Biomedical subjects

K Sakai

Publications and source records attributed to K Sakai.

At least 343 records · Page 19Linked to original sources

PubMed1998

Fluid shear stress attenuates tumor necrosis factor-alpha-induced tissue factor expression in cultured human endothelial cells.

Hemodynamic forces modulate various endothelial cell functions under gene regulation. Previously, we have shown that fibrinolytic activity of endothelial cells is enhanced by the synergistic effects of shear stress and cytokines. In this study, we investigated the effect of shear stress on tumor necrosis factor (TNF)-alpha-induced tissue factor (TF) expression in cultured human umbilical vein endothelial cells (HUVECs), using a modified cone-plate viscometer. Shear stresses at physiological levels reduced TNF-alpha (100 U/mL)-induced TF expression at both mRNA and antigen levels, in a shear-intensity and exposure-time dependent manner, whereas shear stress itself did not induce TF expression in HUVECs. TF expressed on the cell surfaces measured by flow cytometry using an anti-TF monoclonal antibody (HTF-K180) was also decreased to one third by shear force applied at 18 dynes/cm2 for 15 hours before and 6 hours after TNF-alpha stimulation. Furthermore, functional activity of TF, as assessed by the activation of factor X in the presence of FVIIa and Ca2+, was also decreased by shear application. However, the stability of TF mRNA was not decreased in the presence of shear stress. These results suggest that shear force acts as an important regulator of TF expression in endothelium at the transcriptional level.

Aspirin↗

PubMed1998

Allelic loss analysis of gamma-ray-induced mouse thymic lymphomas: two candidate tumor suppressor gene loci on chromosomes 12 and 16.

A total of 429 gamma-ray-induced thymic lymphomas were obtained from F1 and backcross mice between BALB/c and MSM strains, about a half of which carried a p53-deficient allele. A genome-wide allelic loss analysis has revealed two loci exhibiting frequent allelic losses but no allelic preference, one is localized within a 2.9 cM region between D12Mit53 and D12Mit279 loci on chromosome 12, and the other is near the D16Mit122/D16Mit162 loci on chromosome 16. The frequency of allelic loss in the D12Mit279 region is 62% and does not differ in tumors between the presence and absence of the p53-deficient allele. In contrast, the loss frequency of D16Mit122 is raised by the existence of p53-deficient allele: 62% for p63(-/+) and 13% for p53(+/+), suggesting co-operative function of the two losses. The D12Mit279 and D16Mit122 regions probably harbor different types of tumor suppressor gene that play key roles in lymphoma development.

Animals↗

PubMed1998

Construction and performance test of SGM-TRAIN at UVSOR.

A new spherical-grating monochromator with translational and rotational assembly including a normal-incidence mount (SGM-TRAIN) has been constructed at BL5A of the UVSOR facility. The SGM-TRAIN is an advanced version of a constant-length SGM with the following improvements: (i) a wide energy range of 5-250 eV; (ii) a high resolving power; (iii) use of linear and circular polarization; (iv) reduction of second-order light; (v) two computer-controlled driving modes. Part of the performance tests are reported along with a detailed description of the design.

Journal Article↗

PubMed1998

Regulation of human mast cell beta-tryptase: conversion of inactive monomer to active tetramer at acid pH.

At neutral pH, human mast cell beta-tryptase is stabilized in its enzymatically active, tetrameric form by heparin, and resists inhibition by biologic protease inhibitors. After dissociation of beta-tryptase from heparin, active tetramers rapidly convert to inactive monomers in an isotonic, neutral pH environment. Although reversible transition states probably exist during this conversion, once inactive monomers form, addition of heparin fails to reconstitute active tetramer at neutral pH. The current study shows that complete reactivation of inactive monomers can occur at acidic pH in a heparin-independent manner. The respective rate-determining steps for formation of tetramer and active enzyme from inactive monomers exhibit second and first order kinetics based on an analysis of initial reaction rates. The optimal pH for tetramer formation and reactivation is about 6, suggesting His residues play a critical role. The optimal ionic strength equivalent is 160 mM NaCl; and the optimal temperature range is 22 degrees C to 37 degrees C. We propose a sequential three-step reactivation process at acidic pH, dimerization of monomers (rate-determining second order step), rapid formation of inactive tetramers, and slow formation of active tetramers (overall rate-determining first order step). Whether reactivation of human beta-tryptase occurs at extracellular or intracellular sites, where the pH is acidic in vivo, should be considered.

Chymases↗

PubMed1998

Protein-bound acrolein: potential markers for oxidative stress.

Acrolein (CH2==CH---CHO) is known as a ubiquitous pollutant in the environment. Here we show that this notorious aldehyde is not just a pollutant, but also a lipid peroxidation product that could be ubiquitously generated in biological systems. Upon incubation with BSA, acrolein was rapidly incorporated into the protein and generated the protein-linked carbonyl derivative, a putative marker of oxidatively modified proteins under oxidative stress. To verify the presence of protein-bound acrolein in vivo, the mAb (mAb5F6) against the acrolein-modified keyhole limpet hemocyanin was raised. It was found that the acrolein-lysine adduct, Nepsilon-(3-formyl-3, 4-dehydropiperidino)lysine, constitutes an epitope of the antibody. Immunohistochemical analysis of atherosclerotic lesions from a human aorta demonstrated that antigenic materials recognized by mAb5F6 indeed constituted the lesions, in which intense positivity was associated primarily with macrophage-derived foam cells and the thickening neointima of arterial walls. The observations that (i) oxidative modification of low-density lipoprotein with Cu2+ generated the acrolein-low-density lipoprotein adducts and (ii) the iron-catalyzed oxidation of arachidonate in the presence of protein resulted in the formation of antigenic materials suggested that polyunsaturated fatty acids are sources of acrolein that cause the production of protein-bound acrolein. These data suggest that the protein-bound acrolein represents potential markers of oxidative stress and long-term damage to protein in aging, atherosclerosis, and diabetes.

Acrolein↗

PubMed1998

Reciprocal interactions among neuropeptides and adenosine in the cardiovascular system of rats: a role of K(ATP) channels.

Possible reciprocal interactions among vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP) and adenosine were investigated in anesthetized rats. Changes in arterial blood pressure were taken as a parameter to evaluate the interactions. The i.v. bolus injections of VIP (0.3 or 1 microg kg(-1)), CGRP (0.1 or 0.3 microg kg(-1)) and adenosine (1-100 microg kg(-1)), like acetylcholine (0.1 microg kg(-1)), produced reductions of blood pressure, accompanied by slight changes (less than 5% except for 100 microg kg(-1) adenosine) in heart rate (HR). The vasodepressor responses to VIP and CGRP were significantly augmented by i.v. infusion of adenosine (3 microg kg(-1) min(-1)). The vasodepressor responses to adenosine and CGRP by VIP (0.03 microg kg(-1) min(-1)), and those to adenosine and VIP by CGRP (1 ng kg(-1) min(-1)) were also enhanced. The response to acetylcholine remained unchanged before and during i.v. infusion of either VIP, CGRP or adenosine. The i.v. infusion of cromakalim (0.1 microg kg(-1) min(-1)) also augmented the responses to VIP, CGRP and adenosine, but not to acetylcholine, whereas a single bolus i.v. injection of glibenclamide (20 mg kg(-1)) significantly attenuated each one of them. The present results suggest that endogenous vasodilators, such as VIP, CGRP and adenosine, reciprocally interact in the body, at least partly through ATP-sensitive K+ channels.

Acetylcholine↗

PubMed1998

Quantification of concanavalin A binding to rat brain microsomal membranes detected by fluorescence polarization technique.

A high sensitive method for detecting the change of microsomal membrane surface oligosaccharides was developed to study the regulatory role of lipid- or peptide-linked mannoside of endoplasmic reticulum in synaptic functions. The binding of concanavalin A to the microsomal membrane surface was measured quantitatively using a microgram-order of rat brain microsomal proteins. The fluorescence polarization of concanavalin A (Con A)-fluorescein isothiocyanate (FITC) conjugate bound to the membrane was analyzed to quantitate the change of binding constant and the number of binding sites. As a control, the non-specific binding of bovine serum albumin-FITC conjugate was measured by the same technique. We measured the change of fluorescence intensity of membrane-bound FITC conjugates by the flow cytometry and found that the intensity of FITC conjugate bound to the membrane increased more than that of free form of the probe. We observed that the alpha-mannosidase-treatment of rat brain microsomes resulted in the increase of binding constant of Con A to the microsomal surface without significant loss of binding sites.

Animals↗

PubMed1998

Synergistic effect of calcitonin gene-related peptide on adenosine-induced vasodepression in rats.

The action of calcitonin gene-related peptide (CGRP) on the vasodepressor response to adenosine was investigated in anesthetized rats. I.v. bolus injections of adenosine (1-100 microg/kg), acetylcholine (0.05-0.4 microg/kg), isoproterenol (1-30 ng/kg), nitroglycerin (0.3-10 microg/kg) and diltiazem (10-300 microg/kg) produced dose-dependent decreases in blood pressure, accompanied by changes in heart rate. Only the vasodepressor response elicited by adenosine, among the agents tested, was significantly enhanced by i.v. infusion of either CGRP (1 ng/kg per min) or cromakalim (0.1 microg/kg per min), which possesses glibenclamide-sensitive K+ channel opening activity. After i.v. treatment with glibenclamide (20 mg/kg), the vasodepressor responses not only to adenosine but also to CGRP (0.5 microg/kg) and cromakalim (30 microg/kg) were significantly reduced, while those to acetylcholine and isoproterenol remained unchanged. The result indicates that the enhancement of the adenosine-induced vasodepression by CGRP, like that elicited by cromakalim, seems to be mediated at least partly through ATP-sensitive K+ channel activation.

Adenosine↗

PubMed1998

Localization of D-aspartic acid in elongate spermatids in rat testis.

In the current study, localization of D-aspartic acid (D-Asp) in rat testis was studied by immunohistochemical and biochemical techniques. Immunohistochemical staining of this tissue using specific polyclonal antibody to D-Asp revealed D-Asp immunoreactivity (IR) in the cytoplasm of germ cells, especially around the region rich in elongate spermatids, the most mature of the germ cells. Weak IR was also noted in cytoplasm of spermatocytes and round spermatids; however, it was negligible in interstitial cells and Sertoli cells. The intensity of immunostaining in each seminiferous tubule differed according to its distinct germ cell composition. In testis of young rats, seminiferous tubules lack elongated spermatids, and D-Asp was found to be localized in spermatocytes, the most mature population of germ cells at that age. We used various toxicants to destroy specific testicular cell populations and to confirm the localization of D-Asp in rat testis. Administration of ethane dimethane sulfonate induced a selective destruction of all Leydig cells in this tissue. This resulted in a significant decrease in the D-Asp level, which was probably due to a drop in testosterone brought about by this treatment, and this was followed by a modulation of spermatogenesis. Three days after treatment with methoxyacetic acid (MAA), many seminiferous tubules were found to lack or to have severe depletions of pachytene spermatocytes, but not of elongate spermatids. This caused reductions in protein content and in the total amount of L-Asp, but not that of D-Asp. Twenty days after treatment with MAA, the depleted population of germ cells progressed through the spermatogenic cycle from pachytene spermatocytes to elongate spermatids. At this time, the level of D-Asp decreased significantly, as did that of L-Asp and protein, consistent with D-Asp localization in elongate spermatids. This decrease in the D-Asp level was also seen with immunostaining.

Acetates↗

PubMed1998

Transition of brain activation from frontal to parietal areas in visuomotor sequence learning.

We studied the neural correlates of visuomotor sequence learning using functional magnetic resonance imaging (fMRI). In the test condition, subjects learned, by trial and error, the correct order of pressing two buttons consecutively for 10 pairs of buttons (2 x 10 task); in the control condition, they pressed buttons in any order. Comparison between the test condition and the control condition revealed four brain areas specifically related to learning: the dorsolateral prefrontal cortex (DLPFC), the presupplementary motor area (pre-SMA), the precuneus, and the intraparietal sulcus (IPS). We found that the time course of activation during learning was different between these areas. To normalize the individual differences in the speed of learning, we classified the performance of each subject into three learning stages: early, intermediate, and advanced stages. Both the relative increase of signal intensity and the number of activated pixels within the four areas showed significant changes across the learning stages, with different time courses. The two frontal areas, DLPFC and pre-SMA, were activated in the earlier stages of learning, whereas the two parietal areas, precuneus and IPS, were activated in the later stages. Specifically, DLPFC, pre-SMA, precuneus, and IPS were most highly activated in the early stage, in both the early and intermediate stages, in the intermediate stage, and in both the intermediate and advanced stages, respectively. The results suggest that the acquisition of visuomotor sequences requires frontal activation, whereas the retrieval of visuomotor sequences requires parietal activation, which might reflect the transition from the declarative stage to the procedural stage.

Adult↗

PubMed1998

Primary motor cortex isolation: complete paralysis with preserved primary motor cortex.

We present a left-sided hemiplegic patient with a cerebrovascular lesion involving the medial part of the right frontal and parietal lobes and the corpus callosum, but sparing the hand area of right primary motor cortex (M1). Several studies using transcranial magnetic stimulation demonstrated functional integrity of the efferent pathways from the right M1, intact sensory afferents to M1, an impairment of transcallosal connection between the bilateral motor cortices, and reduced ipsilateral cortico-cortical inhibition within the right M1. Based on these results, we conclude that the paralysis of this patient was caused by disconnection of the intact M1 from any structures requisite for initiation of movements. The present patient also suggests the importance of various afferents to M1 in voluntary movement. We propose a term of 'primary motor cortex isolation' to designate the paralysis reported here.

Electromyography↗

PubMed1998

A non-sense mutation at Arg95 is predominant in complement 9 deficiency in Japanese.

Deficiency of the ninth component of complement (C9D) is one of the most common genetic abnormalities in Japan, with an incidence of one homozygote in 1000. Although C9D individuals are usually healthy, it has been shown that they have an significantly increased risk of developing meningococcal meningitis. In the present study we report the molecular bases for C9D in 10 unrelated Japanese subjects. As a screening step for mutations, exons 2 to 11 of the C9 gene were analyzed using exon-specific PCR/single-strand conformation polymorphism analysis, which demonstrated aberrantly migrating DNA bands in exon 4 in all the C9D subjects. Subsequent direct sequencing of exon 4 of the C9D subjects revealed that eight of the 10 C9D subjects were homozygous for a C to T transition at nucleotide 343, the first nucleotide of the codon CGA for Arg95, leading to a TGA stop codon (R95X). R95X is a novel mutation different from those recently identified in a Swiss family with C9D. Cases 6 and 7 were heterozygous for the R95X mutation. Family study in case 10 confirmed the genetic nature of the defect. In case 6, the second mutation for C9D of the C9 gene was identified to be the substitution of Cys to Tyr at amino acid residue 507 (C507Y), while the genetic defect(s) in the other allele in case 7 remains unknown. Our results indicate that a novel mutation, R95X, is present in most cases of C9D in Japan.

Adult↗

PubMed1998

Potential withdrawal of rheumatoid synovium by the induction of apoptosis using a novel in vivo model of rheumatoid arthritis.

OBJECTIVE: To investigate whether Fas-mediated apoptosis has potential as a new therapeutic strategy in rheumatoid arthritis (RA) by use of a novel model of RA in which human RA tissue is grafted into SCID mice. METHODS: Fresh rheumatoid synovial tissue including joint cartilage was grafted subcutaneously into the backs of SCID mice. Six weeks after engraftment, anti-Fas monoclonal antibody was injected intraperitoneally. Time-related apoptotic changes caused by anti-Fas monoclonal antibody in grafted synovium were evaluated by nick end-labeling histochemistry. RESULTS: Thirty-six hours after the injection, diffuse apoptotic changes were observed in the grafted synovia. Four weeks after the injection, rheumatoid synovial tissue diminished. CONCLUSION: This is the first report concerning the present effectiveness of anti-Fas monoclonal antibody in diminishing rheumatoid synovium in vivo, and suggests the possibility of a new strategy for treating rheumatoid arthritis by inducing Fas-mediated apoptosis.

Animals↗

PubMed1998

High-Resolution Terahertz Spectroscopy by a Compact Radiation Source Based on Photomixing with Diode Lasers in a Photoconductive Antenna

We demonstrate generation of continuous-wave terahertz radiation and its application to molecular spectroscopy. The radiation source is based on the photomixing of two diode laser beams in a low-temperature-grown GaAs photoconductive antenna, which offered output power of several tens of nanowatts at frequencies up to 2 THz with its long term frequency stability being about 5 MHz. The pure rotational spectra of CH3CN and isotopomers of CO were measured with this source, and 1% of absorption was clearly detected with a simple amplitude modulation technique. This indicates that the present system has a potential capability for high-resolution and high-sensitivity molecular spectroscopy. Copyright 1998 Academic Press. Copyright 1998Academic Press

Journal Article↗

PubMed1998

Basilar bifurcation aneurysms associated with persistent primitive hypoglossal artery.

We report two cases of ruptured basilar bifurcation aneurysm associated with a persistent primitive hypoglossal artery. Angiograms revealed low-positioned aneurysms; in both cases bilateral vertebral arteries and posterior communicating arteries were hypoplastic or aplastic. Both aneurysms were successfully clipped via subtemporal transtentorial approach through the craniotomy ipsilateral to the side of the primitive hypoglossal artery. The ipsilateral craniotomy and exposure of the cervical carotid artery were helpful for obtaining the proximal control of the basilar artery needed to perform the clipping procedure with safety.

Adult↗

PubMed1998

Sinus histiocytosis with massive lymphadenopathy: a case of multiple dural involvement.

An exceptional case of Rosai-Dorfman disease (sinus histiocytosis with massive lymphadenopathy) arising from the meninges in a 60-year-old Japanese man is presented. Computerized tomographic scans and magnetic resonance images demonstrated well-circumscribed tumorous lesions that were homogeneously enhanced with contrast medium. Systemic examination revealed no abnormalities except for a cervical lymphadenopathy and diabetes mellitus. Microscopic examination of the resected specimens showed proliferated histiocytosis and infiltration of plasma cells and lymphocytes. The histology was characterized by the presence of histiocytes demonstrating lymphophagocytosis and immunoreactivity for S-100 protein staining. Immunohistochemical studies and electron microscopy were useful in confirming the diagnosis. The clinical and histopathological features of this disease are discussed.

Brain Diseases↗

PubMed1998

Cellular characterization of pituitary adenoma cell line (AtT20 cell) transfected with insulin, glucose transporter type 2 (GLUT2) and glucokinase genes: insulin secretion in response to physiological concentrations of glucose.

We investigated the mechanisms of insulin secretion by transfecting into a pituitary adenoma cell line (AtT20) a combination of genes encoding human insulin (HI), glucose transporter type 2 (GLUT2) and glucokinase (GK), followed by studying the characteristics of these cells. In static incubation, a cell line transfected with insulin gene alone (AtT20HI) secreted mature human insulin but this was not in a glucose-dependent manner. Other cell lines transfected with insulin and GLUT2 genes (AtT20HI-GLUT2-3) or with insulin and GK genes (AtT20HI-GK-1) secreted insulin in response to glucose concentrations of only less than 1 mmol/l. In contrast, cell lines transfected with insulin, GLUT2 and GK genes (AtT20HI-GLUT2-GK-6, AtT20HI-GLUT2-GK-7 and AtT20HI-GLUT2-GK-10) showed a glucose-dependent insulin secretion up to 25 mmol/l glucose. Glucose utilization and oxidation were increased in AtT20HI-GLUT2-GK cell lines but not in AtT20HI, AtT20HI-GLUT2-3 and AtT20HI-GK-1 cells at physiological glucose concentrations, compared with AtT20 cells. Diazoxide, nifedipine and 2-deoxy glucose suppressed (p < 0.05) glucose stimulated insulin secretion in AtT20HI-GLUT2-GK-6 cells. Glibenclamide, KCl or corticotropin releasing factor (CRF) stimulated (p < 0.05) insulin secretion both in AtT20HI and AtT20HI-GLUT2-GK-6 cells. Insulin secretion stimulated by glibenclamide, KCl or CRF was further enhanced by the addition of 25 mmol/l glucose in AtT20HI-GLUT2-GK-6 cells but not in AtT20HI cells. In perifusion experiments, a stepwise increase in glucose concentration from 5 to 25 mmol/l stimulated insulin secretion in AtT20HI-GLUT2-GK cell lines but the response lacked a clear first phase of insulin secretion. Our results suggest that both GLUT2 and glucokinase are necessary for the glucose stimulated insulin secretion in at least rodent cell lines, and that other element(s) are necessary for a biphasic insulin secretion typically observed in beta cells.

Adenoma↗

PubMed1998

Localizing the site of magnetic brain stimulation by functional MRI.

In order to locate the site of action of transcranial magnetic stimulation (TMS) within the human motor cortices, we investigated how the optimal positions for evoking motor responses over the scalp corresponded to the hand and leg primary-motor areas. TMS was delivered with a figure-8 shaped coil over each point of a grid system constructed on the skull surface, each separated by 1 cm, to find the optimal site for obtaining motor-evoked potentials (MEPs) in the contralateral first dorsal interosseous (FDI) and tibialis anterior (TA) muscles. Magnetic resonance imaging scans of the brain were taken for each subject with markers placed over these sites, the positions of which were projected onto the cortical region just beneath. On the other hand, cortical areas where blood flow increased during finger tapping or leg movements were identified on functional magnetic resonance images (fMRI), which should include the hand and leg primary-motor areas. The optimal location for eliciting MEPs in FDI, regardless of their latency, lay just above the bank of the precentral gyrus, which coincided with the activated region during finger tapping in fMRI studies. The direction of induced current preferentially eliciting MEPs with the shortest latency in each subject was nearly perpendicular to the course of the precentral gyrus at this position. The optimal site for evoking motor responses in TA was also located just above the activated area during leg movements identified within the anterior portion of the paracentral lobule. The results suggest that, for magnetic stimulation, activation occurs in the primary hand and leg motor area (Brodmann area 4), which is closest in distance to the optimal scalp position for evoking motor responses.

Brain Mapping↗