Biomedical subjects

Y Tashiro

Publications and source records attributed to Y Tashiro.

At least 55 records · Page 3Linked to original sources

PubMed1996

Membrane topology and retention of microsomal aldehyde dehydrogenase in the endoplasmic reticulum.

Microsomal aldehyde dehydrogenase (msALDH) is anchored to the endoplasmic reticulum (ER) membrane by the hydrophobic domain at its carboxyl terminus, and most of the molecule is exposed to the cytoplasm (Masaki, R., Yamamoto, A., and Tashiro, Y.(1994) J. Cell Biol. 126, 1407-1420). To determine the membrane topology and the intracellular localization of msALDH, the amino-terminal region of bovine opsin containing N-glycosylation sites was fused to the carboxyl terminus of msALDH, and three chimeric proteins with extensions of different sizes were expressed in COS cells. Indirect immunofluorescence microscopy showed the ER localization of all of the chimeric proteins similar to wild-type msALDH. Immunoblotting revealed that the two chimeric proteins containing longer extensions, those with the N-glycosylation site at distances of 13 and 21 amino acids from the membrane anchor, respectively, were glycosylated. These results indicate that the membrane binding domain of msALDH spans the bilayer of the ER. The carbohydrate chain of the chimeras was sensitive to endoglycosidase H but resistant to endoglycosidase D. Upon treatment of transfected COS cells with brefeldin A, the carbohydrate chain was processed to an endoglycosidase H-resistant form, presumably by cis/medial Golgi-specific enzymes redistributed in the ER. These biochemical results in addition to immunofluorescence microscopic observations suggest that msALDH is retained in the ER by blockading of the exit from the ER.

Aldehyde Dehydrogenase↗

PubMed1996

Synthetic nucleosides and nucleotides. 37. Antisense oligodeoxynucleotides bearing 5-(phenylethyl)-2'-deoxyuridylate at the 3'-terminus: exonuclease-resistant molecule with natural phosphodiester backbone.

5-(Phenylethyl)-2'-deoxyuridine has been incorporated into an oligodeoxynucleotide (ODN) by using normal cyanoethyl phosphoramidite chemistry on a DNA synthesizer. For introduction of the modified residue at the 3'-end position of the ODN, we designed and synthesized a new nucleoside phosphoramidite derivative, which connected the 3'-hydroxyl group and phosphoramidite moiety by an alkaline-labile linker. The 3'-end could be substituted in ODNs by using commercially available supports as a starting material following standard NH4OH treatment. The ODN carrying 5-(phenylethyl)-2'-deoxyuridine at the 3'-end position showed about 3-fold resistance to nucleolytic degradation in human plasma without precluding its specific base-pairing activity.

Deoxyuridine↗

PubMed1996

Inhibition of vesicle-mediated protein transport by nordihydroguaiaretic acid.

Nordihydroguaiaretic acid (NDGA) blocks intra-Golgi protein transport in a cell-free system and prolactin secretion from GH3 cells [Tagaya, M., Henomatsu, N., Yoshimori, T., Yamamoto, A., Tashiro, Y., and Fukui, T. (1993) FEBS Lett. 324, 201-204]. To determine which intracellular secretory pathway(s) is inhibited by NDGA, we investigated its effect on the transport of the vesicular stomatitis virus-encoded glycoprotein in BHK-21 cells. NDGA blocked protein transport from the endoplasmic reticulum to the Golgi apparatus, and from the trans-Golgi network to the plasma membrane. In addition, it retarded the brefeldin A-induced retrograde transport of mannosidase II to the endoplasmic reticulum. Although NDGA had an inhibitory effect on protein synthesis, it induced the expression of BiP, a chaperone located in the endoplasmic reticulum. The induction of BiP may be a consequence of the inhibition of protein transport by NDGA.

Animals↗

PubMed1996

Formation of crystalloid endoplasmic reticulum in COS cells upon overexpression of microsomal aldehyde dehydrogenase by cDNA transfection.

When rat liver microsomal aldehyde dehydrogenase (msALDH) was overexpressed in COS-1 cells by cDNA transfection, large granular structures containing both msALDH and endogenous protein disulfide isomerase appeared (Masaki et al. (1994) J. Cell Biol. 126, 1407-1420). Confocal laser microscopy revealed that these granular structures are dispersed throughout the cytoplasm. Electron microscopy showed that the structures are composed of regularly arranged crystalloid smooth endoplasmic reticulum (ER). The formation of the crystalloid ER was accompanied by a remarkable proliferation of smooth ER, which appeared occasionally continuous to the rough ER. We suggest that the smooth ER, proliferated from the rough ER, is transformed and assembled into the crystalloid ER by head-to-head association of the msALDH molecules on the apposed smooth ER membranes. In order to understand the molecular mechanism of the crystalloid ER formation, we asked which portions of the msALDH molecules are needed for the crystalloid ER formation by expressing deletion mutants or chimera protein of msALDH in COS-1 cells. The overexpression of msALDH molecules lacking the stem region preceding the membrane spanning region, although they were exclusively localized in the ER, did not induce the formation of crystalloid ER. More detailed analysis showed that the amino acid sequence FFLL, located in the stem region, is necessary to form the crystalloid ER. The chimera protein containing the last 35 amino acids of msALDH at the carboxyl terminus of chloramphenicol acetyltransferase was localized to the ER, but did not induce the formation of the crystalloid ER. These results suggest that at least two regions, the bulky amino-terminal region and the FFLL sequence in the stem region of msALDH molecules are required for the formation of the crystalloid ER.

Aldehyde Dehydrogenase↗

PubMed1996

Localization of HPC-1/syntaxin 1 in developing rat cerebellar cortex.

In adult rat cerebellum, HPC-1/syntaxin 1 is detected at high density on the plasma membrane of the non-synaptic region of parallel fibers in addition to the synaptic terminal membranes and the synaptic vesicles (Koh, S., Yamamoto, A., Inoue, A., Inoue, Y., Akagawa, Y., Kawamura, Y., Kawamoto, Y., and Tashiro, Y. (1993). J. Neurocytology 22: 995-1005). To assess the possibility that HPC-1/syntaxin 1 participates in the morphogenesis of the nervous system, we examined changes in the localization of HPC-1/syntaxin 1 during postnatal development of the molecular layer of the rat cerebellum. HPC-1/syntaxin 1 appeared in the granule cells in the outer granule cell layer in 3-days-old rat cerebellum when the formation of synapses and the appearance of a synaptic vesicle protein, synaptophysin, had not yet been observed in the molecular layer. At this stage, the granule cells began to form parallel fibers. Confocal laser microscopy and immuno-electron microscopy showed that HPC-1/syntaxin 1 was localized on the extruding plasma membrane of the granule cells to form parallel fibers. In 8-days-old rats, synapses formed between the parallel fibers and the developing dendrites of Purkinje cells, and the HPC-1 immunoreactivity appeared on the axons of parallel fibers and on the synapses. In 21-days-old rats, the HPC-1/syntaxin is involved in the formation of the molecular layer, especially in the axonal growth of the parallel fibers.

Animals↗

PubMed1996

Establishment of a series of alien monosomic addition lines of Japanese bunching onion (Allium fistulosum L.) with extra chromosomes from shallot (A. cepa L. aggregatum group).

Forty one plants of alien monosomic addition lines of Allium fistulosum L. with extra chromosomes from A. cepa L. Aggregatum group (FF + nA) were produced through the second backcross of amphidiploids between these two species to A. fistulosum. Identification of the extra chromosomes in the 16 plants by elaborate karyotype analyses indicate that a complete series (eight different types) of the alien monosomic addition lines was established in Allium for the first time in this study. Chromosomal locations of malate dehydrogenase (MDH) gene, triosephosphate isomerase (TPI) gene and 5S rDNA of A. cepa Aggregatum group were determined using the series; The gene locus Mdh-1 was located on 4A, Tpi-1 on 3A and a 5S rDNA locus on 7A. Our previous and present studies using the alien monosomic addition lines revealed 11 genetic markers (isozyme and 5S rDNA) assigned to all eight chromosomes of A. cepa Aggregatum group, and these markers reconfirmed the completion of the series. Extra chromosomes of 25 other plants were examined by means of simple analyses of the chromosome markers and karyotypes. Of the total 41 plants, frequencies of the alien monosomic addition lines with extra chromosomes 1A to 8A were as follows: 1A, 5 plants; 2A, 3; 3A, 5; 4A, 9; 5A, 4; 6A, 2; 7A, 11; and 8A, 2.

Chromosome Mapping↗

PubMed1996

Role of endocrine cell microvesicles in intercellular chemical transduction.

Microvesicles (MVs) in endocrine cells are morphologically similar to neuronal synaptic vesicles. MVs were shown to contain proteins involved in neurotransmitter storage such as vacuolar H(+)-ATPase and neurotransmitter transporters, and ones in vesicular trafficking such as synaptobrevins and N-ethylmaleimide-sensitive fusion protein. Isolated MVs accumulate cell-specific neurotransmitters in an energy-dependent manner. Upon stimulation, the MVs may fuse with the plasma membrane and secrete the internal neurotransmitters. Thus, endocrine cells possess an MV-mediated secretion system as an intercellular signal transducing system.

Animals↗

PubMed1996

[A histopathological study of corneal amyloidosis secondary to trichiasis].

We present a case of secondary corneal amyloidosis whose etiological mechanism was investigated by immunohistochemistry and electron microscopy. A 48-year-old woman had suffered from trichiasis in the right eye for 35 years, and developed secondary corneal amyloidosis, a phenomenon previously described but whose etiological mechanism has not been explained. Slitlamp examination of the cornea revealed a white excrescence with a diameter of 2 mm. The lesion was excised and examined by light and electron microscopy. Large deposits of an amorphous eosinophilic material were observed beneath the atrophic epithelium. Amyloid was detected in these deposits using Congo red stain, polarized light, and electron microscopy. Neither vascularization nor infiltration of inflammatory cells was observed. Immunohistochemical tests for protein AL, protein AA, prealbumin, beta 2-microglobulin and cytokeratin in paraffin sections were all negative. Characteristic findings were observed in the border zone between the basal cells and the deposits. Numerous digitiform cell processes and membrane-bound globular fragments of basal cells were seen in the superficial region of the deposits. The cell membrane of some globules was interrupted and the contents appeared to have been discharged into the stroma. These findings suggest that basal cells of the corneal epithelium provide an amyloid precursor on the stroma.

Amyloidosis↗

PubMed1996

[Lymphangiomyomatosis with chylous ascites treatment successfully by peritoneo-venous shunting].

A 49-year-old woman was admitted to our hospital because of coughing and dyspnea. A chest roentogenogram showed emphysematous changes and a diffuse reticular shadow. A high-resolution CT scan of the chest showed many small cysts throughout the lungs. Lymphangiomyomatosis was diagnosed after examination of a specimen obtained by transbronchial biopsy. Abdominal distention due to chylous ascites developed during the hospital stay despite anti-estrogen therapy. Because the ascites was resistant to conservative therapy, we decided to begin peritoneo-venous shunting with a Denver Shunt system. After the operation, the abdominal distention was controlled for 1 year and 11 months, at which time the patient died of respiratory and heart failure with pneumonia. At autopsy, the shunt was patent and functional although about 900 ml of serous ascites fluid was present. An adenocarcinoma was found in the upper lobe of the right lung, but it may not have been related to the lymphangiomyomatosis. Peritoneovenous shunting with a Denver Shunt can be used to treat chylous ascites due to lymphangiomyomatosis when conservative therapy is insufficient.

Adenocarcinoma↗

PubMed1995

Localization of N-ethylmaleimide-sensitive fusion protein in pinealocytes.

N-ethylmaleimide-sensitive fusion protein (NSF), a protein necessary for vesicular docking and/or fusion, was detected immunohistochemically in pinealocytes. NSF was distributed similarly to synaptophysin and vacuolar-type H(+)-ATPase (V-ATPase), marker proteins for synaptic-like microvesicles (MVs) abundantly present in pinealocytes. A subcellular fractionation study indicated that .> 95% of NSF was present as a membrane-bound form and that some NSF was associated with MVs. Like neuronal NSF, the protein was not solubilized from membranes with either 2 mM Mg-ATP or 2% sodium carbonate, suggesting that NSF was tightly bound to the membranes. NSF was also detected in purified MVs from bovine posterior pituitaries. Since MVs are the organelles in which transmitters are stored, these results suggest that NSF is involved in the MV-mediated exocytosis of transmitters from endocrine cells.

Adrenal Medulla↗

PubMed1995

Lack of p53 point mutations in chemically induced mouse hepatoblastomas: an end-stage, highly malignant hepatocellular tumor.

Inactivation of the p53 tumor suppressor gene appears to be an important event in the progression of many types of human neoplasms; however its role in rodent experimental tumorigenesis is controversial. Previous studies have shown that a wide array of chemically induced and spontaneous mouse liver tumors lack p53 mutations within the evolutionarily conserved regions of exons 5-8. However, since p53 inactivation in human neoplasms occurs relatively late in tumor progression, it is possible that the mouse liver tumors evaluated previously were not suitably advanced to incur p53 aberrations. In the present study, we examined an end-stage, highly malignant embryonal mouse liver tumor known as the hepatoblastoma (HB) for p53 mutations utilizing the highly sensitive 'cold' single-strand conformation polymorphism (SSCP) technique. In addition, several of the HBs were examined by direct nucleotide sequencing. No aberrations of the p53 gene were detected within exons 5-8 of any of the 16 HBs examined. These results confirm that the p53 gene plays a minimal role in the development or malignant progression of hepatocellular tumors in mice.

Animals↗

PubMed1995

Microvesicles isolated from bovine posterior pituitary accumulate norepinephrine.

Histochemical study indicated that the posterior pituitary possesses numerous microvesicles (MVs) containing synaptophysin, a marker protein specific for brain synaptic vesicles (Navone, F., Di Gioia, G., Jahn, R., Browning, M., Greengard, P., and De Camilli, P. (1989) J. Cell Biol. 109, 3425-2433). By monitoring cross-reactivity with anti-synaptophysin antibody, the MVs were highly purified from bovine posterior pituitaries by a combination of differential and sucrose density gradient centrifugations. The purified MVs had an average diameter of about 60 nm and were associated with synaptophysin as revealed by immunoelectron microscopy. The vesicles contained ATPase activity partially sensitive to bafilomycin A1 and to vanadate. The membrane fraction immunoisolated with anti-synaptophysin antibody also exhibited similar ATPase activity. The two ATPases could be purified separately; the vandate-sensitive enzyme was identified as a 115-kDa polypeptide immunochemically similar to chromaffin granule P-ATPase (forming phosphoenzyme intermediate), and the bafilomycin A1-sensitive ATPase showed essentially the same properties as those of vacuolar type H(+)-ATPases. Upon addition of ATP, the MVs formed an electrochemical gradient of protons and took up norepinephrine in a reserpine-sensitive manner, indicating the presence of secondary monoamine transporter coupled with vacuolar type H(+)-ATPase. No uptake of L-glutamate, gamma-aminobutyrate, glycine, or acetylcholine was observed. The identification of MVs as organelles responsible for storage of monoamines is important for understanding the physiological function of the posterior pituitary.

Acetylcholine↗

PubMed1995

Neurotransmitters partially restore glucose sensitivity of insulin and glucagon secretion from perfused streptozotocin-induced diabetic rat pancreas.

To elucidate the mechanisms of insensitivity of hormone secretion to glucose in streptozotocin-induced diabetic rat islets, we investigated the effects of acetylcholine (ACh) and norepinephrine on insulin and glucagon secretion in response to changes in glucose concentration, using perfused pancreas preparations. Basal insulin secretion at a blood glucose level of 5.6 mmol/l was significantly higher and basal glucagon secretion significantly lower in streptozotocin-induced diabetic rats than in controls, and neither high (16.7 mmol/l) nor low (1.4 mmol/l) blood glucose concentrations influenced insulin or glucagon secretion. Addition of 10(-6) mol/l ACh to the perfusate increased glucose-stimulated insulin secretion. Also, 10(-6) mol/l ACh, 10(-7) mol/l norepinephrine, as well as a combination of both, induced marked glucagon secretion, this was suppressed by high blood glucose level. Although simultaneous addition of 10(-6) mol/l ACh and 10(-7) mol/l norepinephrine induced only a slight increase in glucagon secretion in response to glucopenia, there was a significant increase in glucagon secretion in conjunction with an ambient decrease in insulin. Histopathological examination revealed a marked decline in acetylcholinesterase and monoamine-oxidase activities in the islets of streptozotocin-induced diabetic rats. We speculate that reduction of the potentiating effects of ACh and norepinephrine lessens glucose sensitivity of islet beta and alpha cells in this rat model of diabetes.

Acetylcholine↗

PubMed1995

Effects of fibrin on the angiogenesis in vitro of bovine endothelial cells in collagen gel.

The effect of fibrin on angiogenesis in vitro was investigated using an experimental model of tube formation by bovine capillary endothelial cells (BCEs) in type I collagen gel. One milligram per milliliter of fibrin added into type I collagen gel significantly increased the length of the tubular structures formed by BCEs in the gel by about 180% compared with type I collagen only. The facilitating effect of fibrin on tube formation by BCEs was inhibited by either anti-basic fibroblast growth factor (bFGF) IgG (25 micrograms/ml) or anti-urokinase type plasminogen activator (uPA) IgG (10 micrograms/ml) added to the gel and culture medium, but not by anti-tissue type plasminogen activator (uPA) IgG (10 micrograms/ml) added to the gel and culture medium, but not by anti-tissue type plasminogen activator (10 micrograms/ml) or non-immune IgG. The Arg-Gly-Asp (RGD) containing peptides (100 micrograms/ml) added to the culture medium also suppressed tube formation by BCEs in fibrin-containing type I collagen gel, but not in type I collagen gel. These results suggest that the increased release of bFGF and uPA by BCEs therefore plays a role in the angiogenic effect of fibrin in vitro, and the angiogenic effect of fibrin is mediated by the RGD sequence in fibrin, probably via the function of integrin receptor of the BCEs.

Amino Acid Sequence↗

PubMed1995

A subpopulation of large ganglion neurons express IsK protein mRNA: an in situ hybridization analysis in the rat eye.

Rat IsK protein is a 130-amino acid membrane protein and subserves generation of K+ outward current. The expression of this putative K+ ion channel mRNA was explored in the eye with in situ hybridization. The ganglion cell layer of the retina expressed IsK mRNA in a subpopulation of ganglion cells composed of large cell bodies. The hybridization-positive cells were scattered throughout the retina. The hybridization signal was also localized to the epithelial cells throughout the cornea. The results provided evidence for IsK message in neuronal cells. Expression of IsK message in the eye was thus shown to be restricted to particular cell types.

Animals↗

PubMed1995

Defective insulin and glucagon secretion in isolated perfused pancreata of diabetic WBN/Kob rats.

To elucidate the pathophysiology of diabetes mellitus in male WBN/Kob rats, we performed pancreatic perfusion experiments and histopathological studies. Intraperitoneal glucose tolerance tests showed a diabetic pattern in 12-month-old WBN/Kob rats. In perfused pancreata of WBN/Kob rats, both the first and the second phases of insulin secretion in response to a 16.7 mM glucose challenge were markedly reduced compared with those in age-matched Wistar rats (p < 0.01, respectively). Furthermore, the insulin secretion rate in response to glucopenia (1.4 mM) was significantly higher (p < 0.05) and the decrement in insulin secretion was significantly lower (p < 0.05) in WBN/Kob rats. The decrement in glucagon secretion with 16.7 mM glucose was significantly blunted (p < 0.001), and the glucagon secretion rate in response to glucopenia was also significantly lower in WBN/Kob rats than in controls (p < 0.01). Although insulin secretion in response to 10 mM arginine was also moderately reduced in WBN/Kob rats (p < 0.05), the glucagon secretion rates in response to 10 mM arginine were similar in the two groups. Histopathological examination revealed widespread disappearance of acinar cells and islets, inflammatory changes, and marked fibrosis in the pancreata of WBN/Kob rats. Immunohistochemical studies showed decreased numbers of B cells in the islets of WBN/Kob rats. These findings suggest that this WBN/Kob rat strain is a useful model for studying not only pathogenesis, but also pathophysiology, i.e., defective hormonal secretion, in some types of human diabetes mellitus.

Animals↗

PubMed1995

Nasal glioma: an immunohistochemical and ultrastructural study.

A case of a 14 month old Japanese female infant presenting with nasal glioma is reported. The tumor had been noticed at the nasal radix since birth and had slowly and progressively enlarged. There was no communication between the tumor and the cranial cavity on radiological examination. The tumor was macroscopically anchored to the nasal septum by a fibrous stalk, and histologically consisted of nests or trabeculae of either polygonal or spindle cells with plump eosinophilic cytoplasm and oval nuclei, separated by vascular-rich connective tissue intermingled with multinucleated giant cells. These tumor cells were immunohistochemically positive for glial fibrillary acidic protein as well as for S-100 protein and vimentin. An electron microscopic examination revealed collagen fibers and basal lamina between the tumor cells and the fibroblasts. Tumor cells possessed abundant intermediate filaments, which showed occasional Rosenthal fiber-like structures, in their cytoplasm and processes. A few oligodendrocytes and cilia of 9 microtubule doublets either with or without 2 central microtubules were also noted. These clinicopathological findings suggested that this tumor was once an encephalo(meningo)cele, which probably degenerated as a result of the loss of intracranial communication and then appeared to be isolated from the intracranial tissue.

Brain↗

PubMed1995

Pulmonary oncocytoma: report of a case in conjunction with an immunohistochemical and ultrastructural study.

An intra-pulmonary mass in a 51 year old Japanese woman was incidentally discovered in the right middle lobe. The resected tumor was grossly well-demarcated, solid, light yellowish white in color and measured 3.0 x 2.0 x 1.5 cm in size. It was composed of a diffuse proliferation of large polygonal cells with an abundant, granular cytoplasm, and round to irregular nuclei with prominent nucleoli. Smaller eosinophilic cells with hyperchromatic nuclei and larger vacuolated cells were also observed. However, no mucin production was detected. There were neither argyrophilic nor argentaffin cells, and no serotonin-positive cells. They showed an immunoreactivity to cytokeratin and vimentin but not to alpha-actin. On electron microscopy, abundant microvilli, which have never been previously described in pulmonary oncocytomas, were observed. Occasional desmosomes and myelin figures as well as numerous mitochondria were also seen. No neurosecretory granules were present. These findings suggested that this tumor might have an epithelial origin from the bronchial serous gland with subsequent cellular degeneration.

Adenoma, Oxyphilic↗