Biomedical subjects

M Fukuda

Publications and source records attributed to M Fukuda.

At least 289 records · Page 16Linked to original sources

PubMed1999

Dual roles of sialyl Lewis X oligosaccharides in tumor metastasis and rejection by natural killer cells.

Aberrant expression of cell surface carbohydrates such as sialyl Lewis X is associated with tumor formation and metastasis. In order to determine the roles of sialyl Lewis X in tumor metastasis, mouse melanoma B16-F1 cells were stably transfected with alpha1, 3-fucosyltransferase III to express sialyl Lewis X structures. The transfected B16-F1 cells, B16-FTIII, were separated by cell sorting into three different groups based on the expression levels of sialyl Lewis X. When these transfected cells were injected into tail veins of C57BL/6 mice, B16-FTIII.M cells expressing moderate amounts of sialyl Lewis X in poly-N-acetyllactosamines produced large numbers of lung tumor nodules. Surprisingly, B16-FTIII.H cells expressing the highest amount of sialyl Lewis X in shorter N-glycans died in lung blood vessels, producing as few lung nodules as B16-FTIII.N cells which lack sialyl Lewis X. In contrast, B16-FIII.H cells formed more tumors in beige mice and NK cell-depleted C57BL/6 mice than did B16-FTIII.M cells. B16-FTIII.H cells bound to E-selectin better than did B16-FTIII.M cells, but both cells grew at the same rate. These results indicate that excessive expression of sialyl Lewis X in tumor cells leads to rejection by NK cells rather than tumor formation facilitated by attachment to endothelial cells.

Animals↗

PubMed1999

Direct interaction between a quinoline derivative, MS-209, and multidrug resistance protein (MRP) in human gastric cancer cells.

MS-209 is a novel quinoline derivative reversing P-glycoprotein-mediated multidrug resistance (MDR). We investigated the interaction between MS-209 and multidrug resistance protein (MRP) in MRP-overexpressing human gastric cancer cells. We measured [3H]leukotriene C4 uptake into the membrane vesicles of the cells and intracellular calcein and [3H]vincristine accumulation with or without MS-209. In multi-drug-resistant MKN45R0.8 cells selected by doxorubicin, MS-209 dose dependently reduced MRP-mediated [3H]leukotriene C4 uptake and increased calcein accumulation. In both resistant and unselected cell lines expressing the MRP gene, MS-209 increased [3H]vincristine accumulation in proportion with the level of MRP mRNA expression and enhanced the cytotoxicity of etoposide, doxorubicin, and vincristine. The reversal effects correlated with the level of MRP mRNA expression in these cells. Our results indicate that MS-209 effectively reverses intrinsic and acquired MRP-mediated MDR of gastric cancer cells by interacting directly with MRP.

ATP-Binding Cassette Transporters↗

PubMed1999

Overexpression of nucleoside diphosphate kinases induces neurite outgrowth and their substitution to inactive forms leads to suppression of nerve growth factor- and dibutyryl cyclic AMP-induced effects in PC12D cells.

Whether nucleoside diphosphate kinase (NDPK) is involved in neuronal differentiation was investigated with special reference to its enzyme activity. Neurite outgrowth of PC12D cells induced by nerve growth factor or a cyclic AMP analog was suppressed to some extent when inactive NDPKs (the active site histidine 118 was replaced with alanine), not active forms, were transiently overexpressed. This suppression was more definite in their stably expressed clones. NDPKbeta-transfected clones and, to a lesser extent, NDPKalpha-transfected clones, but not inactive NDPK-transfected clones, extended neurites without differentiation inducers. These results imply that NDPKs may play a role by exerting their enzyme activity during differentiation of PC12 cells.

Alanine↗

PubMed1999

A novel endogenous inhibitor of phenoloxidase from Musca domestica has a cystine motif commonly found in snail and spider toxins.

Phenoloxidase inhibitor (POI), found in the hemolymph of housefly pupae, is a novel dopa-containing and cystine-rich peptide that competitively inhibits phenoloxidase with a Ki in the nanomolar range. [Tyr32]POI is a potential precursor molecule also found in the hemolymph that may be posttranslationally oxidized to the dopa-containing peptide after creation of a rigid structure. By employing both a solid-phase peptide synthesis system based on a 9-fluorenylmethoxycarbonyl strategy and a specific air oxidation technique to ensure correct folding, we have been able to synthesize [Tyr32]POI. The synthetic [Tyr32]POI was confirmed to be identical to the native [Tyr32]POI by coelution high-performance liquid chromatography analysis and by enzymatic analysis using the phenoloxidase inhibition assay. To determine the disulfide pairings within the peptides, a series of enzyme hydrolyses and partial reduction/alkylation steps were performed. Three cystine pairs (Cys11-Cys25, Cys18-Cys29, and Cys24-Cys36) were determined by identification of the resulting peptides. The disulfide pairings of the two adjacent Cys residues (Cys11-Cys25 and Cys24-Cys36) were unambiguously assigned by comparing the derived fragments with the two possible isomers synthesized through a novel disulfide-linking technique. The arrangement of the disulfide bridges in POI was found to be topologically identical to those found for several peptides within the inhibitor cystine knot structural family. Although these peptides share a low primary sequence homology and display a diversity of biological functions, they nonetheless share similarities in their cystine motifs and tertiary structure. The tertiary structure model of POI, which was derived through molecular dynamics and energy minimization studies using restraints with determined disulfide connectivities, suggests that POI is a new class member of the inhibitor cystine-knot structural family.

Amino Acid Sequence↗

PubMed1999

Patients with temporal lobe epilepsy show an increase in brain-derived neurotrophic factor protein and its correlation with neuropeptide Y.

Model studies on animal seizures have proposed potential involvement of the neurotrophins, BDNF and NGF, in human epilepsy. However, their biological significance in this disease itself remains to be evaluated. Here we demonstrate that patients with intractable temporal lobe epilepsy show a marked increase in protein levels of BDNF (2.6-fold, p<0.01) but not other neurotrophins. Moreover, the specific BDNF increase was significantly correlated with contents of neuropeptide Y. Thus, these results indicate the activity-dependent expression of BDNF in human subjects and its potential contribution to the pathophysiology of human epilepsy via neuropeptide Y.

Adolescent↗

PubMed1999

Molecular cloning and expression of a novel beta-1, 6-N-acetylglucosaminyltransferase that forms core 2, core 4, and I branches.

Mucin-type O-glycans are classified according to their core structures. Among them, cores 2 and 4 are important for having N-acetyllactosamine side chains, which can be further modified to express various functional oligosaccharides. Previously, we discovered by cloning cDNAs that the core 2 branching enzyme, termed core 2 beta-1,6-N-acetylglucosaminyltransferase-leukocyte type (C2GnT-L), is highly homologous to the I branching beta-1, 6-N-acetylglucosaminyltransferase (IGnT) (Bierhuizen, M. F. A., Mattei, M.-G., and Fukuda, M. (1993) Genes Dev. 7, 468-478). Using these homologous sequences as probes, we identified an expressed sequence tag in dbEST, which has significant homology to C2GnT-L and IGnT. This approach, together with 5'and 3' rapid amplification of cDNA ends, yielded a human cDNA that encompasses a whole coding region of an enzyme, termed C2GnT-mucin type (C2GnT-M). C2GnT-M has 48.2 and 33.8% identity with C2GnT-L and IGnT at the amino acid levels. The expression of C2GnT-M cDNA directed the expression of core 2 branched oligosaccharides and I antigen on the cell surface. Moreover, a soluble chimeric C2GnT-M had core 4 branching activity in addition to core 2 and I branching activities. A soluble chimeric C2GnT-L, in contrast, almost exclusively contains core 2 branching activity. Northern blot analysis demonstrated that the C2GnT-M transcripts are heavily expressed in colon, small intestine, trachea, and stomach, where mucin is produced. In contrast, the transcripts of C2GnT-L were more widely detected, including the lymph node and bone marrow. These results indicate that the newly cloned C2GnT-M plays a critical role in O-glycan synthesis in mucins and might have distinctly different roles in oligosaccharide ligand formation compared with C2GnT-L.

Amino Acid Sequence↗

PubMed1999

Mismatch negativity and N2b attenuation as an indicator for dysfunction of the preattentive and controlled processing for deviance detection in schizophrenia: a topographic event-related potential study.

The present study compares the amplitudes and topographic patterns of mismatch negativity (MMN) and N2b in schizophrenic patients and normal controls. Twenty-one schizophrenic outpatients and 19 normal volunteers participated in the study. Event-related potentials (ERPs) were recorded in a selective attention task. During the task, subjects were required to focus on one ear, counting deviant stimuli, those deviating in duration from a sequence of standard stimuli. MMN was significantly attenuated in the schizophrenics as compared with the normals in the frontocentral regions. In addition to MMN, the N2b amplitude was also reduced, which showed a significant correlation with the MMN amplitude in the schizophrenics. The late negativity elicited by the deviant stimuli in the unattended condition showed different topographical features between the groups. Whereas the normals showed a lateralized distribution with an ear-related asymmetry, similar to that of the N2b, the schizophrenics showed a frontal dominance, coinciding with the sustained negativity reported by Näätänen et al. (1982), which reflects the automatic preparation for detecting possible subsequent stimulus changes. The amplitude of the sustained negativity was significantly correlated with the performance level in the schizophrenics. The results indicated that although both preattentive and controlled processings are impaired, schizophrenic patients, presumably due to the deficient controlled processing, owe much to automatic processing in the deviant stimulus detection process.

Acoustic Stimulation↗

PubMed1999

Apoptosis in breast cancer and its relationship to clinicopathological characteristics and prognosis.

BACKGROUND AND OBJECTIVES: Apoptosis is essential to maintain homeostasis in living organisms and occurs in a variety of tissues in response to both physiological and pathological stimuli. In breast cancer, most cytotoxic drugs and hormonal treatments induce apoptosis. We studied the relationships between apoptosis and clinicopathological variables or prognosis in 143 patients with operable breast cancer. METHODS: Apoptosis was numerically graded in 5 consecutive 40x high-power fields (HPF) of hematoxylin-eosin (H&E) stained sections, since we showed that there was a significant correlation of H&E staining with terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining. RESULTS: The average number of apoptotic cells was 19.9 (0 approximately 168)/5 HPF, and cases were classified into 3 groups based on the number of apoptotic cells/5HPF: 0 to 10, 11 to 30, and 31+. The level of apoptosis increased with increasing size of the tumor, and apoptosis was rarely seen in tumors with positive ER or lower proliferative activity, as assessed by DNA polymerase alpha. As shown by DNA content analysis, apoptotic cells were observed more frequently in tumors with low G1 and high S-phase fractions. In addition, apoptosis was correlated with overexpression of p53 and poor prognosis. Although apoptosis did not correlate with EIC (extensive intraductal component) status, tumors with comedo component had higher values of apoptosis than those without comedo component. CONCLUSIONS: In breast cancer, apoptosis might reflect biological behavior, namely a higher degree of biological aggressiveness and unfavorable prognosis.

Adult↗

PubMed1999

Prognostic value of internal tandem duplication of the FLT3 gene in childhood acute myelogenous leukemia.

BACKGROUND: Recently, an internal tandem duplication of the FLT3 gene (FLT3/ITD) was found in 20% of adult cases of acute myelogenous leukemia (AML), and this length abnormality was suggested to be associated with leukemic progression. PROCEDURE: We examined the mRNA expression of the FLT3 gene by using reverse transcription-polymerase chain reaction (RT-PCR) in 64 children with AML, and further abnormal transcripts were cloned and sequenced. RESULTS: An unexpected longer product was found in seven patients (11%) by RT-PCR of the FLT3 gene. Sequence analysis of these abnormal products revealed the presence of tandemly duplicated fragments in all seven patients. Three factors were identified to be associated with high incidence of FLT3/ITD; older patients (> or = 10 years) (P = 0.049), high WBC count (> or = 50,000/microl) at presentation (P = 0.002), and M3 in FAB subtypes (P = 0.002). Induction failure was observed in 3 (43%) of 7 patients with FLT3/ITD. Only FLT3/ITD was identified as a significant risk factor for induction failure by univariate analysis (P = 0.013), although it was not significant by multivariate analysis (P = 0.11). The Kaplan-Meier estimate of event-free survival rate at 5 years was 14% for patients with FLT3/ITD, which was significantly lower in comparison with 69% for patients without FLT3/ITD (P = 0.003). This finding was also identified by multivariate analysis (P = 0.01). CONCLUSIONS: In this study, FLT3/ITD was observed in 11% of childhood AML and identified to be associated with poor prognosis. A large prospective study with uniform treatment is necessary to confirm our results.

Adolescent↗

PubMed1999

Intracellular transport of acid beta-glucosidase and lysosome-associated membrane proteins is affected in Gaucher's disease (G202R mutation).

Gaucher's disease (GD) is caused by an inherited deficiency of acid beta-glucosidase with storage of glucosylceramides in the lysosomes of macrophages. This study identifies a G202R mutation in the acid beta-glucosidase gene in an infant with severe neuronopathic (type 2) GD and only slightly reduced acid beta-glucosidase activity. Western blot analysis, pulse chase experiments, and the thin frozen section immunogold method were used to analyse the implications of this mutation on the pathogenesis, clinical heterogeneity and diagnostic evaluation of GD. The results show that acid beta-glucosidase persists in the patient's fibroblasts as a mannose-rich polypeptide in the endoplasmic reticulum and is not transported to the lysosomes. By contrast, high expression of the lysosome-associated membrane proteins LAMP-1 and LAMP-2, saposin C, and cathepsin D was observed in the patient's lysosomes. Immunogold labelling of the integral membrane proteins LAMP-1 and LAMP-2 increases significantly at the cell surface of Kupffer cells and fibroblasts as well as at the apical membrane of hepatocytes. In addition, LAMP-1 and LAMP-2 associate with the bilayer of stored glucosylceramide. It is concluded that defective intracellular transport of mutant acid beta-glucosidase from the endoplasmic reticulum to lysosomes leads to a more severe clinical phenotype than the residual enzyme activity may indicate. Furthermore, the detection of LAMP in the tubular bundles of undigested glucosylceramides, as well as their increased concentration at the surfaces of the affected cells, suggests that these proteins play a role in the storage or removal of substrate in GD. Intracellular targeting of acid beta-glucosidase and LAMP contributes to the broad phenotypic heterogeneity of GD.

Antigens, CD↗

PubMed1999

A family with an atonic variant of paroxysmal kinesigenic choreoathetosis and hypercalcitoninemia.

We report a family with an incompletely atonic variant of paroxysmal kinesigenic choreoathetosis (PKC). Three members of the family experienced attacks of muscle weakness which resembled the choreoathetotic attacks that occur in PKC in terms of their kinesigenicity and duration, clarity of consciousness during the attacks, good therapeutic response to low doses of phenytoin, and familial transmission, but differed from choreoathetotic attacks in PKC in that they were atonic. All three affected individuals were hypercalcitoninemic.

Adult↗

PubMed1999

Cataract formation through the polyol pathway is associated with free radical production.

The relationship between sugar cataract formation and radical production was investigated. The in vivo formation of free radicals in the lenses of rats fed on a diet containing 25% and 50% galactose was studied using the electron spin resonance (ESR) spin trapping method. Effects of treatment with aldose reductase inhibitor (ARI) SNK-860 on free radical formation were determined in 25% and 50% galactose fed rats. Hydroxyl radical (*OH) adduct of the spin trap 5,5'-dimethyl-1-pyrroline- N -oxide (DMPO) was directly detected in the superficial cortical cataract obtained from 25% and 50% galactose-fed rats. *OH production was completely inhibited by ARI SNK-860 in both galactose groups. Polyol accumulated in rat lenses given 50% galactose with a peak within the first 2 weeks, and was significantly inhibited by SNK-860. The increase in *OH production was considered with the polyol accumulation in both galactose groups. The dose of SNK-860 to inhibit *OH by 50% level was estimated at 3 m by the method of kinetic competition in vitro experiment. SNK-860 is not an effective *OH scavenger compared to other *OH scavengers. The results of the present study suggest that *OH is indirectly inhibited by SNK-860 resulting from decreasing polyol and *OH formation is related to sugar cataract formation in early stages, possibly via the Fenton reaction involving H2O2 produced from the activated polyol pathway. We suppose that *OH may accelerate damage to the cell membrane of lens fibers resulting from polyol accumulation *OH may play an important role in the early stage of sugar cataract process.

Aldehyde Reductase↗

PubMed1999

Corneal endothelial changes in patients with chronic renal failure.

PURPOSE: To evaluate the morphologic changes of the corneal endothelium in patients with chronic renal failure. METHODS: Twenty corneas of 20 patients with chronic renal failure undergoing hemodialysis were examined by wide-field specular microscopy. Twenty normal corneas from 20 healthy age-matched subjects were enrolled as control corneas. RESULTS: Despite normal endothelial cell density, the corneal endothelium of patients with chronic renal failure showed marked polymegethism and pleomorphism. CONCLUSION: Polymegethism and pleomorphism of the corneal endothelium are found in patients with chronic renal failure.

Adolescent↗

PubMed1999

Role of synaptotagmin, a Ca2+ and inositol polyphosphate binding protein, in neurotransmitter release and neurite outgrowth.

Synaptotagmin I (or II), a possible Ca(2+)-sensor of synaptic vesicles, has two functionally distinct C2 domains: the C2A domain binds Ca2+ and the C2B domain binds inositol high polyphosphates (IP4, IP5, and IP6). Ca(2+)-regulated exocytosis of secretory vesicles is proposed to be activated by Ca2+ binding to the C2A domain and inhibited by inositol polyphosphate binding to the C2B domain. Synaptotagmins now constitute a large family and are thought to be involved in both regulated and constitutive vesicular trafficking. They are classified from their distribution as neuronal (synaptotagmin I-V, X, and XI) and the ubiquitous type (synaptotagmin VI-IX). Among them, synaptotagmins III, V, VI and X are deficient in IP4 binding activity due to the amino acid substitutions in the C-terminal region of the C2B domain, suggesting that these isoforms can work for vesicular trafficking even in the presence of inositol high polyphosphates. Synaptotagmin I is also known to be present in neuronal growth cone vesicles. Antibody against the C2A domain (anti-C2A) that inhibits Ca(2+)-regulated exocytosis also blocked neurite outgrowth of the chick dorsal root ganglion (DRG) neuron, suggesting that Ca(2+)-dependent synaptotagmin activation is also crucial for neurite outgrowth.

Amino Acid Sequence↗

PubMed1999

Splenectomy attenuates superoxide anion release into the hepatic sinusoids after lipopolysaccharide challenge.

BACKGROUND/AIMS: The aim of this study was to determine whether the spleen contributes to superoxide anion release into the hepatic sinusoids and subsequent damage to endothelial cells of the hepatic sinusoids after lipopolysaccharide challenge. METHODS: Rats were given 2 mg/kg body weight lipopolysaccharide. Three hours after the treatment, superoxide anion release into the hepatic sinusoids was examined in a liver perfusion model using the cytochrome C method. Damage to endothelial cells of the hepatic sinusoids was assessed from the purine nucleoside phosphorylase/glutamic-pyruvic transaminase ratio in the liver perfusate. To further characterize the mechanisms behind these changes, these studies were done in rats given superoxide dismutase or an anti-TNFalpha antibody. To study whether the spleen plays a role in the mechanisms, experiments with splenectomized rats were performed. RESULTS: Lipopolysaccharide challenge resulted in superoxide anion release into the hepatic sinusoids and damage to endothelial cells of the hepatic sinusoids. These changes were significantly attenuated by the treatments with superoxide dismutase or an antibody against TNFalpha, as well as by splenectomy. The hepatic macrophage and Kupffer cell populations after lipopolysaccharide challenge were significantly smaller in the rats given splenectomy than in those given a sham operation. There were no significant differences in the neutrophil populations between the two groups. Levels of TNFalpha were significantly lower in the former than the latter, whereas there were no significant differences in levels of Interleukin-8 between the two groups. CONCLUSIONS: Splenectomy reduced the superoxide anion release into the hepatic sinusoids caused by the lipopolysaccharide challenge and subsequent damage to endothelial cells of the hepatic sinusoids. This supports the view that splenectomy has a protective effect in lipopolysaccharide-induced liver injury.

Animals↗

PubMed1999

Relationship between joint effusion, joint pain, and protein levels in joint lavage fluid of patients with internal derangement and osteoarthritis of the temporomandibular joint.

PURPOSE: The purpose of this study was to investigate the relationship between the presence of joint effusion, joint pain, and protein levels in joint lavage fluid (JL) of patients with internal derangement (ID) and osteoarthritis (OA) of the temporomandibular joint (TMJ). PATIENTS AND METHODS: Thirty-eight joints in 26 patients with ID and OA of the TMJ were studied. Magnetic resonance imaging (MRI) evidence of joint effusion was evaluated in T2-weighted images. Samples of JL were collected from the superior joint space during pumping manipulation, and the protein concentration was measured. The presence of pain was based on joint tenderness or a complaint of pain in the preauricular region during mouth opening or closing. RESULTS: Joint effusion was demonstrated in 20 of 25 (80%) painful joints; a significantly higher incidence than in pain-free joints (5 of 13, 38.5%). The mean protein concentration (2.15 mg/mL) in JL from painful joints was significantly higher than in pain-free joints (1.22 mg/mL) (P < .05). Furthermore, the mean protein concentration (2.12 mg/mL) in JL from joints with effusion was significantly higher than in joints without joint effusion (1.27 mg/mL) (P < .05). CONCLUSIONS: These data demonstrate that painful joints are more likely to show joint effusion on MRI, and the protein levels in JL recovered from these joints is higher than in pain-free joints. These data also suggested that joint effusion may be related to the inflammatory changes seen in patients with ID and OA.

Adolescent↗

PubMed1999

Functional involvement of synaptotagmin I/II C2A domain in neurite outgrowth of chick dorsal root ganglion neuron.

Synaptotagmin I or II (Syt I/II) is involved in Ca2+-regulated exocytosis of secretory vesicles, probably serving as a Ca2+-sensor via its C2A domain. Synaptotagmin is also known to be expressed in neuronal growth cone vesicles, but its functional involvement in neurite outgrowth remains largely unknown. In this study, we examined the function of Syt I/II in neurite outgrowth in cultured chick dorsal root ganglion neurons using an anti-synaptotagmin I and II C2A domain (anti-STI/II-C2A) antibody that inhibits Ca2+-regulated exocytosis. Immunoblots confirmed the high specificity of the anti-STI/II-C2A antibody and showed the expression of synaptotagmin I or II in chick dorsal root ganglion neurons. Immunocytochemistry revealed that synaptotagmin I or II is enriched at the growth cone region of chick dorsal root ganglion neurons, in both lamellipodia and filopodia. Whole or Fab-fragment of the anti-STI/II-C2A antibody loaded into dorsal root ganglion neurons by trituration significantly inhibited neurite outgrowth, whereas preimmune immunoglobulin G had no effect. These results showed that the C2A domain of synaptotagmin I or II plays a crucial role in neurite outgrowth.

Animals↗

PubMed1999

A novel, high endothelial venule-specific sulfotransferase expresses 6-sulfo sialyl Lewis(x), an L-selectin ligand displayed by CD34.

L-selectin mediates lymphocyte homing by facilitating lymphocyte adhesion to unique carbohydrate ligands, sulfated sialyl Lewis(x), which are expressed on high endothelial venules (HEV) in secondary lymphoid organs. The nature of the sulfotransferase(s) that contribute to sulfation of such L-selectin counterreceptors has been uncertain. We herein describe a novel L-selectin ligand sulfotransferase, termed LSST, that directs the synthesis of the 6-sulfo sialyl Lewis(x) on L-selectin counterreceptors CD34, GlyCAM-1, and MAdCAM-1. LSST is predominantly expressed in HEV and exhibits striking catalytic preference for core 2-branched mucin-type O-glycans as found in natural L-selectin counterreceptors. LSST enhances L-selectin-mediated adhesion under shear compared to nonsulfated controls. LSST therefore corresponds to an HEV-specific sulfotransferase that contributes to the biosynthesis of L-selectin ligands required for lymphocyte homing.

Acetylglucosamine↗