubMed2000

Fournier's gangrene caused by Candida species as the primary organism.

Fournier's gangrene is a rare entity caused by polymicrobial aerobic and anaerobic bacteria. We report a case of Fournier's gangrene caused by Candida as the primary organism. A 65-year-old man presented with perineal soft-tissue infections. He underwent surgical debridement and suprapubic cystostomy with both antifungal and antimicrobial therapy. The histopathologic examination revealed necrotizing fasciitis with Candida species as the sole initial pathogen. The case suggests that primary fungal pathogens should be considered as a causative organism of Fournier's gangrene.

Aged↗
PubMed2000

Front-cell-specific expression of membrane-type 1 matrix metalloproteinase and gelatinase A during cohort migration of colon carcinoma cells induced by hepatocyte growth factor/scatter factor.

Migration of tumor cells is usually assessed as single cell locomotion in vitro using Boyden chamber type assays. In vivo, however, carcinoma cells frequently invade the surrounding tissue as coherent clusters or nests of cells. We have called this type of movement "cohort migration" and developed a two-dimensional in vitro cohort migration model, in which human rectal well-differentiated adenocarcinoma cells (L-10) migrate from piled-up cell islands as coherent sheets of cells when stimulated with hepatocyte growth factor/scatter factor. In this study, we examined whether there is a cohort migration-specific way of expression of matrix metalloproteinases (MMP) and whether degradation of extracellular matrix is necessary for this type of migration. Production of membrane-type 1-MMP (MT1-MMP) and gelatinase A (MMP-2) by L-10 cells was demonstrated by gelatin zymography, immunoblotting, and reverse transcription-PCR. When cohort migration was induced with hepatocyte growth factor/scatter factor, MT1-MMP and MMP-2 were immunolocalized predominantly in the leading edges of the front cells of migrating cell sheets, with the following cells being negative. In addition, during the cohort migration on gelatin-coated substratum, the gelatin matrix was degraded by the cells, in a very organized manner, causing radially arrayed lysis of gelatin matrix at the sites of leading edges. BB94, a synthetic inhibitor specific to MMPs, tissue inhibitor of metalloproteinases-1 and -2, and the COOH-terminal hemopexin-like domain of MMP-2 inhibited the migration on gelatin matrix. Thus, these data demonstrate that gelatin matrix is reorganized to suit cell migration via leading-edge-of-front-cell-specific localization of MT1-MMP and MMP-2 during cohort migration and suggest that the reorganization is essential for this type of migration.

Adenocarcinoma↗