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M Aoki

Publications and source records attributed to M Aoki.

At least 55 records · Page 3Linked to original sources
PubMed2001

Genomic organization of the dysferlin gene and novel mutations in Miyoshi myopathy.

OBJECTIVE: Mutations in the skeletal muscle gene dysferlin cause two autosomal recessive forms of muscular dystrophy: Miyoshi myopathy (MM) and limb girdle muscular dystrophy type 2B (LGMD2B). The purpose of this study was to define the genomic organization of the dysferlin gene and conduct mutational screening and a survey of clinical features in 21 patients with defined molecular defects in the dysferlin gene. METHODS: Genomic organization of the gene was determined by comparing the dysferlin cDNA and genomic sequence in P1-derived artificial chromosomes (PACs) containing the gene. Mutational screening entailed conformational analysis and sequencing of genomic DNA and cDNA. Clinical records of patients with defined dysferlin gene defects were reviewed retrospectively. RESULTS: The dysferlin gene encompasses 55 exons spanning over 150 kb of genomic DNA. Mutational screening revealed nine novel mutations associated with MM. The range of onset in this patient group was narrow with a mean of 19.0 +/- 3.9 years. CONCLUSION: This study confirms that the dysferlin gene is mutated in MM and LGMD2B and extends understanding of the timing of onset of the disease. Knowledge of the genomic organization of the gene will facilitate mutation detection and investigations of the molecular biologic properties of the dysferlin gene.

Adolescent↗
PubMed2001

Oncogenic transformation induced by membrane-targeted Akt2 and Akt3.

The kinases Akt2, Akt3 and their myristylated variants, Myr-Akt2 and Myr-Akt3 were expressed by the RCAS vector in chicken embryo fibroblasts (CEF). Myr-Akt2 and Myr-Akt3 were strongly oncogenic, inducing multilayered foci of transformed cells. In contrast, wild-type Akt2 and Akt3 were only poorly transforming, their efficiencies of focus formation were more than 100-fold lower; foci appeared later and showed less multilayering. Addition of the myristylation signal not only enhanced oncogenic potential but also increased kinase activities. Myr-Akt2 and Myr-Akt3 also induced hemangiosarcomas in the animal, whereas wild type Akt2 and Akt3 were not oncogenic in vivo. Furthermore, Akt2, driven by the lck (lymphocyte specific kinase) promoter in transgenic mice, induced lymphomas. The oncogenic effects of Akt2 and Akt3 described here are indistinguishable from those of Akt1. The downstream targets relevant to oncogenic transformation are therefore probably shared by the three Akt kinases.

Animals↗
PubMed2001

Novel low molecular weight spirodiketopiperazine derivatives potently inhibit R5 HIV-1 infection through their antagonistic effects on CCR5.

Novel low molecular weight spirodiketopiperazine derivatives which potently inhibit R5 human immunodeficiency virus type 1 (HIV-1) infection through their antagonistic effects on CCR5 were identified. One such compound E913 (M(r) 484) specifically blocked the binding of macrophage inflammatory protein-1alpha (MIP-1alpha) to CCR5 (IC(50) 0.002 microm) and MIP-1alpha-elicited cellular Ca(2+) mobilization (IC(50) approximately 0.02 microm). E913 potently inhibited the replication of laboratory and primary R5 HIV-1 strains as well as various multidrug-resistant monocyte/macrophage tropic (R5) HIV-1 at IC(50) values of 0.03 to 0.06 microm. E913 was inactive against T cell tropic (X4) HIV-1; however, when combined with a CXCR4 antagonist AMD-3100, E913 potently and synergistically inhibited the replication of dualtropic HIV-1 and a 50:50 mixture of R5 and X4 HIV-1. Antagonism in anti-HIV-1 activity was not seen when E913 was combined with the reverse transcriptase inhibitor zidovudine or protease inhibitors. E913 proved to compete with the binding of antibodies to CCR5 which recognize the C-terminal half of the second extracellular loop (ECL2B) of CCR5. E913 and its analogs are acid-resistant and orally bioavailable in rodents. These data warrant that spirodiketopiperazine derivatives be further developed as potential therapeutics for HIV-1 infection.

Animals↗
PubMed2001

Concentration of vascular endothelial growth factor in the tumour tissue as a prognostic factor of soft tissue sarcomas.

Previous studies have shown that the prognosis of patients who have tumours with high microvessel density (MVD) is worse than that of patients who have a lower density in a variety of cancers. In this study, we investigated the clinical relevance of neovascularity assessed by MVD and the concentration of vascular endothelial growth factor (VEGF) in the tumour tissue of patients with soft tissue sarcoma in comparison with major clinicohistologic parameters by univariate and multivariate analysis. In 115 patients with soft tissue sarcoma, MVD was measured by counting vessels stained with factor VIII antibody. The concentration of VEGF in the tumour tissue was determined by enzyme-linked immunosorbent assay. These parameters were then compared with disease outcome. The concentration of VEGF in the tumour tissue, but not MVD, was found to be correlated with disease outcome in patients with soft tissue, sarcoma. VEGF concentration in the tumour tissue showed a relationship with the clinical stage and histologic grade of the tumour. There was no significant difference in the levels of tissue VEGF concentration and MVD among soft tissue sarcomas classified according to histologic type. The level of tissue VEGF concentration in patients who had subsequent local recurrence and metastasis were significantly higher than the respective values in patients who did not have such disease outcome. No significant correlation existed between MVD and the concentration of VEGF in the tumour tissue. Univariate analysis showed that a high tissue VEGF concentration was associated with poor overall survival of the patient and a greater probability that local recurrence and metastasis had occurred. Multivariate analysis revealed that the tissue concentration of VEGF is an independent prognostic factor for the disease outcome of patients with soft tissue sarcoma. VEGF concentration in the tumour tissue, but not MVD, is an additional prognostic parameter for disease outcome in patients with soft tissue sarcoma, regardless of histologic type.

Adult↗
PubMed2001

Missense mutation of the interleukin-12 receptor beta1 chain-encoding gene is associated with impaired immunity against Mycobacterium avium complex infection.

Interleukin-12 (IL-12) plays an important role in the production of interferon gamma (IFN-gamma) and is essential for protection against intracellular pathogens such as Mycobacterium and Salmonella. A 31-year-old man had disseminated Mycobacterium avium complex (MAC) infection. The production of IFN-gamma by peripheral blood mononuclear cells stimulated with phytohemagglutinin (PHA-PBMCs) was found severely impaired (40.7 pg/mL compared with 833 +/- 289 pg/mL for the patient's and healthy subjects' (n = 3) PHA- PBMCs, respectively), and the patient's PHA-PBMCs completely lacked surface IL-12 receptor beta1 (IL-12Rbeta1) chain. The IL-12Rbeta1 gene transcript in his PHA-PBMCs had an R213W substitution in each allele. Family history showed that both parents were heterozygotes in the R213W substitution. Transfection of a human embryonal kidney cell line 293 (HEKC293) with wild-type IL-12Rbeta1wt gene led to cell surface IL-12Rbeta1 expression; however, no expression was seen in HEKC293 transfected with the mutated IL-12Rbeta1R213W gene. The IL-12Rbeta1 gene transcript, but no IL-12Rbeta1 protein, was detected in PHA-PBMCs and T cells, suggesting a post-translational event(s), most likely a shortened turnover of the protein. The R213W substitution was not detected in the cells of 32 healthy persons or of 25 patients with tuberculosis or MAC infection. Six amino acid substitutions (Q214R, M365T, G378R, H438Y, A525T, and G594E) were identified, but the incidences of such substitutions were not significantly different between the groups. The Q214R substitution is reportedly linked to IL-12Rbeta1 deficiency; however, the study showed that 19 and 10 of 57 Japanese and 6 and 4 of 33 healthy white persons were heterozygous and homozygous for Arg-214, respectively, suggesting that the Q214R substitution represents a polymorphism and is not related to IL-12Rbeta1 deficiency but that the R213W substitution is responsible for IL-12Rbeta1 deficiency.

Adult↗
PubMed2001

2-Acylimino-3H-thiazoline derivatives: a novel template for platelet GPIIb/IIIa receptor antagonists.

In the course of our research for the low-molecular weight RGD peptide mimics, we have found that a rigid 2-acylimino-3H-thiazoline structure is suitable for the peptide backbone mimics. Introduction of amidinophenyl and beta-alanine moiety as arginine and aspartic acid side-chain surrogates to this backbone mimic resulted in a highly potent fibrinogen receptor antagonist 2-(4-amidinobenzoylimino)-3,4-dimethyl-N-(2-carboxyethyl)-3H-thiazoline-5-carboxamide (7c), namely PS-028 (Ki = 46.5 +/- 5.8 microM).

Animals↗
PubMed2001

Coumarin and chromen-4-one analogues as tautomerase inhibitors of macrophage migration inhibitory factor: discovery and X-ray crystallography.

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine released from T-cells and macrophages. Although a detailed understanding of the biological functions of MIF has not yet been clarified, it is known that MIF catalyzes the tautomerization of a nonphysiological molecule, D-dopachrome. Using a structure-based computer-assisted search of two databases of commercially available compounds, we have found 14 novel tautomerase inhibitors of MIF whose K(i) values are in the range of 0.038-7.4 microM. We also have determined the crystal structure of MIF complexed with the hit compound 1. It showed that the hit compound is located in the active site of MIF containing the N-terminal proline which plays an important role in the tautomerase reaction and forms several hydrogen bonds and undergoes hydrophobic interactions. A crystallographic study also revealed that there is a hydrophobic surface which consists of Pro-33, Tyr-36, Trp-108, and Phe-113 at the rim of the active site of MIF, and molecular modeling studies indicated that several more potent hit compounds have the aromatic rings which can interact with this hydrophobic surface. To our knowledge, our compounds are the most potent tautomerase inhibitors of MIF. One of these small, drug-like molecules has been cocrystallized with MIF and binds to the active site for tautomerase activity. Molecular modeling also suggests that the other hit compounds can bind in a similar fashion.

Binding Sites↗
PubMed2001

Synthesis and antifungal activities of novel 1,3-beta-D-glucan synthase inhibitors. Part 1.

Highly potent 1,3-beta-D-glucan synthase inhibitors 10, 11 and 13 have been identified by the chemical modification of the fungicidal macrocyclic lipopeptidolactone, RO-09-3655 (1), isolated from the cultured broth of Deuteromycotinia spp. D-Ornithine derivative (10) showed improved antifungal activity in the systemic candidiasis model in mice and reduced hepatotoxicity in vitro, as compared with 1.

Animals↗
PubMed2001

Amyotrophic lateral sclerosis-linked glutamate transporter mutant has impaired glutamate clearance capacity.

We have investigated the functional impact of a naturally occurring mutation of the human glutamate transporter GLT1 (EAAT2), which had been detected in a patient with sporadic amyotrophic lateral sclerosis. The mutation involves a substitution of the putative N-linked glycosylation site asparagine 206 by a serine residue (N206S) and results in reduced glycosylation of the transporter and decreased uptake activity. Electrophysiological analysis of N206S revealed a pronounced reduction in transport rate compared with wild-type, but there was no alteration in the apparent affinities for glutamate and sodium. In addition, no change in the sensitivity for the specific transport inhibitor dihydrokainate was observed. However, the decreased rate of transport was associated with a reduction of the N206S transporter in the plasma membrane. Under ionic conditions, which favor the reverse operation mode of the transporter, N206S exhibited an increased reverse transport capacity. Furthermore, if coexpressed in the same cell, N206S manifested a dominant negative effect on the wild-type GLT1 activity, whereas it did not affect wild-type EAAC1. These findings provide evidence for a role of the N-linked glycosylation in both cellular trafficking and transport function. The resulting alteration in glutamate clearance capacity likely contributes to excitotoxicity that participates in motor neuron degeneration in amyotrophic lateral sclerosis.

ATP-Binding Cassette Transporters↗
PubMed2001

A role of the kinase mTOR in cellular transformation induced by the oncoproteins P3k and Akt.

The oncoproteins P3k (homolog of the catalytic subunit of class IA phosphoinositide 3-kinase) and Akt (protein kinase B) induce oncogenic transformation of chicken embryo fibroblasts. The transformed cells show constitutive phosphorylation of the positive regulator of translation p70S6 kinase (S6K) and of the eukaryotic initiation factor 4E-BP1 binding protein (4E-BP1), a negative regulator of translation. Phosphorylation activates S6K and inactivates 4E-BP1. A mutant of Akt that retains kinase activity but does not induce phosphorylation of S6K or of 4E-BP1 fails to transform chicken embryo fibroblasts, suggesting a correlation between the oncogenicity of Akt and phosphorylation of S6K and 4E-BP1. The macrolide antibiotic rapamycin effectively blocks oncogenic transformation induced by either P3k or Akt but does not reduce the transforming activity of 11 other oncoproteins. Rapamycin inhibits the kinase mTOR, an important regulator of translation, and this inhibition requires binding of the antibiotic to the immunophilin FKBP12. Displacement of rapamycin from FKBP12 relieves the inhibition of mTOR and also restores P3k-induced transformation. These data are in accord with the hypothesis that transformation by P3k or Akt involves intervention in translational controls.

Animals↗
PubMed2001

Inhibition of neointimal formation after balloon injury by cilostazol, accompanied by improvement of endothelial dysfunction and induction of hepatocyte growth factor in rat diabetes model.

AIMS/HYPOTHESIS: Cilostazol, a well-known phosphodiesterase type 3 (PDE3) inhibitor for the treatment of peripheral arterial disease, has vasodilator properties and an anti-proliferative action on the growth of vascular smooth muscle cells. In this study, we tested whether cilostazol inhibits neointimal formation and improves endothelial dysfunction after balloon injury in non-diabetic and diabetic rats. METHODS: Cilostazol or vehicle was administered to non-diabetic and streptozotocin-induced diabetic rats from 7 days before to 14 days after balloon injury of the carotid artery. We focused on the expression of hepatocyte growth factor to explore how cilostazol improved endothelial dysfunction. Also, we studied the effects of cilostazol on hepatocyte growth factor production in in vitro experiments. RESULTS: At 14 days after injury, the ratio of neointimal to medial area was decreased in rats treated with cilostazol in non-diabetic and diabetic animals. The impaired response to acetylcholine in balloon injured vessels was improved by cilostazol in non-diabetic and diabetic rats (p < 0.05). Vascular hepatocyte growth factor concentration was decreased in injured vessels of non-diabetic rats compared to uninjured vessels. Moreover, hepatocyte growth factor was further decreased in injured vessels of diabetic rats as compared to those of non-diabetic rats (p < 0.05). Of note, administration of cilostazol attenuated the decrease in hepatocyte growth factor concentration in injured vessels of both non-diabetic and diabetic rats (p < 0.01). Increase in vascular hepatocyte growth factor by cilostazol was confirmed by in vitro experiments showing that cilostazol increased hepatocyte growth factor concentration in cultured human vascular smooth muscle cells, accompanied by cAMP accumulation. CONCLUSION/INTERPRETATION: Our study shows that the increase in vascular hepatocyte growth factor by cilostazol could improve abnormal growth of vascular smooth muscle cells and endothelial dysfunction through rapid regeneration of endothelial cells.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed2001

Neck muscle responses to abrupt vertical acceleration in the seated human.

The control of neck muscle during marked changes in "g" loading must be to protect head-trunk orientation. However, little is known about the organization of reflexes. We therefore investigated the shortest latency in neck muscle evoked by abrupt ascending and descending vertical acceleration with a stroke of 20 cm and peak acceleration 0.4xg in six healthy subjects. The subjects were seated upright and restrained on a chair driven by a hydraulic servo system. Ascent induced small responses in sternocleidomastoid muscles (SCMs) with a latency of 24.0 ms (SE 1.7) from onset of head acceleration, followed by larger responses of 100 ms in duration. In comparison the responses in SCMs evoked by descent were significantly delayed, with greater interindividual variations at 44.1 ms (SE 3.1) and smaller in amplitude than responses in ascent. Latencies were consistent and showed no habituation. In order to eliminate stretch components, we measured the neck responses to vertical acceleration with the head fixed by cervical collar. The latency of the fast response of SCM evoked by ascent was not significantly different from the latency of SCMs without the collar. These results may indicate that the fast responses of SCMs to sudden ascent may be composed of a vestibular-collic reflex for making the neck and head rigid for defense to sudden gravitational change.

Acceleration↗
PubMed2001

Covered Gianturco stent for tracheal stricture: application of polychlorovinylidene and polyurethane as covering materials.

The aim of this study was to assess the efficacy and safety of thin membranes of polychlorovinylidene (PCV) or polyurethane (PU) as covering materials for Gianturco stents in the treatment of severe tracheal stricture caused by intraluminal tumor. Manufactured Gianturco stents covered with PCV or PU membrane were used to treat six malignant and one benign tracheal stricture. The initial results, complications, clinical follow-up, bronchoscopic findings, and three autopsy microscopic examinations were reviewed. Informed consent was obtained after the nature of the treatment had been fully explained before every procedure. The stents successfully dilated the tracheal strictures, providing immediate relief of respiratory symptoms in all patients with no major complications. During the follow-up period, the covering materials prevented tumor ingrowth until death or intraluminal granuloma formation. Autopsies proved that no histological change occurs in the tracheal mucosa in response to the presence of PCV or PU; however, ulcer formation occurred in one patient and penetration of the stent struts into the tracheal wall in two. Bloody sputum with ulcer formation, minimal granuloma formation at the distal end of the stent, and abnormal bacterial load in the sputum were long-term complications. The Gianturco stent covered with PCV or PU membrane is a useful option as a palliative treatment for malignant and an emergent salvage for benign tracheal stricture, because both materials are thinner and less voluminous than the others. However, the indications for its use are limited to patients with poor prognoses, because hemoptysis, granuloma formation, and bacterial propagation remain problematic.

Aged↗
PubMed2001

Effects of carnitine on cardiac function after cardioplegic ischemia in neonatal rabbit hearts.

BACKGROUND: Ischemia immediately impairs myocardial fatty acid metabolism and reduces the concentration of carnitine which is an essential cofactor for fatty acid metabolism in the mitochondria. The purpose of this study was to investigate the effects of carnitine administration on recovery of cardiac function after cardioplegic ischemia in the neonatal heart where fatty acid metabolism is not a predominant source of adenosine triphosphate. METHODS: Isolated blood-perfused neonatal rabbit hearts underwent 3 hours of cold cardioplegic ischemia. The control group (n = 10) was reperfused with unmodified diluted blood. The carnitine group (n = 10) was reperfused with the blood containing 5 mM/L of carnitine. Before ischemia (base line) and after 15 and 30 minutes reperfusion, left ventricular (LV) function and LV compliance were measured using a intraventricular conductance catheter combined with an isovolumic balloon. Coronary blood flow was measured and myocardial oxygen consumption was calculated. RESULTS: Carnitine significantly improved not only LV systolic function but also LV diastolic function (p < 0.05) as well as LV compliance after ischemia. Coronary blood flow and myocardial oxygen consumption were significantly improved after ischemia in the carnitine group compared with the control group (p < 0.05). CONCLUSIONS: These results suggest that carnitine strikingly improves LV functional recovery and aerobic metabolism after cold cardioplegic arrest, and may improve cardiac performance in neonates after open heart surgery.

Animals↗
PubMed2001

Carnitine affects fatty acid metabolism after cardioplegic arrest in neonatal rabbit hearts.

BACKGROUND: Fatty acid (FA) metabolism and the contribution of carnitine to metabolism after cardioplegic arrest still remain unclear, especially in the neonatal heart where beta-oxidation is not a predominant source of adenosine triphosphate. METHODS: FA metabolism and the effects of carnitine administration were evaluated using a newborn (7-day-old) rabbit blood-perfused Langendorff model subjected to cold cardioplegic arrest. The hearts were divided into five groups; (1) perfused with unmodified diluted blood (n = 9), (2) subjected to 180 minutes of cold cardioplegic arrest and reperfused with the blood (n = 9), (3) subjected to the same ischemia and reperfused with the blood containing 40 microM/L (n = 9), (4) 0.5 mM/L (n = 5), and (5) 5 mM/L of carnitine (n = 5). During reperfusion, FA metabolism was assessed by iodine-123-labeled 15-(p-iodophenyl)-3-(R,S)-methylpentadecanoic acid, a fatty acid. The myocardial time-radioactivity curve was then determined and a mathematical compartment analysis of the external detection was used to elucidate FA metabolism in the cardiac myocyte. RESULTS: Cold cardioplegic arrest resulted in significantly impaired FA metabolism following reperfusion. Compartment analysis suggested that FA activation in the cytosol and beta-oxidation were impaired. Carnitine supplementation in groups 3 and 4 improved FA metabolism during reperfusion. In contrast, supplementation in group 5 had no beneficial effect on FA metabolism. CONCLUSIONS: These results suggest that FA metabolism is impaired after cold cardioplegic arrest and that carnitine supplementation may improve aerobic metabolism in neonates after open heart surgery.

Animals↗