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Hitoshi Takahashi

Publications and source records attributed to Hitoshi Takahashi.

At least 55 records · Page 3Linked to original sources
PubMed2005

Analysis of the 5' end structure of HCV subgenomic RNA replicated in a Huh7 cell line.

OBJECTIVE: Recently, HCV subgenomic RNA that replicates in vitro in a certain cell line have been elucidated. Since the 5' end of the genome of positive strand RNA viruses is often modified with a cap structure or a covalently linked protein, we have assessed structural feature of the HCV genome obtained from Huh7 cells in which HCV subgenomic RNA has been shown to efficiently self-replicate. METHODS: HCV subgenomic RNA was obtained from the Huh7 and was analyzed for its 5' end. RESULTS: Phosphorylation of the genomic RNA by polynucleotide kinase was observed only after treatment with phosphatase. The labeling efficiency of the genome with polynucleotide kinase was not enhanced by treatment with pyrophosphatase. CONCLUSION: It is suggested that the 5' end of HCV genomic RNA obtained from HCV replicon cells is not modified except phosphorylation. Furthermore, analysis of the 5' end of the HCV RNA obtained from the HCV subgenome self-replicating cells revealed the presence of two types of subgenomic RNA that contained either guanylate or adenylate at the 5' end. This result indicates that the 5' end of the subgenome in Huh7 cells is redundant and there is no significant evolutionary advantage between the two genomes.

Adenosine Monophosphate↗
PubMed2005

Pathologic features of dysplasia and accompanying alterations observed in surgical specimens from patients with intractable epilepsy.

Malformations caused by abnormalities of cortical development, or cortical dysplasias, were examined in surgical specimens from 108 patients with medically intractable epilepsy to determine the scope of histopathologic changes. The relevance of the clinical findings was also evaluated. Various types and degrees of dysplastic features were observed in various combinations, including architectural abnormalities, an increased number of neurons in the molecular layer and/or cortical layer II, neuronal clustering, an increased number of satellite oligodendrocytes, abnormal gyration, single and/or aggregates of heterotopic neurons in the white matter, and the appearance of cytologically abnormal cells, such as giant or dysmorphic neurons and balloon cells. In the temporal lobe specimens, microdysgenesis (corresponding to mild malformations caused by abnormalities of cortical development and type IA/B focal cortical dysplasias) was more frequently observed than Taylor-type focal cortical dysplasia (type IIA/B), whereas in the frontal lobe specimens, the frequency of occurrence of both types was even. The ages at seizure onset and surgery of patients with the latter type were significantly lower than those of patients with the former. On the other hand, prominent astrocytosis in the cortex and white matter was evident in all cases, and many corpora amylacea and neurofibrillary tangle-like inclusions were observed in a subset of cases. An ultrastructural investigation revealed dilatation of the postsynaptic dendritic spines and shafts in the cortex and features indicating the occurrence in the white matter of demyelination followed by remyelination. Thus, with regard to the epileptogenic lesions, although dysplastic changes constitute the pathogenetic basis, the overlapping subsequent degenerative processes involving synapses, dendrites, and axons might contribute to the development of epileptogenic processes. Astrocytes might also actively participate in the development of the pathogenesis of epilepsy.

Adolescent↗
PubMed2005

A new preadipocyte cell line, AP-18, established from adult mouse adipose tissue.

Establishing preadipocyte cell lines from mature adipose tissues could help lead to a better understanding of adipogenesis. We have established a unique preadipocyte cell line, AP-18, derived from the subepidermal layer of ear skin from an adult C3H/HeM mouse. AP-18 cells exhibit fibroblast-like morphology, slow growth, and contact inhibition. The doubling time of AP-18 cells is 50-60 h, which is about 2-fold longer than that of well-known 3T3-L1 cells derived from mouse embryos. A small population of AP-18 cells spontaneously differentiates into adipocytes by 8 days after confluence, as judged by the accumulation of triglyceride droplets. Treatment of confluent AP-18 preadipocytes with adipogenic agents, containing dexamethasone, 3-methyl-1-isobutylxanthine, and insulin, increased triglyceride contents about 5-fold compared to the contents in untreated cells. We also analyzed mRNA expression profiles for key transcription factors involved in adipocyte differentiation, peroxisome proliferator-activated receptor (PPAR)gamma and the CCAAT/enhancer binding protein (C/EBP) family, and for differentiation markers, aP2, adipocyte-specific fatty acid-binding protein and adipsin, adipocyte-specific serine protease. AP-18 preadipocytes express mRNAs for C/EBPbeta, C/EBPalpha, PPARgamma, and aP2 before differentiation, but not adipsin mRNA. Expression of aP2 mRNA was increased in fully differentiated AP-18 cells. Likewise, expression of adipsin mRNA was increased after induced differentiation of AP-18 cells and reached the highest level in fully differentiated adipocytes. Thus, differentiation of AP-18 cells is associated with the increased expression of aP2 and adipsin mRNAs. The newly established AP-18 cell line provides a useful model for investigating adipocyte differentiation and adipogenesis.

3T3 Cells↗
PubMed2005

Isolation and characterization of an N-linked oligosaccharide that is increased in glioblastoma tissue and cell lines.

We have isolated and characterized N-linked oligo-saccharides that are significantly increased in glioblastoma tissue and cell lines. The structures of N-linked oligosaccharides present in 3 human normal brain tissues, 15 patients with glio-blastoma and 3 glioma cell lines were analyzed by partially automated technique for the isolation and fluorescent labeling of N-linked sugar chains from glycoproteins. Characterization of the sugar chains was achieved with the use of a combination of HPLC columns and a highly sensitive fluorescence detector at femtomole levels. By collecting peaks which accounted for 0.1% or more, sixteen different oligosaccharide structures were characterized from glioblastoma tissue and cell lines. The 16 oligosaccharide structures accounted for 48.9% of the total N-linked oligosaccharides present in glioblastoma tissue. The major components of total oligosaccharides were similar to those of normal brain tissue. The amount of a biantennary bigalactosylated structure with one core fucosylation (A2G2F) was present in increased levels in glioblastoma tissue (mean = 2.90%) and glioma cell lines (mean = 5.60%), while being less than 0.1% in normal brain tissue. Expression of highly branched tetra-antennary N-glycans that are usually detected in lungs or hepatocellular cancer was not observed. Tissue glioma cells and cultured cells also displayed strong LCA-lectin binding, which binds to sugar chains with core fucose (including A2G2F), while normal brain tissue did not. Moreover, LCA lectin inhibited proliferation of glioma cells through induction of apoptosis. A2G2F on glioma specimens may provide a novel marker and target for the diagnosis and treatment of glioblastoma, respectively.

Adolescent↗
PubMed2004

Remarkable effect of milnacipran, a serotonin-noradrenalin reuptake inhibitor (SNRI), on depressive symptoms in patients with Parkinson's disease who have insufficient response to selective serotonin reuptake inhibitors (SSRIs): two case reports.

The authors present here two cases of Parkinson's disease with depression refractory to SSRIs treatment, who experienced a complete remission after replacing the ongoing SSRIs with a serotonin-noradrenalin reuptake inhibitor (SNRI), milnacipran. The case reports suggest that milnacipran may be one of the treatment options for depression in patients with Parkinson's disease who had inadequate response to SSRIs. Further studies are warranted to confirm this observation.

Adrenergic Uptake Inhibitors↗
PubMed2004

Cerebral lipoma and the underlying cortex of the temporal lobe: pathological features associated with the malformation.

Intracranial lipomas are believed to be congenital malformations rather than true neoplasms, resulting from the abnormal differentiation of the meninx primitiva, the undifferentiated mesenchyme. We report here the surgical pathological features of a lipoma that was located on the cerebral surface of an abnormally formed fissure, and the underlying cortex of the middle temporal gyrus of a 20-year-old woman. The mass was composed of typical adipose tissue in which a large number of blood vessels were present. Thick connective tissue associated with the arachnoid membrane covered the cortical surface. The cortex exhibited a polymicrogyric configuration in which the cortical ribbon was abnormally undulated and excessively folded. Reelin-immunolabeled Cajal-Retzius-cell-like cells were observed frequently in the fused molecular layer. The cortical lamination underlying the molecular layer was poorly defined. Along the border between the connective tissue and cortical surface, there was a narrow zone in which the mesenchymal and neuronal tissues were intermingled, and where immunohistochemical and ultrastructural investigations disclosed disruption of the basal lamina, prominent astrocytosis, and abundant axonal and synaptic profiles. These findings suggest that focal disturbances in cerebral cortical development occur in association with the development of lipomas.

Adult↗
PubMed2004

Frontotemporal dementia with co-occurrence of astrocytic plaques and tufted astrocytes, and severe degeneration of the cerebral white matter: a variant of corticobasal degeneration?

We report two patients who exhibited frontotemporal dementia (FTD) with unusual neuropathological features. The ages of the patients at death were 65 and 67 years, the disease durations were 6 and 5 years, and the clinical diagnoses were Pick's disease and corticobasal degeneration (CBD), respectively. At autopsy, both cases exhibited neuropathological findings compatible with those of CBD, including atrophy of the frontal and parietal lobes, neuronal loss and gliosis in the cortical and subcortical regions, and presence of cortical ballooned neurons and astrocytic plaques (APs). In both cases, immunoblotting of insoluble tau exhibited the pattern of selective accumulation of four-repeat tau, a finding that is also compatible with CBD. However, severe degeneration was evident in the frontal and parietal white matter in both cases. Moreover, a striking finding was the widespread presence in the affected cortex of tufted astrocytes (TAs), which are characteristic of progressive supranuclear palsy (PSP). Neither co-occurrence of APs and TAs nor severe degeneration of the cerebral white matter is a feature of either CBD or PSP. No mutations were found in the tau gene in either case. In conclusion, the possibility that these two cases represent a new neuropathological phenotype of non-familial FTD rather than simply a variant of CBD cannot be completely excluded.

Amyloid beta-Peptides↗
PubMed2004

Dose-dependent effects of methylmercury administered during neonatal brain spurt in rats.

Rapid brain growth occurs primarily during the third trimester in humans, whereas in rats it occurs after parturition. Therefore, we hypothesized that the effects of methylmercury (MeHg) on the postnatal developing rat nervous system may help in understanding the neurotoxicity on the human fetal brain when the brain is most vulnerable. In the present experiment, the dose-response effects of MeHg treatment during the postnatal developing phase in rats were studied. Male Wistar rats were orally administered 0, 1, 3, and 5 mg/kg/day methylmercury chloride (MMC), respectively, on postnatal day 1 and for 30 consecutive days. The body weight decline began from day 25 and typical symptoms, such as hind-limb crossing and ataxia, were observed in rats treated with 5 mg/kg/day MMC. The weight loss and typical symptoms were not observed in rats treated with 1 and 3 mg/kg/day. Mercury (Hg) concentrations in the brain were 2.6, 4.5, and 9.6 microg/g in the rats treated with 1, 3, and 5 mg/kg/day, respectively, on the day after the final MMC treatment. At 5 to 6 weeks of age, dose-dependent deficits of motor coordination in the rotarod test and learning disability in the passive avoidance response test were observed. Histopathological examination of a proportion of the MeHg-treated rats revealed widespread neuronal degeneration manifested by neuron loss and astrocytosis in the cerebral cortex, striatum, and cerebellum, where severity of the lesions seemed to increase in proportion to the administered dose of MMC. These findings using neonatal rats will be useful for better understanding of the effects of MeHg in the developing human brain during gestation.

Animals↗
PubMed2004

Familial amyotrophic lateral sclerosis with bulbar onset and a novel Asp101Tyr Cu/Zn superoxide dismutase gene mutation.

We describe a patient with familial amyotrophic lateral sclerosis (FALS) in whom we identified a novel missense mutation in exon 4 (Asp101Tyr) of the Cu/Zn superoxide dismutase (SOD1) gene. The disease started with a bulbar symptom (rapidly progressive hoarseness) and at autopsy showed degenerative changes restricted to the upper and lower motor neuron systems (more strictly, with lower motor predominance, showing the most severe degeneration in the nucleus ambiguus). Occasional intracytoplasmic Lewy-body-like hyaline inclusions that were immunoreactive for ubiquitin and SOD1, but immunonegative for neurofilament protein, were found in the lower motor neurons. This is the first report of hoarseness as the initial manifestation of FALS. This SOD1 gene mutation may be associated with a particular clinicopathological phenotype.

Amyotrophic Lateral Sclerosis↗
PubMed2004

Disclosure of a pro-apoptotic glyceraldehyde-3-phosphate dehydrogenase promoter: anti-dementia drugs depress its activation in apoptosis.

Overexpression and subsequent nuclear accumulation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is involved in neuronal apoptosis induced by several stimuli in which GAPDH antisense oligonucleotides specifically block the increment (2 approximately 3 fold) of GAPDH mRNA contents occurring prior to neuronal death. However, these agents do not affect the basal, constitutive mRNA contents. This suggests that there may be distinct gene regulations for GAPDH mRNA expression. Herein, we cloned two types of promoter regions upstream of this gene; viz., #104 (1.02-kb) and #302 (2.46-kb). These fragments were inserted into the pGL3 luciferase reporter system and transiently transfected into cultured cerebellar neurons undergoing cytosine arabinonucleoside-induced apoptosis. The functional analysis of these constructs revealed that #104, but not #302, increased luciferase activity in response to the apoptotic stimulus. Deletion and replacement mutation analysis of the #104 fragment disclosed the promoter core harbored between the 154-bp and 84-bp domains (3.5-fold activity of the control). Furthermore, anti-dementia drugs (such as Cognex and Aricept) markedly depress the expression of this pro-apoptotic GAPDH promoter activity. Interestingly, immunocytochemical examination of human post-mortem materials from patients with Alzheimer's disease revealed nuclear aggregated GAPDH in neurons of the affected brain regions, implying an association with apoptotic cell death. The current findings indicate that induction of the pro-apoptotic protein GAPDH is genetically regulated at the level of promoter activation, and this protein may be an important molecular target for developing anti-apoptotic therapeutic agents in certain neurological illnesses.

Aged↗
PubMed2004

CCL2, a product of mice early after systemic inflammatory response syndrome (SIRS), induces alternatively activated macrophages capable of impairing antibacterial resistance of SIRS mice.

Infection associated with systemic inflammatory response syndrome (SIRS) is a major cause of morbidity and mortality in patients with major surgery, polytrauma, and severe burn injury. In previous studies, mice with severe pancreatitis (a mouse model of SIRS, SIRS mice) have been shown to be greatly susceptible to various infections. In the present study, a mechanism involved in the impaired resistance of SIRS mice to infectious complications was investigated. Sera from SIRS mice impaired the resistance of normal mice to infectious complications induced by cecal ligation and puncture (CLP). CC chemokine ligand 2 (CCL2) was detected in sera of SIRS mice. Resident macrophages (RMphi) cultured with SIRS mouse sera converted to alternatively activated macrophages (AAMphi), which were also demonstrated in mice treated with recombinant murine CCL2. However, AAMphi were not demonstrated in mice injected with SIRS mouse sera and anti-CCL2 monoclonal antibody (mAb) in combination. Furthermore, normal mice that received SIRS mouse sera and anti-CCL2 mAb resisted CLP-induced infectious complications. These results indicate that the resistance of SIRS mice to infectious complications is impaired by AAMphi generated from RMphi in response to SIRS-associated CCL2 production.

Animals↗
PubMed2004

Alterations in human tau transcripts correlate with those of neurofilament in sporadic tauopathies.

Abnormalities in tau mRNA splicing cause frontotemporal dementia and parkinsonism linked to chromosome 17, and similar alterations are suggested in sporadic tauopathies such as progressive supranuclear palsy (PSP) or corticobasal degeneration (CBD). We have analyzed the expression of three-repeat (3R) and four-repeat (4R) tau isoforms in brains with familial and sporadic tauopathies. By RT-PCR analysis, decreased levels of 3R tau mRNA were detected not only in severely affected cases with progressive supranuclear palsy or corticobasal degeneration but also in cases with Alzheimer's disease or Pick's disease. Levels of 3R tau transcripts were closely correlated with levels of neurofilament transcripts. By contrast, expressions of glial fibrillary acidic protein and myelin basic protein were similar in all brains. These results suggest that decrease of 3R tau mRNA associated with loss of neuronal element may largely contribute to the increased ratio of 4R/3R tau mRNA in sporadic tauopathies.

Basic Helix-Loop-Helix Proteins↗
PubMed2004

A new modified DNA enzyme that targets influenza virus A mRNA inhibits viral infection in cultured cells.

DNA enzymes are RNA-cleaving single-stranded DNA molecules. We designed DNA enzymes targeting the PB2 mRNA translation initiation (AUG) region of the influenza A virus (A/PR/8/34). The modified DNA enzymes have one or two N3'-P5' phosphoramidate bonds at both the 3'- and 5'-termini of the oligonucleotides, which significantly enhanced their nuclease resistance. These modified DNA enzymes had the same cleavage activity as the unmodified DNA enzymes, determined by kinetic analyses, and reduced influenza A virus replication by more than 99%, determined by plaque formation. These DNA enzymes are highly specific; their protective effect was not observed in influenza B virus (B/Ibaraki)-infected Madin-Darby canine kidney cells.

Animals↗
PubMed2004

Accumulation of phosphorylated alpha-synuclein in the brain and peripheral ganglia of patients with multiple system atrophy.

We immunohistochemically examined the brain and peripheral sympathetic ganglia from eight patients with multiple system atrophy (MSA), using an antibody specific for phosphorylated alpha-synuclein (anti-PSer129). Phosphorylated alpha-synuclein was deposited in five cellular locations: oligodendroglial cytoplasm and nucleus, and neuronal cytoplasm, processes and nucleus. Many neuronal cytoplasmic inclusions (NCIs) were found in the pontine and inferior olivary nuclei and, to a lesser extent, in the substantia nigra, locus ceruleus, and neocortical and hippocampal neurons. NCIs were also found in the sympathetic ganglia in two out of the eight cases. Moreover, anti-PSer129 immunohistochemistry revealed extensive neuropil pathology; swollen neurites were abundant in the pontine nucleus, delicate neurites were observed in the deeper layers of the cerebral cortex and thalamus, and neuropil threads and dot-like structures were distributed in the basal ganglia and brainstem. Diffuse neuronal cytoplasmic staining (pre-NCI) was frequently found in the pontine and inferior olivary nuclei. Thus, the widespread accumulation of phosphorylated alpha-synuclein in both glial and neuronal cells is a pathological feature in patients suffering from MSA.

Aged↗
PubMed2004

Aprataxin, the causative protein for EAOH is a nuclear protein with a potential role as a DNA repair protein.

Early-onset ataxia with ocular motor apraxia and hypoalbuminemia (EAOH) is an autosomal recessive neurodegenerative disorder characterized by early-onset ataxia, ocular motor apraxia, and hypoalbuminemia. Recently, the causative gene for EAOH, APTX, has been identified. Of the two splicing variants of APTX mRNA, the short and the long forms, long-form APTX mRNA was found to be the major isoform. Aprataxin is mainly located in the nucleus, and, furthermore, the first nuclear localization signal located near the amino terminus of the long-form aprataxin is essential for its nuclear localization. We found, based on the yeast two-hybrid and coimmunoprecipitation experiments, that the long-form but not the short-form aprataxin interacts with XRCC1 (x-ray repair cross-complementing group 1). Interestingly the amino terminus of the long-form aprataxin is homologous with polynucleotidekinase-3'-phosphatase, which has been demonstrated to be involved in base excision repair, a subtype of single-strand DNA break repair, through interaction with XRCC1, DNA polymerase beta, and DNA ligase III. These results strongly support the possibility that aprataxin and XRCC1 constitute a multiprotein complex and are involved in single-strand DNA break repair, and furthermore, that accumulation of unrepaired damaged DNA underlies the pathophysiological mechanisms of EAOH.

Alternative Splicing↗
PubMed2004

SCA17 homozygote showing Huntington's disease-like phenotype.

We report a homozygous case of spinocerebellar ataxia type 17 with 48 glutamines. The age of the patient at disease onset was not lower than those of heterozygotes with the same CAG-repeat sizes, but the clinical manifestations were rapidly progressive dementia and chorea. Neuronal loss was relatively restricted and most prominent in the Purkinje cell layer and striatum; however, intranuclear neuronal polyglutamine accumulation was widespread, with a high frequency in the cerebral cortex and striatum.

Adult↗
PubMed2004

Increased norepinephrine production associated with burn injuries results in CCL2 production and type 2 T cell generation.

The susceptibility of thermally injured mice (TI-mice) to various infections is markedly influenced by burn-associated type 2 T cell responses, which are common with severe thermal injuries. Previously, we have reported that CC chemokine ligand 2/monocyte chemoattractant protein-I (CCL2) is produced in mice within 1 day of thermal injury, and the subsequent development of burn-associated type 2 T cell responses are triggered by this chemokine produced early after thermal injury. In this study, influence of norepinephrine (NE) on CCL2 production in mice early after thermal injury (TI) was investigated. Peripheral blood mononuclear cells (PBMC) and peritoneal macrophages (PMphi) from TI-mice produced CCL2, but the same cell preparations from normal mice did not. This chemokine was not produced by PBMC and PMphi from TI-mice previously treated with 6-hydroxydopamine (6-OHDA), which destroys sympathetic nerve termini. NE production was increased in circulation of TI-mice, and treatment of TI-mice with 6-OHDA resulted in the inhibition of NE secretion. When PBMC from normal mice were treated with NE, they acquired the ability to produce CCL2. Splenic T cells from TI-mice produced IL-4 into their culture fluids, while the cytokine was not produced by splenic T cells from TI-mice previously treated with 6-OHDA. These results indicate that NE may have an important role on early CCL2 production and the subsequent development of burn-associated type 2 T cell responses.

Animals↗