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Biomedical subjects

R Fukuda

Publications and source records attributed to R Fukuda.

At least 37 records · Page 2Linked to original sources
PubMed2001

T cell-mediated cytotoxicity via Fas/Fas ligand signaling in Helicobacter pylori-infected gastric corpus.

BACKGROUND: Helicobacter pylori infection induces T helper-1 immune responses in inflamed mucosa. However, the role of T cell-mediated cytotoxicity in the induction of epithelial apoptosis is still unclear. The aim of this study was to investigate the involvement of the Fas/Fas ligand (Fas/Fas-L) system in the H. pylori-infected gastric corpus. MATERIALS AND METHODS: Gastric fundic biopsy specimens were taken from patients with and without H. pylori infection. The expression of Fas and Fas-L was examined by immunohistochemistry and RT-PCR. Subsets of gastric infiltrating T cells in the biopsy specimens were studied by immunohistochemistry and flow cytometry. In histological sections, apoptosis was detected by the TUNEL method. We studied the in vitro expression of Fas-L in peripheral T cells after stimulation with H. pylori antigen and interferon-gamma (IFN-gamma). The Fas-mediated in vitro cytotoxicity of activated T cells was assessed by flow cytometry. RESULTS: The numbers of CD4+ and CD8+ T cells were greater in H. pylori-infected subjects. Fas expression was abundantly increased on fundic gland epithelium, and Fas-L was detected on lamina propria mononuclear cells in H. pylori-infected mucosa. TUNEL-positive epithelial cells were also increased in H. pylori-infected subjects. H. pylori antigen and IFN-gamma induced Fas-L mRNA expression in both CD4+ and CD8+ T cells. In cytotoxic assay, activated T cells induced apoptosis in AGS cells, which could be significantly inhibited by neutralizing Fas-L antibody. CONCLUSIONS: T cell-mediated cytotoxicity via Fas/Fas-L signaling may contribute to the induction of apoptosis in gastric epithelial cells during H. pylori infection.

Adult↗
PubMed2001

Functional magnetic resonance imaging of odor identification: the effect of aging.

BACKGROUND: Sense of smell declines with age and impairment in olfaction has been observed in some neurodegenerative disorders such as Alzheimer's disease. Functional neuroimaging techniques enable researchers to observe brain regions activated by olfactory stimuli. METHODS: We gave three mainly olfactory-mediated odors (limonene, methylsalicylate, and eugenol) to six young and six elderly subjects and observed the areas activated by using blood oxygen level dependent contrast functional magnetic resonance imaging. RESULTS: The group mapping of young subjects showed extensive activation in the orbitofrontal cortex, commonly believed to be the olfactory cortex, some limbic areas (the hippocampus and the thalamus), regions involved with gustatory sensation (the anterior insula and the inferior postcentral gyrus), superior and inferior temporal gyri, and cerebellum. In the elderly group, only the left inferior temporal gyrus and the primary visual cortex reached accepted significance levels. CONCLUSIONS: We have therefore confirmed previous reports of brain regions involved in olfactory processing in young volunteers and demonstrated decreased activation in elderly volunteers.

Adult↗
PubMed2001

Clinical relevance of precore and basal core promoter variants of hepatitis B virus during natural hepatitis B e antigen seroconversion may be overstated.

BACKGROUND: Clinical relevance of nucleotide changes in precore and basal core promoters in the hepatitis B virus genome during hepatitis B e antigen seroconversion may be overstated. The authors investigated the existence and changes in the relative proportion of variants to wild virus that occur with seroconversion. METHODS: Sera from 30 school-aged long-term hepatitis B virus carriers, including 11 tested before and after seroconversion during 1 to 8 years of follow-up, were evaluated for variations in nucleotide sequences of the basal core promoter (T1762 and A1764), precore region (A1869), and carboxyl-terminus of the X region of the hepatitis B virus genome using an amplification refractory mutation detection system with mutant-specific primers. RESULTS: All variants were found to already exist before seroconversion at various wild-type/mutant ratios. The positive rates of these variants were not changed with loss of hepatitis B e antigen. Although there was a relative increase in the concentration of these mutants in wild-type/mutant mixed populations, most patients with only a wild-type population maintained the same pattern after loss of hepatitis B e antigen. CONCLUSIONS: Our results indicate that hepatitis B virus exists as a quasi species, and correlations of nucleotide sequences with clinical and serologic findings must be done with caution.

Adolescent↗
PubMed2001

Effects of mutation in hepatitis C virus nonstructural protein 5A on interferon resistance mediated by inhibition of PKR kinase activity in mammalian cells.

The IFN-induced double-stranded RNA (dsRNA)-activated protein kinase PKR is one of the key molecules in the antiviral effects of IFN. To clarify the effects of hepatitis C virus nonstructural protein 5A (NS5A) on antiviral activity of IFN, in particular on PKR kinase activity, in mammalian cells, we established inducible NS5A-expressing cell lines derived from human osteosarcoma (Saos-2). The cells expressing NS5A derived from an IFN-resistant clone (NS5A-lb) that interacted with endogenous PKR in vitro, showed a suppressive effect on IFN function as determined by interference with vesicular stomatitis virus (VSV) infection, whereas NS5A (NS5A-2a) from an IFN-sensitive clone did not block the antiviral effect of IFN. A mutant with deletion of the IFN sensitivity determining region (ISDR) in NS5A-1b (NS5A-AISDR) also interacted with PKR and suppressed its activity in vitro. However, neither NS5A-2a nor the C-terminal truncated mutant of NS5A-1b (NS5A-deltaC) blocked PKR activity. These observations confirmed the previous report that the inhibitory effect of NS5A on IFN activity is mediated at least in part by the repression of PKR. In addition, we showed that IFN sensitivity was determined not only by the ISDR but that the involvement of the C-terminal region of NS5A-1b is important for the suppression of PKR activity.

Amino Acid Sequence↗
PubMed2001

Structural abnormalities in frontal, temporal, and limbic regions and interconnecting white matter tracts in schizophrenic patients with prominent negative symptoms.

OBJECTIVE: Imaging studies of schizophrenia have repeatedly demonstrated global abnormalities of cerebral and ventricular volumes. However, pathological changes at more local levels of brain organization have not yet been so clearly characterized because of the few brain regions of interest heretofore included in morphometric analyses as well as heterogeneity of patient samples. METHOD: Dual echo magnetic resonance imaging (MRI) data were acquired at 1.5 T from 27 right-handed patients who met DSM-IV criteria for schizophrenia with enduring negative symptoms and from 27 healthy comparison subjects. Between-group differences in gray and white matter volume were estimated at each intracerebral voxel after registration of the images in standard space. The relationship between clinical symptom scores and brain structure was also examined within the patient group. Spatial statistics and permutation tests were used for inference. RESULTS: Significant deficits of gray matter volume in the patient group were found at three main locations: 1) the left superior temporal gyrus and insular cortex, 2) the left medial temporal lobe (including the parahippocampal gyrus and hippocampus), and 3) the anterior cingulate and medial frontal gyri. The volume of these three regions combined was 14% lower in the patients relative to the comparison subjects. White matter deficits were found in similar locations in the left temporal lobe and extended into the left frontal lobe. The patient group showed a relative excess of gray matter volume in the basal ganglia. Within the patient group, basal ganglia gray matter volume was positively correlated with positive symptom scores. CONCLUSIONS: Anatomical abnormalities in these schizophrenic patients with marked negative symptoms were most evident in left hemispheric neocortical and limbic regions and related white matter tracts. These data are compatible with models that depict schizophrenia as a supraregional disorder of multiple, distributed brain regions and the axonal connections between them.

Adult↗
PubMed2001

Gastrointestinal stromal tumor with a novel mutation of KIT proto-oncogene.

The majority of human gastrointestinal mesenchymal tumors are gastrointestinal stromal tumors (GISTs). Recent reports have shown the existence of gain-of-function mutations in the juxta-membrane domain of receptor tyrosine kinase (KIT) in GISTs. We present a 77-year-old woman with GIST diagnosed by positive immunostaining of cluster designation (CD) 34 and KIT. This case had a novel mutation at codon 576 located in the juxta-membrane domain of KIT. Our results indicate the importance of mutations in this KIT region and suggest the possibility of the existence of other types of mutations in this region in GISTs.

Aged↗
PubMed2000

Compartmental specificity of cellular membrane fusion encoded in SNARE proteins.

Membrane-enveloped vesicles travel among the compartments of the cytoplasm of eukaryotic cells, delivering their specific cargo to programmed locations by membrane fusion. The pairing of vesicle v-SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) with target membrane t-SNAREs has a central role in intracellular membrane fusion. We have tested all of the potential v-SNAREs encoded in the yeast genome for their capacity to trigger fusion by partnering with t-SNAREs that mark the Golgi, the vacuole and the plasma membrane. Here we find that, to a marked degree, the pattern of membrane flow in the cell is encoded and recapitulated by its isolated SNARE proteins, as predicted by the SNARE hypothesis.

Biological Transport↗
PubMed2000

Topological restriction of SNARE-dependent membrane fusion.

To fuse transport vesicles with target membranes, proteins of the SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) complex must be located on both the vesicle (v-SNARE) and the target membrane (t-SNARE). In yeast, four integral membrane proteins, Sed5, Bos1, Sec22 and Bet1 (refs 2-6), each probably contribute a single helix to form the SNARE complex that is needed for transport from endoplasmic reticulum to Golgi. This generates a four-helix bundle, which ultimately mediates the actual fusion event. Here we explore how the anchoring arrangement of the four helices affects their ability to mediate fusion. We reconstituted two populations of phospholipid bilayer vesicles, with the individual SNARE proteins distributed in all possible combinations between them. Of the eight non-redundant permutations of four subunits distributed over two vesicle populations, only one results in membrane fusion. Fusion only occurs when the v-SNARE Bet1 is on one membrane and the syntaxin heavy chain Sed5 and its two light chains, Bos1 and Sec22, are on the other membrane where they form a functional t-SNARE. Thus, each SNARE protein is topologically restricted by design to function either as a v-SNARE or as part of a t-SNARE complex.

Carrier Proteins↗
PubMed2000

Functional architecture of an intracellular membrane t-SNARE.

Lipid bilayer fusion is mediated by SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) located on the vesicle membrane (v-SNAREs) and the target membrane (t-SNAREs). The assembled v-SNARE/t-SNARE complex consists of a bundle of four helices, of which one is supplied by the v-SNARE and the other three by the t-SNARE. For t-SNAREs on the plasma membrane, the protein syntaxin supplies one helix and a SNAP-25 protein contributes the other two. Although there are numerous homologues of syntaxin on intracellular membranes, there are only two SNAP-25-related proteins in yeast, Sec9 and Spo20, both of which are localized to the plasma membrane and function in secretion and sporulation, respectively. What replaces SNAP-25 in t-SNAREs of intracellular membranes? Here we show that an intracellular t-SNARE is built from a 'heavy chain' homologous to syntaxin and two separate non-syntaxin 'light chains'. SNAP-25 may thus be the exception rather than the rule, having been derived from genes that encoded separate light chains that fused during evolution to produce a single gene encoding one protein with two helices.

Escherichia coli↗
PubMed2000

Myopathy phenotype of transgenic mice expressing active site-mutated inactive p94 skeletal muscle-specific calpain, the gene product responsible for limb girdle muscular dystrophy type 2A.

A defect of the gene for p94 (calpain 3), a skeletal muscle-specific calpain, is responsible for limb girdle muscular dystrophy type 2A (LGMD2A), or 'calpainopathy', which is an autosomal recessive and progressive neuromuscular disorder. To study the relationships between the physiological functions of p94 and the etiology of LGMD2A, we created transgenic mice that express an inactive mutant of p94, in which the active site Cys129 is replaced by Ser (p94:C129S). Three lines of transgenic mice expressing p94:C129S mRNA at various levels showed significantly decreased grip strength. Sections of soleus and extensor digitorum longus (EDL) muscles of the aged transgenic mice showed increased numbers of lobulated and split fibers, respectively, which are often observed in limb girdle muscular dystrophy muscles. Centrally placed nuclei were also frequently found in the EDL muscle of the transgenic mice, whereas wild-type mice of the same age had almost none. There was more p94 protein produced in aged transgenic mice muscles and it showed significantly less autolytic degradation activity than that of wild-type mice. Although no necrotic-regenerative fibers were observed, the age and p94:C129S expression dependence of the phenotypes strongly suggest that accumulation of p94:C129S protein causes these myopathy phenotypes. The p94:C129S transgenic mice could provide us with crucial information on the molecular mech-anism of LGMD2A.

Animals↗
PubMed2000

Interactions of phosphatidylinositol 3-kinase Src homology 3 domain with its ligand peptide studied by absorption, circular dichroism, and UV resonance raman spectroscopies.

Absorption, circular dichroism (CD), and UV resonance Raman (UVRR) spectroscopies were applied to selectively examine the environmental and structural changes of Trp and Tyr residues in the phosphatidylinositol 3-kinase (PI3K) SH3 domain induced by ligand association. Comparison of the spectra of PI3K SH3 in the presence or absence of its ligand peptide RLP1 (RKLPPRPSK) indicated that RLP1 binding changed the environment of Trp55 of the SH3 to be more hydrophilic and its H bonding weaker and that of Tyr residues to be more hydrophobic. The D21N mutant (Asp21 --> Asn) of the SH3 yielded a UV CD distinct from that of the wild type, and its spectral changes induced by RLP1 binding were smaller and different from those of the wild type in absorption, CD, and UVRR spectra, suggesting that the mutation of conserved Asp21 affected the conformation of the ligand binding cleft and thus might lead to the decrease in the ligand affinity. These data provide direct evidence for the occurrence of environmental and structural changes of PI3K SH3 by the association of a ligand and the D21N mutation.

Absorption↗
PubMed2000

Factors affecting serum haloperidol level assessed by longitudinal therapeutic monitoring.

1. Since the development of a kit for the assay of haloperidol by enzyme immunoassay, therapeutic monitoring of haloperidol has been utilised in Japan for several years. By retrospectively analysing the accumulated data, this study was carried out to investigate the factors affecting haloperidol level. Especially, the effects of enzyme-inducing comedications were analysed in relation to serum gamma-glutamyltransferase (GGT) level, which has been measured simultaneously with haloperidol. Serum haloperidol level measurements were obtained from medical records of inpatients on multiple medications (n = 102). For each subject, average haloperidol level was computed during the same prescription and doses. The effects of age, sex, smoking status, and the coadministration of carbamazepine and barbiturates (including phenobarbital, amobarbital and pentobarbital) were analysed using correlation, between-group comparison and multiple regression analysis. Separately, the effect of comedications on haloperidol and GGT levels were analysed individually in a small number of patients (n = 5) who had received those comedications intermittently. Significant lowering of serum haloperidol level by the coadministration of carbamazepine and/or barbiturates was observed. The coadministration was also correlated with the elevated GGT level in between-group comparison. In the separate analysis, the change in haloperidol level was correlated with the change in GGT level in some individuals but not in others. None of the other clinical factors investigated in the study showed significant effect on haloperidol level. This study suggests that the lowering of haloperidol level and the elevation of GGT level may often occur coincidentally by the coadministration of enzyme-inducing drugs. However, whether there is a causal relationship between these phenomena and whether elevated serum GGT level could serve as a clinically useful marker need to be clarified by further basic pharmacological research.

Adult↗
PubMed2000

Diagnostic accuracy of a new non-invasive enzyme immunoassay for detecting Helicobacter pylori in stools after eradication therapy.

BACKGROUND: Helicobacter pylori eradication therapy has been commonly performed for patients with peptic ulcer. An inexpensive, reliable, non-invasive test would be useful for evaluation of the effectiveness of eradication therapy. AIM: To evaluate the diagnostic potential of a new non-invasive enzyme immunoassay (HpSA) for H. pylori antigen in stools. METHODS: A total of 112 peptic ulcer patients with H. pylori infection (82 male, 30 female; age range 22-81 years, mean 54 years) received a course of eradication therapy. Four weeks after the end of the therapy, stool samples were collected from all patients and tested using the HpSA. The diagnostic accuracy of the HpSA EIA was evaluated in comparison with the results of a 13C-urea breath test. RESULTS: The diagnostic sensitivity of HpSA EIA after eradication therapy was 90%, the specificity 98%, the positive predictive value 82% and the negative predictive value 99%. CONCLUSION: The HpSA stool test is potentially useful for the diagnosis of H. pylori infection 4 weeks after the end of eradication therapy.

Adult↗
PubMed2000

CYP2C19 genotype status and intragastric pH during dosing with lansoprazole or rabeprazole.

BACKGROUND: CYP2C19 has an important role in the catabolism of several proton pump inhibitors. However, the relative contribution of CYP2C19-mediated metabolism varies among the different proton pump inhibitors. AIM: To determine the effect of CYP2C19 genotype status on intragastric pH during dosing with lansoprazole or rabeprazole. SUBJECTS AND METHODS: The subjects were 20 male volunteers without Helicobacter pylori infection. Their CYP2C19 genotype status was determined by a polymerase chain reaction-restriction fragment length polymorphism method. Twenty-four-hour monitoring of intragastric acidity was performed three times: once without medication, once on the last day of a 7-day course of rabeprazole, and once on the last day of a 7-day course of lansoprazole. RESULTS: Subjects were divided into three groups on the basis of their CYP2C19 genotype status: homozygous extensive metabolizers (homo-EMs, n=7), heterozygous extensive metabolizers (hetero-EMs, n=9), and poor metabolizers (PMs, n=4). The median pH during rabeprazole administration was not influenced by CYP2C19 genotype. On the other hand, the median pH in PMs during lansoprazole dosing was higher than in homo-EMs and hetero-EMs. The percentage of time with pH < 4.0 had a similar tendency to that of median pH. CONCLUSION: CYP2C19 genotype status influences gastric acid suppression by lansoprazole, but not by rabeprazole.

2-Pyridinylmethylsulfinylbenzimidazoles↗
PubMed2000

Neutrophil chemoattractant 2 beta regulates expression of the Reg gene in injured gastric mucosa in rats.

BACKGROUND & AIMS: Regenerating (Reg) protein has a trophic effect on gastric mucosal cells. We have shown that Reg gene expression is increased in enterochromaffin-like (ECL) cells during the healing of damaged gastric mucosa around mucosal erosion. This study was designed to explore the stimulants of Reg expression during the healing of gastric mucosal damage. METHODS: Time course changes of the expression of genes for various proinflammatory cytokines and Reg were investigated after induction of gastric mucosal lesions in rats. The direct effect of proinflammatory cytokines on Reg gene expression and Reg protein production were investigated in vitro using counterflow elutriation-enriched rat ECL cells. CXC receptor 2 (CXCR-2) expression was investigated in ECL cells by reverse-transcription polymerase chain reaction. Reg gene expression was also investigated in rats treated by the neutralizing antibody of cytokine-induced neutrophil chemoattractant (CINC-2 beta). RESULTS: During healing, the gene expression of several proinflammatory cytokines and Reg was markedly augmented. Among the proinflammatory cytokines, CINC-2 beta is the only cytokine in which augmented expression preceded the increase of Reg gene expression. In rats treated with CINC-2 beta neutralizing antibody, the augmentation of Reg gene expression was significantly inhibited. When ECL cells were incubated with these proinflammatory cytokines, CINC-2 beta dose-dependently increased Reg messenger RNA and Reg protein in ECL cells. CXCR-2 was identified in isolated ECL cells. CONCLUSIONS: CINC-2 beta, expressed in damaged gastric mucosa, stimulates the production of Reg protein in ECL cells via CXCR-2 and may be involved in the accelerated healing of injured gastric mucosa.

Animals↗
PubMed2000

Expression of peroxisome proliferator-activated receptor (PPAR)gamma in gastric cancer and inhibitory effects of PPARgamma agonists.

Peroxisome proliferator-activated receptor (PPAR) gamma is expressed in human colon cancer, prostate cancer and breast cancer cells, and PPARgamma activation induces growth inhibition in these cells. PPARgamma expression in human gastric cancer cells, however, has not been fully investigated. We report the PPARgamma expression in human gastric cancer, and the effect of PPARgamma ligands on proliferation of gastric carcinoma cell lines. Immunohistochemistry was used to demonstrate the presence of PPARgamma protein in surgically resected specimens from well differentiated, moderately differentiated and poorly differentiated adenocarcinoma. We used reverse transcription-polymerase chain reaction and Northern and Western blot analyses to demonstrate PPARgamma expression in four human gastric cancer cell lines. PPARgamma agonists (troglitazone and 15-deoxy-Delta(12,14)-prostaglandin J2) showed dose-dependent inhibitory effects on the proliferation of the gastric cancer cells, and their effect was augmented by the simultaneous addition of 9- cis retinoic acid, a ligand of RXRalpha. Flow cytometry demonstrated G1 cell cycle arrest and a significant increase of annexin V-positive cells after treatment with troglitazone. These results suggest that induction of apoptosis together with G1 cell cycle arrest may be one of the mechanisms of the antiproliferative effect of PPARgamma activation in human gastric cancer cells.

Apoptosis↗
PubMed2000

Effect of rebamipide on prostaglandin EP4 receptor gene expression in rat gastric mucosa.

Prostaglandin E2 (PGE2) plays an important role in the regulation of gastric mucus secretion. We have previously shown that the prostaglandin EP4 receptor (EP4) gene is abundantly expressed in gastric mucus-producing cells. Furthermore, we have shown that EP4 is present in a rat normal gastric mucosal cell line (RGM1) and that PGE2 increases mucus secretion from these cells via EP4. Rebamipide, an anti-gastric ulcer agent, has been reported to promote gastric PGE2 production and mucus secretion. However, it is unclear whether rebamipide influences mucus secretion by altering expression of the EP4 gene. Therefore, we tested the effect of rebamipide on EP4 gene expression in the gastric mucosa. Seven-week-old Wistar rats received oral rebamipide (100 mg/kg) with and without water-immersion restraint stress (WRS). All rats were killed, and their gastric tissues were used to investigate the expression of mRNA for EP4 and cyclooxygenase types 1 and 2. The thickness of the gastric mucus layer was also measured. The effect of rebamipide on EP4 gene expression and PGE2 production in RGM1 cells was also investigated in vitro. Furthermore, the effect of PGE2 on cyclic adenosine monophosphate (cAMP) production by RGM1 cells with or without rebamipide was studied. Oral rebami-pide significantly increased EP4 gene expression in the gastric antrum but not in the corpus after WRS. Furthermore, it increased surface mucus thickness and suppressed ulcer formation in the gastric mucosa after WRS. In vitro, rebamipide significantly augmented EP4 gene expression in RGM1 cells, and PGE2 significantly increased the cAMP production by RGM1 cells incubated with rebamipide. Rebamipide promotes EP4 gene expression and may consequently increase the gastric mucus secretion via EP4 receptors in the rat antral mucosa.

Alanine↗