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Y Ihara

Publications and source records attributed to Y Ihara.

At least 73 records · Page 4Linked to original sources
PubMed1999

Appearance of sodium dodecyl sulfate-stable amyloid beta-protein (Abeta) dimer in the cortex during aging.

We previously noted that some aged human cortical specimens containing very low or negligible levels of amyloid beta-protein (As) by enzyme immunoassay (EIA) provided prominent signals at 6 approximately 8 kd on the Western blot, probably representing sodium dodecyl sulfate (SDS)-stable Abeta dimer. Re-examination of the specificity of the EIA revealed that BAN50- and BNT77-based EIA, most commonly used for the quantitation of Abeta, capture SDS-dissociable Abeta but not SDS-stable Abeta dimer. Thus, all cortical specimens in which the levels of Abeta were below the detection limits of EIA were subjected to Western blot analysis. A fraction of such specimens contained SDS-stable dimer at 6 approximately 8 kd, but not SDS-dissociable A(beta) monomer at approximately 4 kd, as judged from the blot. This A(beta) dimer is unlikely to be generated after death, because (i) specimens with very short postmortem delay contained the A(beta) dimer, and (ii) until 12 hours postmortem, such SDS-stable A(beta) dimer is detected only faintly in PDAPP transgenic mice. The presence of A(beta) dimer in the cortex may characterize the accumulation of A(beta) in the human brain, which takes much longer than that in PDAPP transgenic mice.

Adult↗
PubMed1999

Stable expression in Chinese hamster ovary cells of mutated tau genes causing frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17).

Extensive neuronal loss and aggregation of tau as cytoplasmic inclusions in neurons and glial cells in selected cortical and subcortical regions is the most striking characteristic of frontotemporal dementia and parkinsonism linked to chromosome 17, which is caused by exonic or intronic mutations in the tau gene. Here, we examined the effects of four exonic mutations in four-repeat tau using stably transfected Chinese hamster ovary cells. The proportion of polymerized tubulin was the largest in the P301L transfectant. G272V and P301L transfectants showed greater instability of microtubules in the presence of Colcemid than wild-type tau, V337M, or R406W transfectants. Thus no distinct phenotypes were shared by the mutant tau transfectants with regard to microtubule assembly and stability. Unexpectedly, R406W showed low and negligible levels of phosphorylation at Thr 231 and Ser 396, respectively, in the transfectant. This presents a sharp contrast to the observation that tau aggregates in R406W-affected brains are heavily phosphorylated at these two sites. This result suggests that hyperphosphorylation at these sites cannot occur in the tau R406W bound to microtubules, and thus that the hyperphosphorylated species of tau may be generated only after disruption of microtubules.

Animals↗
PubMed1999

Induction of interleukin (IL)-6 by hypoxia is mediated by nuclear factor (NF)-kappa B and NF-IL6 in cardiac myocytes.

OBJECTIVES: We have previously reported that interleukin (IL)-6 is induced by hypoxic stimulation in cardiac myocytes. In the present study, we examined the induction of potent transcription factors of IL-6, nuclear factor (NF)-kappa B and NF-IL6, in cardiac myocytes subjected to hypoxia. METHODS: Five different lengths of IL-6 promoter-luciferase reporter plasmids and three mutant plasmids, in which the binding sites of NF-kappa B and/or NF-IL6 were disrupted, were transfected into neonatal rat cardiac myocytes. Luciferase activities after hypoxic stimulation were measured. Electrophoretic mobility shift assays were performed using oligonucleotides containing the binding site for NF-kappa B or NF-IL6 as a probe. RESULTS: Hypoxic stimulation for 4 h increased luciferase activity by 5.7 fold in -179/+12-luciferase reporter plasmid, whereas no significant increase was observed in -60/+12-luciferase plasmid. Decrease in luciferase activity was more prominent when the NF-kappa B binding site was disrupted rather than when the NF-IL6 binding site was disrupted. Moreover, when both sites were disrupted, luciferase activity increased only by 1.5 fold. Electrophoretic mobility shift assays demonstrated enhanced binding activity to oligonucleotides containing the NF-kappa B binding site in hypoxic cardiac myocytes, which displayed a supershift with antibody to its subunit, p50 or p65. The binding activity to the NF-IL6 probe also enhanced and displayed a supershift with antibody to NF-IL6. CONCLUSIONS: Although hypoxic stimulation induced NF-kappa B and NF-IL6 in cardiac myocytes, NF-kappa B may be the primary positive regulator of transcriptional activation of the IL-6 gene in the context of hypoxia.

Animals↗
PubMed1999

Loss of heterozygosity of 14q32 in colorectal carcinoma.

Previous allelotyping studies on colorectal carcinoma suggest that loss of heterozygosity (LOH) on chromosome 14q may be a common genetic alteration in this tumor type. The purpose of this study was to determine precise frequency of LOH at 14q32 region in colorectal carcinomas and to define a minimal region of LOH. LOH at 14q32 in 66 primary colorectal carcinomas were analyzed by polymerase chain reaction-based deletion mapping using six highly polymorphic microsatellites. The average rate of informative data was 68.2% in all cases of colorectal carcinoma analyzed. The average rate of LOH at 14q32 was 44.1%, with the highest frequency of 50.0% at D14S267 locus in the informative cases. The minimal region of LOH was defined by markers D14S65 and D14S250 at 14q32. The present data provide useful information for the isolation of tumor suppressor genes from this region.

Carcinoma↗
PubMed1999

Calnexin discriminates between protein conformational states and functions as a molecular chaperone in vitro.

Although calnexin is thought to function as a molecular chaperone for glycoproteins, a prevalent view is that it cannot distinguish between protein conformational states, binding solely through its lectin site to monoglucosylated oligosaccharides. Using purified components in vitro, calnexin effectively prevented the aggregation not only of glycoproteins bearing monoglucosylated oligosaccharides but also proteins lacking N-glycans, an effect enhanced by ATP. It also suppressed the thermal denaturation of nonglycosylated proteins and enhanced their refolding in conjunction with other cellular components. Calnexin formed stable complexes with unfolded conformers of these proteins but not with the native molecules. Therefore, in addition to being a lectin, calnexin functions as a bona fide molecular chaperone capable of interacting with polypeptide segments of folding glycoproteins.

Adenosine Triphosphate↗
PubMed1999

Hyperglycemia causes oxidative stress in pancreatic beta-cells of GK rats, a model of type 2 diabetes.

Reactive oxygen species are involved in a diversity of biological phenomena such as inflammation, carcinogenesis, aging, and atherosclerosis. We and other investigators have shown that the level of 8-hydroxy-2'-deoxyguanosine (8-OHdG), a marker for oxidative stress, is increased in either the urine or the mononuclear cells of the blood of type 2 diabetic patients. However, the association between type 2 diabetes and oxidative stress in the pancreatic beta-cells has not been previously described. We measured the levels of 8-OHdG and 4-hydroxy-2-nonenal (HNE)-modified proteins in the pancreatic beta-cells of GK rats, a model of nonobese type 2 diabetes. Quantitative immunohistochemical analyses with specific antibodies revealed higher levels of 8-OHdG and HNE-modified proteins in the pancreatic beta-cells of GK rats than in the control Wistar rats, with the levels increasing proportionally with age and fibrosis of the pancreatic islets. We further investigated whether the levels of 8-OHdG and HNE-modified proteins would be modified in the pancreatic beta-cells of GK rats fed with 30% sucrose solution or 50 ppm of voglibose (alpha-glucosidase inhibitor). In the GK rats, the levels of 8-OHdG and HNE-modified proteins, as well as islet fibrosis, were increased by sucrose treatment but reduced by voglibose treatment. These results indicate that the pancreatic beta-cells of GK rats are oxidatively stressed, and that chronic hyperglycemia might be responsible for the oxidative stress observed in the pancreatic beta-cells.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1999

[Multiple risk factor syndrome].

Insulin resistance predisposes to the development of a cluster of abnormalities including glucose intolerance, hyperlipidemia, and hypertension, which may relate to the excess intra-abdominal fat deposits. The cluster of abnormalities increases a risk of atherosclerosis and comprises multiple risk factor syndrome (MRFS). Though many suspectable candidate genes of MRFS was analyzed, there is no compelling candidate gene by itself for MRFS. Simultaneous existence of polygenic and environmental factors may induce MRFS resulting in atherosclerosis.

Glucose Intolerance↗
PubMed1998

Transcriptional repressors are increased in pancreatic islets of type 2 diabetic rats.

To further clarify the mechanism of impaired insulin gene transcription in the diabetic state, we investigated the expression and function of the transcriptional repressor CREM (CRE modulator) in rat pancreatic islets. The CREM gene generates both transcriptional activators and repressors by alternative splicing and an intronic promoter. We isolated a novel alternatively spliced CREM isoform, CREM-17X, which efficiently represses insulin gene transcription, in addition to the three previously reported repressors. We also compared mRNA levels of insulin and the CREM repressors in pancreatic islets of Wistar and GK (Goto-Kakizaki) rats, the well-characterized spontaneous animal model of type 2 diabetes. The CREM repressor levels are increased, and the expression of insulin mRNA is decreased in GK rats, suggesting that increased CREM repressor expression in pancreatic islets could contribute to the decreased insulin gene transcription that results in impaired insulin secretion in type 2 diabetes.

Animals↗
PubMed1998

The presence of amyloid beta-protein in the detergent-insoluble membrane compartment of human neuroblastoma cells.

To investigate the intracellular compartmentalization of amyloid beta-protein (Abeta), human neuroblastoma SH-SY5Y cells were fractionated and the Abeta content in each fraction was quantitated by the well-characterized two-site enzyme-linked immunosorbent assay (ELISA). Subcellular fractionation of the cell revealed two distinct pools of Abeta within the cells: a Triton-soluble and a Triton-insoluble pools with the latter being larger than the former. Because Triton insolubility points to caveolae-like domains, we prepared detergent-insoluble, low-density membrane domains from SH-SY5Y cells using two different protocols. The low-density membrane fraction prepared by either protocol was found to contain a substantial proportion of intracellular Abeta40 and Abeta42. These results indicate that the distinct membrane domains are involved in the generation and/or trafficking of Abeta.

Amyloid beta-Peptides↗
PubMed1998

Bisecting GlcNAc structure is implicated in suppression of stroma-dependent haemopoiesis in transgenic mice expressing N-acetylglucosaminyltransferase III.

Several sugar structures have been reported to be necessary for haemopoiesis. We analysed the haematological phenotypes of transgenic mice expressing beta-1,4 N-acetylglucosaminyltransferase III (GnT-III), which forms bisecting N-acetylglucosamine on asparagine-linked oligosaccharides. In the transgenic mice, the GnT-III activity was elevated in bone marrow, spleen and peripheral blood and in isolated mononuclear cells from these tissues, whereas no activity was found in these tissues of wild-type mice. Stromal cells after long-term cultures of transgenic-derived bone marrow and spleen cells also showed elevated GnT-III activity, compared with an undetectable activity in wild-type stromal cells. As judged by HPLC analysis, lectin blotting and lectin cytotoxicity assay, bisecting GlcNAc residues were increased on both blood cells and stromal cells from bone marrow and spleen in transgenic mice. The transgenic mice displayed spleen atrophy, hypocellular bone marrow and pancytopenia. Bone marrow cells and spleen cells from transgenic mice produced fewer haemopoietic colonies. After lethal irradiation followed by bone marrow transplantation, transgenic recipient mice showed pancytopenia compared with wild-type recipient mice. Bone marrow cells from transgenic donors gave haematological reconstitution at the same level as wild-type donor cells. In addition, non-adherent cell production was decreased in long-term bone marrow cell cultures of transgenic mice. Collectively these results indicate that the stroma-supported haemopoiesis is compromised in transgenic mice expressing GnT-III, providing the first demonstration that the N-glycans have some significant roles in stroma-dependent haemopoiesis.

Acetylglucosamine↗
PubMed1998

Collapsin response mediator protein-2 is associated with neurofibrillary tangles in Alzheimer's disease.

Intraneuronal accumulation of paired helical filaments (PHF) is considered to be closely related to the neuronal loss observed in brains of patients affected with Alzheimer's disease. The central issue is whether PHF formation itself causes or accelerates the neuronal perikaryal and neuritic degeneration or whether they are simply the consequence of preceding degeneration. We sought to address the issue in part by characterizing the PHF-associated molecules and thus raised a number of monoclonal antibodies to neurofibrillary tangles. One monoclonal antibody, 3F4, strongly reacted with neurofibrillary tangles and some plaque neurites but few neuropil threads. This monoclonal antibody labeled a 65-kDa protein, but not tau or ubiquitin, on a Western blot of human brain extract and immunoprecipitated the same protein. The peptides released from the purified 65-kDa protein had the same sequences as those of a newly identified protein, human collapsin response mediator protein-2. Incorporation into neurofibrillary tangles may deplete soluble, cytosolic human collapsin response mediator protein-2 and lead to abnormal neuritic and/or axonal outgrowth of the tangle-bearing neuron, thus accelerating the neuritic degeneration in Alzheimer's disease.

Alzheimer Disease↗
PubMed1998

Ectopic expression of N-acetylglucosaminyltransferase III in transgenic hepatocytes disrupts apolipoprotein B secretion and induces aberrant cellular morphology with lipid storage.

N-Acetylglucosaminyltransferase III (GnT-III) produces "bisecting-GlcNAc" and regulates the branching of N-glycans. GnT-III activity is elevated during hepatocarcinogenesis, which is in contrast to the undetectable level found in normal hepatocytes. To determine the biological significance of GnT-III in hepatocytes, transgenic mice that specifically express GnT-III in the liver were established and characterized. The transgenic hepatocytes had a swollen oval-like morphology, with many lipid droplets. Apolipoprotein B, which contained increased level of bisecting-GlcNAc accumulated in the transgenic hepatocytes. In the transgenic serum, triglycerides, the beta- and pre-beta-lipoprotein fractions, and apolipoprotein B100 were significantly decreased, compared with levels in nontransgenic serum. These abnormal phenotypes were more prominent in the mice with more copies of the transgene and a resulting high GnT-III activity. We demonstrate that aberrant glycosylation, as the direct result of the formation of bisecting-GlcNAc, disrupts the function of apolipoprotein B, leading to the generation of fatty liver. This observation suggests a novel mechanism for the pathogenesis of fatty liver.

Animals↗
PubMed1998

Diffuse plaques associated with astroglial amyloid beta protein, possibly showing a disappearing stage of senile plaques.

To clarify whether senile plaques disappear, we examined amyloid beta protein (A beta) deposits in non-demented subjects, and found novel diffuse plaques associated with astroglial A beta. Formalin-fixed paraffin-embedded sections from cortical areas were immunolabeled with a panel of A beta antibodies, and astroglial and microglial markers. Cerebral A beta deposition was primarily found as diffuse plaques (DP) in these subjects. A subset of DP was associated with clusters of intensely A beta-positive small granules. The clusters, which were located just adjacent to astroglial nucleus, had the characteristics of lipofuscin granules and, therefore, were quite different from "small stellate deposits". Substantial amounts of A beta-positive granules were found inside astrocytes by dual labeling of A beta and glial fibrillary acid protein, and the majority of astroglial A beta immunoreactivity was located on lipofuscin granules. A beta-positive granules lacked immunoreactivity with antisera for the N-terminal region of A beta. These peculiar DP showed a much weaker staining than ordinary DP. The DP associated with astroglial A beta were found in about one third of the subjects, although the density varied widely among individuals. From these findings, we propose that DP, which are associated with the N-terminal truncated A beta in astrocytes, represent the disappearing stage of senile plaques.

Aged↗
PubMed1998

Chloroquine myopathy suggests that tau is degraded in lysosomes: implication for the formation of paired helical filaments in Alzheimer's disease.

We have found that amorphous tau deposits in chloroquine myopathy (CM), a vacuolar myopathy induced by the administration of chloroquine, a well-known lysosomotropic agent. The dynamics of tau in CM and immunocytochemistry strongly suggest that the accumulation of tau is due to defective tau degradation in the lysosomal compartment in the muscle. This observation may offer a new view on the formation of paired helical filaments in Alzheimer's disease: this selective protein degradation pathway may be defective and result in intracellular accumulation of tau, thereby forming the unusual filaments.

Alzheimer Disease↗
PubMed1998

Prenatal diagnosis of prune belly syndrome at 12 weeks of pregnancy: case report and review of the literature.

We present a case of prune belly syndrome in a 12-week fetus whose previous anomaly scan at 10 weeks had been normal. The ultrasound diagnosis was based on the findings of a lower abdominal cystic echo caused by abnormal dilatation of the bladder. Termination was performed at 14 weeks and autopsy confirmed the distended bladder. In addition, there was bilateral hydronephrosis and an absence of abdominal muscles, liver, spleen and diaphragm. A review of the literature indicates that ours may be the earliest reported case of prune belly syndrome.

Adult↗