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Biomedical subjects

K Fukushima

Publications and source records attributed to K Fukushima.

At least 217 records · Page 12Linked to original sources
PubMed1997

A single-point mutation in HCF causes temperature-sensitive cell-cycle arrest and disrupts VP16 function.

The temperature-sensitive BHK21 hamster cell line tsBN67 ceases to proliferate at the nonpermissive temperature after a lag of one to a few cell divisions, and the arrested cells display a gene expression pattern similar to that of serum-starved cells. The temperature-sensitive phenotype is reversible and results from a single missense mutation--proline to serine at position 134--in HCF, a cellular protein that, together with the viral protein VP16, activates transcription of herpes simplex virus (HSV) immediate-early genes. The tsBN67 HCF mutation also prevents VP16 activation of transcription at the nonpermissive temperature. The finding that the same point mutation in HCF disrupts both VP16 function and the cell cycle suggests that HCF plays a role in cell-cycle progression in addition to VP16-dependent transcription.

Amino Acid Sequence↗
PubMed1997

A partial deficiency of dehydrodolichol reduction is a cause of carbohydrate-deficient glycoprotein syndrome type I.

Carbohydrate-deficient glycoprotein (CDG) syndrome type I is a congenital disorder that involves the underglycosylation of N-glycosylated glycoproteins (Yamashita, K., Ideo, H., Ohkura, T., Fukushima, K., Yuasa, I., Ohno, K., and Takeshita, K. (1993) J. Biol. Chem. 268, 5783-5789). In an effort to further elucidate the biochemical basis of CDG syndrome type I in our patients, we investigated the defect in the multi-step pathway for biosynthesis of lipid-linked oligosaccharides (LLO) by the metabolic labeling method using [3H]glucosamine, [3H]mannose, and [3H]mevalonate. The LLO levels in synchronized cultures of fibroblasts from these patients were severalfold lower than those in control fibroblasts in the S phase, and the oligosaccharides released from LLO showed the same structural composition, Glc1 approximately 3.Man9.GlcNAc.GlcNAc, in the case of both the patients and controls. The amount of [3H]mannose incorporated into mannose 6-phosphate, mannose 1-phosphate, and GDP-mannose was greater in fibroblasts from these patients than in the control fibroblasts in the G1 period, although the ratios of these acidic mannose derivatives as indicated by the relative levels of radioactivity were the same for the two types of fibroblasts. Furthermore, upon metabolic labeling with [3H]mevalonate, the level of [3H]dehydrodolichol in fibroblasts from these patients increased in the S phase, and the levels of [3H]dolichol and [3H]dolichol-PP oligosaccharides concomitantly decreased, although the chain length distribution of the respective dolichols and dehydrodolichols was the same in the two types of fibroblasts. These results indicate that the conversion of dehydrodolichol to dolichol is partially defective in our patients and that the resulting loss of dolichol leads directly to underglycosylation.

Cells, Cultured↗
PubMed1997

Mechanistic studies on metabolic chiral inversion of 4-(4-methylphenyl)-2-methylthiomethyl-4-oxobutanoic acid (KE-748), an active metabolite of the new anti-rheumatic agent 2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298), in rats.

The chiral inversion properties of 4-(4-methylphenyl)-2-methylthiomethyl-4-oxobutanoic acid (KE-748), an active metabolite of 2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298), were compared with those of ibuprofen in rats. After administration of R(-)-[2 alpha-2H]KE-748, S(+)-KE-748 was present in the rat plasma, and the deuterium atoms of the S(+)-enantiomer were almost all replaced by hydrogen atoms. After administration of S(+)-[2 alpha-2H]KE-748, the deuterium content of S(+)-KE-748 in the plasma remained intact. In the in vitro study, using a cell-free system and rat liver homogenates, the chiral inversion of ibuprofen was apparent when both CoA and ATP were present; however, KE-748 was not inverted. In the study on isolated rat hepatocytes, the unidirectional chiral inversion from R(-)-to S(+)-enantiomer was observed for both ibuprofen and KE-748. When R(-)-ibuprofen was incubated with medium and long chain fatty acids (carbon chain length C6 to C16), using isolated hepatocytes, the chiral inversion decreased significantly. On the other hand, when R(-)-KE-748 was incubated with short and medium chain fatty acids (carbon chain length C3 to C8), chiral inversion was inhibited markedly. To induce hepatic microsomal long chain fatty acid CoA ligase, rats were treated with clofibric acid (CF rats). In both in vitro and in vivo experiments on CF rats, chiral inversion from R(-)-to S(+)-ibuprofen was enhanced significantly compared with that in controls, whereas the enhancement was not observed in the case of R(-)-KE-748. There was no influence of benzoic acid, a typical substrate on medium chain fatty acid CoA ligase in the mitochondrial matrix, on chiral inversion of R(-)-ibuprofen, using, isolated hepatocytes. In contrast, the chiral inversion from R(-)-to S(+)-KE-748 was strongly inhibited in the presence of benzoic acid. These results indicate that chiral inversion of R(-)-KE-748 may proceed via formation of the CoA-thioester intermediate with loss of the 2 alpha-methine proton, in a manner similar to that seem with R(-)-ibuprofen. However, the enzymes needed to form CoA-thioester of R(-)-KE-748 differ from those for R(-)-ibuprofen.

Animals↗
PubMed1997

c-fos antisense blocks methamphetamine-induced ambulatory activity reversibly.

To investigate the involvement of immediate early genes (IEGs) in the ambulatory activity induced by a low dose of methamphetamine (MAP) in mice, we injected an antisense phosphorothioate oligodeoxynucleotide (ODN) to c-fos mRNA stereotactically into the striatum of ddY male mice. MAP (2 mg kg-1)-induced ambulatory activity was significantly suppressed when the mouse was pretreated by the ODN (2 mM, 1 microliter), and this was accompanied by a decrease in c-Fos-like protein immunoreactivity. Repetitive intermittent administration of MAP at intervals of 3 days confirmed that the suppressive effect was transient and reproducible. These results suggest that c-Fos is at least partly the key molecule regulating the psychomotor stimulation by MAP in mouse brain.

Animals↗
PubMed1997

Familial limb-girdle myasthenia with tubular aggregates.

Two sisters developed slowly progressive limb-girdle weakness in their childhood. The weakness responded to acetylcholinesterase inhibitors. Repetitive nerve stimulation showed decremental responses and single-fiber electromyography demonstrated increased jitter and blocking. Needle electromyography revealed myopathic changes. Antiacetylcholine receptor antibodies were negative. Histologic examinations demonstrated myopathy with tubular aggregates in the muscle fibers while the neuromuscular junctions appeared normal. They were diagnosed with familial limb-girdle myasthenia. This is the first report of this syndrome with morphologic studies of neuromuscular junctions.

Female↗
PubMed1997

Stereoselective disposition and chiral inversion of KE-298, a new antirheumatic drug, in rats.

The present study was an attempt to elucidate the relationship between stereoselective pharmacokinetics and protein binding of KE-298 and its active metabolites, deacetyl-KE-298 (M-1) and S-methyl-KE-298 (M-2). Metabolic chiral inversion was also investigated. The levels of unchanged KE-298 in plasma after oral administration of (+)-(S)-KE-298 to rats were lower than those of (-)-(R)-KE-298, whereas the levels of M-1 and M-2 after administration of (+)-(S)-KE-298 were higher than after (-)-(R)-KE-298. In vitro, rat plasma protein binding of (+)-(S)-KE-298 was lower than that of (-)-(R)-KE-298. In contrast, the binding of (+)-(S)-M-1 and (+)-(S)-M-2 was higher than that of (-)-(R)-M-1 and (-)-(R)-M-2. Displacement studies revealed that the (+)-(S) and (-)-(R)- enantiomers of KE-298 and their metabolites bound to the warfarin binding site on rat serum albumin. These results suggested that the stereoselective plasma levels in KE-298 and its metabolites were closely related to enantiomeric differences in protein binding attributed to quantitative differences in binding to albumin rather than to the different binding sites. Unidirectional chiral inversion was detected after oral administration of either (-)-(R)-KE-298 or (-)-(R)-M-2 to rats both yielding (+)-(S)-M-2.

Animals↗
PubMed1997

B cell subpopulations separated by CD27 and crucial collaboration of CD27+ B cells and helper T cells in immunoglobulin production.

B cell immunoglobulin production is regulated by helper T cells through direct interaction and secreted cytokines. In the present study, we functionally analyzed CD27 in cord and peripheral blood B cells. Adult peripheral blood B cells were separated into CD27+ and CD27- cells, which differed in their morphology. Cord blood B cells did not express CD27, and CD27 expression on peripheral blood B cells increased with age. Only CD27+ B cells had the ability to produce immunoglobulin, which was increased by contact with a tumor necrosis factor-related transmembrane ligand, CD70. Adult peripheral blood CD27+ B cells can be further subdivided into two discrete subtypes: IgD- CD27+ and IgD+ CD27+ B cells. IgD- CD27+ B cells produce IgG, IgM and IgA, whereas IgD+ CD27+ B cells predominantly produce IgM. The addition of activated CD4+ CD45RO T cells expressing CD70 caused down-regulation of CD27 expression on activated B cells, and this down-modulation was completely blocked by anti-CD70 monoclonal antibody, indicating direct T-B cell contact via CD27/CD70. The triggering via CD27 and CD40 additively increased the immunoglobulin production under Staphylococcus aureus Cowan strain plus interleukin-2 stimulation. Taken together, our findings demonstrate that peripheral blood B cells are separated into subpopulations by CD27 and IgD expression and that CD27+ B cells produce large amounts of immunoglobulin by interaction with the CD70 molecule.

Adult↗
PubMed1997

Cholesterol and bile acid metabolism after selective portal vein ligation.

AIM: To examine the regulatory effect of bile acid level on bile acid synthesis in the liver. METHODS: The portal branch perfusing left lateral and median lobes of the liver was ligated in rats and the activities of hepatic microsomal cholesterol 7 alpha-hydroxylase, the rate-limiting enzyme of bile acid synthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, and intrahepatic concentrations of cholesterol and bile acids were determined in the liver lobes deprived of and supplied with portal blood on Days 0, 1, 2, 4, and 7 after selective portal vein ligation (SPVL). RESULTS: In the portal vein (PV)-ligated lobes, liver weight decreased, hepatic cholesterol concentration was unchanged, and microsomal cholesterol concentration increased after SPVL. In the PV-nonligated lobes, liver weight increased, hepatic cholesterol concentration increased, and microsomal cholesterol concentration was unchanged. There were no significant differences in the activities of HMG-CoA reductase and cholesterol 7 alpha-hydroxylase among the PV-ligated and PV-nonligated lobes and the sham-operated controls. Intrahepatic bile acid level increased significantly in the PV-nonligated lobes for 4 days after SPVL, whereas those were essentially constant in the PV-ligated and the sham-operated control liver. Despite significant changes in the concentrations of intrahepatic cholesterol and bile acid, no significant correlations were observed between these concentrations and the activities of cholesterol 7 alpha-hydroxylase. CONCLUSIONS: SPVL causes atrophy and hypertrophy of the PV-ligated and nonligated liver lobes, respectively, without any significant changes in cholesterol and bile acid synthesis. Intrahepatic concentrations of bile acids and cholesterol have no regulatory effect on cholesterol 7 alpha-hydroxylase activity in the SPVL rat model.

Animals↗
PubMed1997

Corticovestibular interactions: anatomy, electrophysiology, and functional considerations.

This review summarizes anatomical and electrophysiological observations related to corticovestibular interactions as a step toward understanding their possible functions. Vestibular information is represented in at least three distinct regions of the cerebral cortex in cats and monkeys: the parietal and somatosensory cortex and the parietoinsular vestibular cortex. In addition, vestibular-related signals are found in more extensive regions, including the motor and premotor regions and frontal eye fields. Most of these regions also project directly to the vestibular nuclei. In monkeys, at least six cortical regions have been identified, including the motor, somatosensory, parietal and temporal areas. Most of these regions receive vestibular projections via the thalamus. Most neurons in those cortical areas respond to head velocity and receive converging vestibular, visual and somatosensory input. Electrical stimulation of some of these cortical areas in anesthetized cats influences the activity of many vestibular nuclear neurons including those projecting to the spinal cord. Lesions of the parietal vestibular regions impair the vestibulo-ocular reflex (VOR) and visual suppression of the VOR as well as vestibular-related cognitive functions such as spatial perception and memory in human subjects. Diffuse cortical damage also results in similar impairment of the VOR and suppression of the VOR and possibly the vestibulo-collic reflex. Such impairments after cortical lesions may well be due in part to interruption of cortico-vestibular connections. Future studies in alert animals should focus on the role of different cortical regions projecting to the vestibular nuclei, specifically on how each affects the processing of vestibular signals that mediate vestibulo-motor reflexes and that are used for vestibular related cognitive processes.

Animals↗
PubMed1997

Technetium-99m methoxyisobutylisonitrile single-photon emission tomography in hepatocellular carcinoma.

Nine lesions in eight patients with hepatocellular carcinoma (HCC) were studied using single-photon emission tomography (SPET) and technetium-99m methoxyisobutylisonitrile (99mTc-MIBI) to evaluate the pattern of uptake of 99mTc-MIBI in the lesions and the relation between the uptake pattern and the histopathology of HCC. All the lesions were diagnosed as HCC by percutaneous needle biopsy. Four of the nine lesions showed positive uptake of 99mTc-MIBI, while the other five showed negative uptake. All of the lesions which showed positive uptake were of the compact type. Of the five lesions that showed negative uptake, four were of the trabecular type while one was of the compact type. These results suggest that the patterns of 99mTc-MIBI accumulation in HCC are divided into positive and negative types and that these uptake patterns are associated with the tissue structure of HCC.

Aged↗
PubMed1997

Production of glucosyltransferases by clinical mutans streptococcal isolates as determined by semiquantitative cross-dot assay.

Forty-four clinical isolates of mutans streptococci were examined by a semiquantitative cross-dot assay for in vitro production of glucosyltransferases GTF-I, GTF-SI and GTF-S of Streptococcus mutans, and GTF-I of Strep. sobrinus, using monospecific antibodies. The isolates were obtained from 12 1.5- to 3-year old children, six caries-active and six caries-free, and from their mothers. The isolates were selected originally from 243 isolates and they represented 35 genetically distinct types as analysed by serotyping and ribotyping. 27 isolates were of serotype c, nine of serotype e and eight of serotype g. Mother child pairs shared nine ribotypes, suggesting vertical transmission. The results showed that, when cultured in Todd-Hewitt broth supplemented with 1% glucose, all Strep. mutans isolates produced GTF-I and GTF-S and all except two produced GTF-SI of Strep. mutans. All Strep. sobrinus isolates produced GTF-I of Strep. sobrinus. The Strep. mutans GTF-I, GTF-SI and GTF-S production of isolates exhibiting a different ribotype showed variability. The variability of GTF-SI and GTF-S production was less pronounced for serotype e isolates. The GTF-I production by Strep. sobrinus isolates did not vary. Transmitted strains produced the same levels of GTFs as strains that were distinct (not transmitted). Strep. mutans isolates of caries-active children produced the same levels of GTF-I and GTF-S, but tended to produce lower levels of GTF-SI than isolates of caries-free children. In conclusion, the results suggested that Strep. mutans isolates exhibiting a different ribotype often had differences in production of GTFs. However, no clear superiority of the high-producer over the low-producer strains was found in regard to their colonization or caries promotion in young children.

Adult↗
PubMed1997

Slow eye movement evoked by sudden appearance of a stationary visual stimulus observed in a step-ramp smooth pursuit task in monkey.

We examined the influence of a peripheral visual stimulus on eye movement while monkeys performed a horizontal step-ramp pursuit task. When an irrelevant visual stimulus was presented before the onset of the target motion, slow eye movement away from the stimulus was observed. When the stimulus appeared during a temporal gap between the offset of the fixation point and the onset of target motion the velocity of the slow eye movement increased. The onset of the movement was time-locked to the onset of the extraneous visual stimulus and its latency was comparable to the latency of smooth pursuit in trials without a peripheral stimulus. The results demonstrate a new form of smooth eye movement, suggesting that the eye movement may be contained in the initial stages of smooth pursuit observed in step-ramp paradigms.

Animals↗
PubMed1997

A human urinary protease inhibitor (ulinastatin) inhibits neutrophil extracellular release of elastase during cardiopulmonary bypass.

OBJECTIVES: To determine the benefits of a human urinary protease inhibitor (ulinastatin) on postoperative pulmonary dysfunction associated with neutrophil activation during cardiopulmonary bypass. DESIGN: A prospective, randomized, clinical study. SETTING: The study was performed at Keio University Hospital, Tokyo. PARTICIPANTS: Eighteen adult patients scheduled for primary cardiac surgery. INTERVENTIONS: The patients were randomly assigned either to the control group (n = 8) or to the group (n = 10) receiving ulinastatin (600,000 U in total). MEASUREMENTS AND MAIN RESULTS: Human neutrophil ability to release elastase in response to N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP) in vitro was measured before and after cardiopulmonary bypass, together with plasma levels of neutrophil elastase complex, interleukin-B, and C3a. Intrapulmonary shunt fraction was then calculated. Neutrophil elastase release in response to fMLP significantly increased in the control group, but remained unchanged in the ulinastatin group. In addition, ulinastatin minimized the increase of plasma neutrophil elastase, independently of the production of interleukin-B or C3a. Simultaneously, ulinastatin ameliorated the increase of intrapulmonary shunt, which was correlated with extracellular elastase release. CONCLUSIONS: Ulinastatin attenuated the elevation of fMLP-induced elastase release, which was associated with the deterioration of gas exchange during cardiopulmonary bypass. The administration of this agent has a potential to lessen the risk of postperfusion lung injury.

Adult↗
PubMed1997

Antigenic similarities to Paracoccidioides brasiliensis in thermo-dependent dimorphic fungi isolated from soil in Botucatu, SP, Brazil.

We compared the antigenic characteristics of two thermo-dependent dimorphic fungi isolated from soil in Botucatu, an endemic area of paracoccidioidomycosis (PCM) and Paracoccidioides brasiliensis. The soil isolates grew as cerebriform colonies at 37 degrees C (yeast form) and as cottonous colonies at 25 degrees C (mycelial form). No pathogenicity for ddY mice or hamsters were observed. In immunodiffusion test, there were precipitation bands between the 2 soil isolates and pooled PCM patient sera. There were also common precipitation bands at 21, 50 and 58 kDa between the soil isolates antigens and PCM patient sera by Western-blotting, but no gp43 kDa band. No gene for gp 43 kDa protein was detected in the soil isolates by PCR. The fact that these isolates were obtained from an endemic area of PCM and there were some antigenic similarities between the soil isolates and P. brasiliensis in immunodiffusion test and Western-blotting may have some importance in epidemiological surveys done with paracoccidioidin as well interfering with the immune response of the exposed population.

Animals↗
PubMed1997

The production of IL-8 in cerebrospinal fluid in aseptic meningitis of children.

Neutrophils accumulate initially in the cerebrospinal fluid (CSF) of aseptic meningitis, perhaps because of increased levels of granulocyte colony-stimulating factor (G-CSF), macrophage inflammatory protein-1alpha (MIP-1alpha), and IL-8 in the subarachnoid space. We studied levels of these cytokines in children with aseptic meningitis using ELISA. When meningeal symptoms existed, IL-8 levels (1399 +/- 1600 ng/l, n = 32) in the CSF were significantly higher than those either after meningeal symptoms disappeared (61 +/- 56 ng/l, n = 18) or in controls (44 +/- 63 ng/l, n = 27) (P < 0.0001). High levels of IL-8 on admission dropped sequentially. Significant correlations were found between IL-8 levels and either neutrophil counts (r = 0.612), G-CSF levels (r = 0.873) or MIP-1alpha levels (r = 0.623) in the CSF of the affected patients (P < 0.0001). IL-8 values in serum were lower than in the corresponding CSF samples from all individuals with meningeal symptoms. The IL-8 mRNA was detectable by reverse-transcribed polymerase chain reaction (PCR)-assisted amplification in fresh leucocytes from the CSF, but not from the peripheral blood of a healthy volunteer. The culture of CSF mononuclear cells produced high levels of IL-8 (approximately 2750 ng/l). These data indicate that IL-8 levels rise transiently at the initial stage of aseptic meningitis, and that mononuclear cells that migrate into the CSF are a cellular source of this chemokine. We suppose that IL-8, in addition to G-CSF and MIP-1alpha, contribute to the localized neutrophil accumulation during the disease.

Adolescent↗
PubMed1997

The highly conserved DAD1 protein involved in apoptosis is required for N-linked glycosylation.

BACKGROUND: The tsBN7 cell line is one of the temperature-sensitive mutants for cell proliferation which have been isolated from the BHK21 cell line derived from the golden hamster. It has a mutation in the DAD1 gene encoding a 12.5kDa highly conserved protein through evolution, and enters apoptosis at the restrictive temperature due to this mutation. RESULTS: DAD1 was recovered in light membrane fractions after differential centrifugation. It could not be released from the membrane, even by carbonate extraction, without a detergent. Upon digestion with proteinase K, both N and C terminal portions-but not the middle portions of DAD1- were released from the membrane. Thus, DAD1 appears to be an integral membrane protein in which both termini are located in the cytosol. DAD1 was localized in the endoplasmic reticulum. In accordance with a similarity to the yeast protein Ost2p, which is a subunit of the oligosaccharyltransferase, at the restrictive temperature, loss of DAD1 function caused a defect of N-linked glycosylation in tsBN7 cells resulting in apoptosis. However, tunicamycin, which is known to inhibit N-linked glycosylation did not induce apoptosis in either tsBN7 or BHK21 cells. CONCLUSION: tsBN7 cells have a defect in N-linked glycosylation caused by the loss of DAD1.

Animals↗
PubMed1997

Synthesis of lipid-linked oligosaccharides is dependent on the cell cycle in rat 3Y1 cells.

Synchronized cultures of rat 3Y1 cells, prepared by the density-arrested method, were used to investigate the relationship of N-glycosylation to the cell cycle. Although total cellular proteins were synthesized independently of the cell cycle, the synthesis of membrane-bound proteins was found to be dependent on the cell cycle, the time of maximum synthesis being just before that of maximum DNA synthesis. The synthesis of lipid-linked oligosaccharides (LLO), which are utilized as intermediates in N-glycosylation, increased in the S phase to a level at least ten times higher than in the G1, G2, and M periods. Moreover, the activities of dehydrodolichyl diphosphate synthase and farnesyl diphosphate synthase, which synthesize the precursors of dolichol, also increased in the S phase concomitantly with LLO synthesis. However, since the cell cycle dependency curves of these two enzyme activities were somewhat broader than that of LLO synthesis, the rate-limiting enzyme in the regulation of LLO synthesis might be another one such as dehydrodolichol reductase. These results suggest that LLO synthesis is regulated by multiple synthetic enzymes which are activated in a cell cycle dependent manner.

Animals↗