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Biomedical subjects

K Takei

Publications and source records attributed to K Takei.

At least 91 records · Page 5Linked to original sources
PubMed1996

Immunogenicity and efficacy of a killed hepatitis A vaccine in day care center children.

The objective of this study was to characterize the immune response of children after the use of two different vaccine doses and to evaluate whether vaccination benefits children attending day care centers in areas with high anti-HAV seroprevalence. The study was conducted in a day care center with a stable population in São Paulo, Brazil. Two groups of 20 children, all seronegative for hepatitis A antibodies, were assigned randomly to receive three times 0.5 and 1.0 ml of the vaccine, the second and third dose 1 and 6 months after the first dose, respectively. There were 27 children in the control group. All children in both vaccinated groups had protective levels of antibodies in the serum after two inoculations, and serious adverse reactions were not observed. In the eighth month of follow-up, a hepatitis A outbreak occurred in the day care center. Five children in the control group had high titers of IgM class anti-HAV, four with clinical manifestations of acute hepatitis. None of the vaccinated children developed symptoms or signs of hepatitis (P = 0.0125), and the estimate of vaccine efficacy was 100%. Two nonstudy children from the center also had clinical and serological evidence of acute hepatitis A. It is concluded that vaccination represents an important method for prevention of hepatitis A transmission in day care centers. The results of this pilot study justify further testing in larger groups.

Child Day Care Centers↗
PubMed1996

Yeast protein translocation complex: isolation of two genes SEB1 and SEB2 encoding proteins homologous to the Sec61 beta subunit.

A yeast gene (cDNA clone) was isolated in a screen for suppressors of secretion-defective sec15-1 mutation. This gene encodes a protein homologous to the beta subunit of the mammalian Sec61 protein complex functioning in protein translocation into the endoplasmic reticulum (ER). The predicted protein, Seb1p, consists of 82 amino acids and contains one potential membrane-spanning region at the C-terminus but no N-terminal signal sequence. Seb1p shows 30% identity to the mammalian Sec61 beta subunit and 34% identity to the Arabidopsis thaliana Sec61 beta subunit. Overexpression of SEB1 from a multicopy plasmid suppressed the temperature sensitivity of sec61-2 and sec61-3 mutants. Immunofluorescence and immunoelectron microscopy indicated that Seb1p resides in the ER membranes with the hydrophilic N-terminus exposed to the cytoplasm. The in vitro translated Seb1p was post-translationally inserted into microsomal membranes. As the chromosomal disruption of the SEB1 gene was not lethal, potential homologous genes were screened by heterologous hybridization. The SEB1 homologue thus isolated, SEB2, encodes a protein 53% identical to Seb1p. Disruption of the chromosomal SEB2 was not lethal whereas the double disruption of SEB1 and SEB2 resulted in a temperature-sensitive phenotype. This study further emphasizes the evolutionary conservation of the ER protein translocation apparatus and provides novel genetic tools for its functional analysis.

Amino Acid Sequence↗
PubMed1996

Isolation and characterization of a cDNA that encodes maize uroporphyrinogen III methyltransferase, an enzyme involved in the synthesis of siroheme, which is prosthetic group of nitrite reductase.

A full-length cDNA clone, pZmSUMT1, encoding an S-adenosyl-L-methionine-dependent uroporphyrinogen III C-methyltransferase (SUMT; EC 2.1.1.107) of maize was isolated from a root cDNA library. pZmSUMT1 had an insert of 1.7 kb and the amino acid sequence deduced from the open reading frame of the cDNA was similar to that of SUMT from various bacteria and also to the SUMT catalytic region of siroheme synthase (cysG) from Escherichia coli. Overproduction of ZmSUMT1 in a cysG mutant of E. coli eliminated the requirement of the strain for cysteine. Transcripts for ZmSUMT1 accumulated rapidly in both rots and leaves in response to the addition of nitrate to the culture medium. The effects of biochemical inhibitors on the nitrate-dependent induction of the gene for ZmSUMT1 coincided with the effects on the genes for other nitrate-assimilatory enzymes, nitrate reductase and nitrite reductase. An import experiment in vitro suggested that the gene product might be located in plastids. The results together indicate that ZmSUMT1 might be involved in the synthesis of siroheme, a prosthetic group of nitrite reductase, and that the expression of its gene is co-regulated with that of other nitrate-assimilatory genes.

Amino Acid Sequence↗
PubMed1996

The synaptic vesicle cycle: a single vesicle budding step involving clathrin and dynamin.

Strong evidence implicates clathrin-coated vesicles and endosome-like vacuoles in the reformation of synaptic vesicles after exocytosis, and it is generally assumed that these vacuoles represent a traffic station downstream from clathrin-coated vesicles. To gain insight into the mechanisms of synaptic vesicle budding from endosome-like intermediates, lysed nerve terminals and nerve terminal membrane subfractions were examined by EM after incubations with GTP gamma S. Numerous clathrin-coated budding intermediates that were positive for AP2 and AP180 immunoreactivity and often collared by a dynamin ring were seen. These were present not only on the plasma membrane (Takei, K., P.S. McPherson, S.L.Schmid, and P. De Camilli. 1995. Nature (Lond.). 374:186-190), but also on internal vacuoles. The lumen of these vacuoles retained extracellular tracers and was therefore functionally segregated from the extracellular medium, although narrow connections between their membranes and the plasmalemma were sometimes visible by serial sectioning. Similar observations were made in intact cultured hippocampal neurons exposed to high K+ stimulation. Coated vesicle buds were generally in the same size range of synaptic vesicles and positive for the synaptic vesicle protein synaptotagmin. Based on these results, we suggest that endosome-like intermediates of nerve terminals originate by bulk uptake of the plasma membrane and that clathrin- and dynamin-mediated budding takes place in parallel from the plasmalemma and from these internal membranes. We propose a synaptic vesicle recycling model that involves a single vesicle budding step mediated by clathrin and dynamin.

Adaptor Protein Complex 2↗
PubMed1996

Histopathologic study of coexistent pathologic states in pancreatic fibrosis in patients with chronic alcohol abuse: two distinct pathologic fibrosis entities with different mechanisms.

The distribution and clinicopathologic features of pancreatic fibrosis were studied histopathologically in 137 autopsy cases of chronic alcohol abuse. Fibrosis was observed in 90 of the cases and was classified as perilobular sclerosis (PS) and intralobular sclerosis (IS). Fibrosis of the PS type was irregular and sometimes patchy and extended into the intralobular area in advanced cases. In some advanced cases, complete replacement of the pancreatic tissue by extensive fibrosis was seen. Fibrosis of the IS type was uniformly distributed. The tissues in some cases showed prominently periacinar fibrosis. In these cases, the pancreatic parenchyma had not been completely replaced by extensive fibrosis. Clinicopathologic comparisons revealed the following results: accompanying liver cirrhosis was greater in the IS than in the PS of fibrosis. However, a higher frequency of protein plugs, pancreatic stones, extensive fibrosis replacement, peripancreatic fibrosis, splenic vein involvement, choledochus involvement, pseudocyst, and ductal hyperplasia was found in the PS type compared to the IS type. In conclusion, the findings on the perilobular and intralobular distribution of fibrosis and differences in various components or accompanying diseases in pancreatic fibrosis suggest that this entity shows two distinct pathologic patterns with differing mechanisms.

Adult↗
PubMed1996

p53 and Ki-67 immunoreactivity and nuclear morphometry of 'carcinoma-in-adenoma' and adenoma of the gall-bladder.

Twenty intramucosal tumors of "carcinoma-in-adenoma' and 43 adenomas (39 pyloric gland type, 4 intestinal type) of the gall-bladder were studied to establish more precise histological criteria of carcinoma or adenoma in cases of "carcinoma in pyloric gland type adenoma', to compare carcinoma in adenoma with pure, that is, without adenomatous components, carcinoma, and to confirm the benign nature of spindle cell foci in the adenomas. Ki-67 and p53 immunostaining and nuclear morphometry were used. Eight pure intramucosal cancers were used as controls. The formalin-fixed, paraffin-embedded sections were stained with p53 and Ki-67 antibodies. Spindle cell foci were observed only in the adenoma area of the pyloric gland type, with a frequency of 23% in 39 adenomas, and of 45% in 20 tumors of carcinoma-in-adenoma. Ki-67 staining was negative in 129 of 130 spindle cell foci examined, regardless of their size, and positive in only one focus (550 microns in size, Ki-67 index 0.2%). All of the spindle cell foci were negative for p53 stain. The Ki-67 positive index was 36.6 +/- 5.6% in the 8 pure carcinomas, and 12.5 +/- 1.9% in the cancer areas of 16 tumors with carcinoma-in-adenoma, while it was 7.9 +/- 1.7% in the adenoma areas of 16 tumors with carcinoma-in-adenoma and 4.9 +/- 0.5% in the 32 pure pyloric gland adenomas. The p53-protein over-expression was found in seven of eight pure intramucosal cancers, and in one of 16 cancer components of carcinoma-in-adenoma. However, it was not found in any of 16 adenoma components of carcinoma-in-adenoma, and 35 adenomas. Cells of the cancer tissue of carcinoma-in-adenoma showed a significantly larger nuclear area and a larger nuclear minor axis than those of the pyloric gland type adenomas, as well as other architectural and cytologic abnormalities differing from the features of adenomas. These results suggest that clustered spindle cells do not indicate a malignant transformation of adenoma cells and that carcinomas in carcinoma-in-adenoma are different from pyloric gland type adenomas in terms of morphology and proliferative activity. Moreover, the results of the present study indicate that carcinomas in carcinoma-in-adenoma are lower in malignancy than pure carcinomas, and that their genetic abnormality may differ from that of pure carcinomas.

Adenocarcinoma↗
PubMed1996

p53 and Ki-67 immunoreactivity and nuclear morphometry of 'carcinoma-in-adenoma' and adenoma of the gall-bladder.

Twenty intramucosal tumors of 'carcinoma-in-adenoma' and 43 adenomas (39 pyloric gland type, 4 intestinal type) of the gall-bladder were studied to establish more precise histological criteria of carcinoma or adenoma in cases of 'carcinoma in pyloric gland type adenoma', to compare carcinoma in adenoma with pure, that is, without adenomatous components, carcinoma, and to confirm the benign nature of spindle cell foci in the adenomas. Ki-67 and p53 immunostaining and nuclear morphometry were used. Eight pure intramucosal cancers were used as controls. The formalin-fixed, paraffin-embedded sections were stained with p53 and Ki-67 antibodies. Spindle cell foci were observed only in the adenoma area of the pyloric gland type, with a frequency of 23% in 39 adenomas, and of 45% in 20 tumors of carcinoma-in-adenoma. Ki-67 staining was negative in 129 of 130 spindle cell foci examined, regardless of their size, and positive in only one focus (550 microns in size, Ki-67 index 0.2%). All of the spindle cell foci were negative for p53 stain. The Ki-67 positive index was 36.6 +/- 5.6% in the 8 pure carcinomas, and 12.5 +/- 1.9% in the carcinoma area of 16 tumors with carcinoma-in-adenoma, while it was 7.9 +/- 1.7% and in the adenoma areas of 16 tumors with carcinoma-in-adenoma and 4.9 +/- 0.5% in the 32 pure pyloric gland adenomas. The p53-protein overexpression was found in seven of eight pure intramucosal cancers, and in one of 16 cancer components of carcinoma-in-adenoma. However, it was not found in any of 16 adenoma components of carcinoma-in-adenoma, and 35 adenomas. Cells of the cancer tissue of carcinoma-in-adenoma showed a significantly larger nuclear area and a larger nuclear minor axis than those of the pyloric gland type adenomas, as well as other architectural and cytologic abnormalities differing from the features of adenomas. These results suggest that clustered spindle cells do not indicate a malignant transformation of adenoma cells and that carcinomas in carcinoma-in-adenoma are different from pyloric gland type adenomas in terms of morphology and proliferative activity. Moreover, the results of the present study indicate that carcinomas. In carcinoma-in-adenoma are lower in malignancy than pure carcinomas, and that their genetic abnormality may differ from that of pure carcinomas.

Adenocarcinoma↗
PubMed1996

Survey of risk factors for expulsive choroidal hemorrhage: case reports. Substantiation of the risk factors and their incidence.

To evaluate the interaction of risk factors for expulsive hemorrhage, we evaluated 3 cases of expulsive hemorrhage and 2 of acute intraoperative choroidal effusion that occurred at the University of Tsukuba Hospital over a 16-year-period and investigated the common findings in these 5 cases. We also selected randomly 500 cataractous cases operated at the University of Tsukuba Hospital and investigated the probabilities of risk factors for expulsive hemorrhage in these control cases. The 5 affected patients exhibited more than four risk factors of expulsive hemorrhage. The probability of more than four risk factors being involved in any cataract operation was approximately 0.33% (0.17 approximately 0.67%; 95% confidence interval). The incidence of expulsive hemorrhage at our hospital was approximately 0.2%, which is similar to the probability rate. Thus, we concluded that the development of expulsive hemorrhage was proved to be related to an accumulation of the risk factors for expulsive hemorrhage.

Aged↗
PubMed1996

Piccolo, a novel 420 kDa protein associated with the presynaptic cytomatrix.

In this study, we describe a novel 420 kDa protein, called Piccolo, found at a wide variety of adult rat brain synapses. High protein levels in the cerebellum, the olfactory bulb and the hippocampus were frequently observed to be associated with asymmetric type 1 synapses. Piccolo is selectively enriched in presynaptic terminals, but is not a component of synaptic vesicles (SVs). Immunogold electron microscopy revealed that Piccolo localizes to the amorphous material among SVs at the presynaptic plasma membrane. Biochemical studies showed that it is very tightly bound to this structure. Thus, we speculate that Piccolo is a structural component of the presynaptic cytomatrix which anchors SVs to the presynaptic plasmalemma.

Animals↗
PubMed1996

[A specular microscopic study on irradiated rabbit corneas--report I. Early injury of the cornea].

To evaluate the effects of ionizing radiation on the cornea, one eye of healthy mature albino rabbits (n = 5) was irradiated with 20 Gy of X-rays (250 kV, 12 mA). Corneal observation was done with a slit-lamp biomicroscope and a specular microscope before irradiation, and follow-up observations were done at one week and 4 weeks after irradiation. Specular microscopic photographs were analyzed with a digitizer. Although slit-lamp biomicroscopy did not show any abnormal findings in the cornea, specular microscopy revealed a significant enlargement of the mean cell area of the superficial corneal epithelium at 4 weeks after irradiation. There were no significant changes in the corneal endothelium during this study. These results strongly suggest that high doses of X-ray irradiation could induce corneal epithelial injury at the early phase, even when by slit-lamp biomicroscopy shows no abnormal findings in the cornea.

Animals↗
PubMed1995

Asymmetric retraction of growth cone filopodia following focal inactivation of calcineurin.

The neuronal growth cone is thought to be the site of decision making in nerve growth and guidance. One likely mechanism of how the growth cone translates various extracellular cues into directed motility involves rises in intracellular calcium. A variety of physiological cues, such as adhesion molecules and neurotransmitters, increases intracellular calcium, and artificial manipulations of growth cone calcium levels affect growth cone morphology and neurite outgrowth. The molecular events downstream of calcium fluxes are incompletely understood. Here we show that calcineurin, a protein phosphatase enriched in growth cones that is dependent on calcium ions and calmodulin, functions in neurite outgrowth and directed filopodial motility in cultured chick dorsal root ganglia neurons. Cyclosporin A and FK506, inhibitors of calcineurin, delayed neuritogenesis and inhibited neurite extension. Chromophore-assisted laser inactivation of calcineurin in regions of growth cones causes localized filopodial and lamellipodial retraction and influences the direction of subsequent outgrowth. We suggest that a spatial distribution of calcineurin activity within the growth cone can regulate motility and direct outgrowth.

Animals↗
PubMed1995

Tubular membrane invaginations coated by dynamin rings are induced by GTP-gamma S in nerve terminals.

The mechanisms through which synaptic vesicle membranes are reinternalized after exocytosis remain a matter of debate. Because several vesicular transport steps require GTP hydrolysis, GTP-gamma S may help identify intermediates in synaptic vesicle recycling. In GTP-gamma S-treated nerve terminals, we observed tubular invaginations of the plasmalemma that were often, but not always, capped by a clathrin-coated bud. Strikingly, the walls of these tubules were decorated by transverse electron-dense rings that were morphologically similar to structures formed by dynamin around tubular templates. Dynamin is a GTPase implicated in synaptic vesicle endocytosis and here we show that the walls of these membranous tubules, but not their distal ends, were positive for dynamin immunoreactivity. These findings demonstrate that dynamin and clathrin act at different sites in the formation of endocytic vesicles. They strongly support a role for dynamin in the fission reaction and suggest that stabilization of the GTP-bound conformation of dynamin leads to tubule formation by progressive elongation of the vesicle stalk.

Animals↗
PubMed1995

Increase in choroidal blood flow in rabbits with endothelin-1 induced transient complete obstruction of retinal vessels.

BACKGROUND: In a previous paper, we reported that retinal blood flow (RBF) ceased immediately after injection of 1 nmol endothelin-1 (ET-1) and no recovery of RBF was detected for at least 50 min. In this study, we confirmed the same duration of RBF cessation and measured choroidal blood flow (CBF) for 180 min. METHODS: We measured CBF in a rabbit model of transient complete obstruction of retinal vessels induced by intravitreal injection of a high dose of ET-1, using the hydrogen clearance method. We also investigated the effects of intravitreal injection of ET-1 on intraocular pressure (IOP), blood pressure, pulse rate and blood gases. RESULTS: CBF was significantly greater in the ET-1-injected eyes than in the control eyes 40-130 min after injection of ET-1 (P < 0.05). The maximal CBF ratio in the ET-1-injected eyes was 128 +/- 7.4% at 40 min. CBF decreased to the pre-injection level at 140 min after the injection of ET-1. There was no significant change in blood pressure, pulse rate and blood gases throughout this experiment, and there was no significant difference in IOP between ET-1-injected eyes and control eyes. CONCLUSION: It seems likely that the increase in CBF resulted from some local mechanisms of control that compensated for the decrease in RBF induced by intravitreal injection of ET-1. This model may be useful for investigation of the regulatory system of intraocular circulation, including endothelin receptors.

Animals↗
PubMed1995

Genotyping of isolates of Taylorella equigenitalis from thoroughbred brood mares in Japan.

Profiles of the genomic DNA of 104 strains of T. equigenitalis isolated from brood mares with contagious equine metritis in Hokkaido during the breeding seasons from 1980 to 1993, as well as those of five strains (SS28, EQ56, EQ59, EQ70 and HH139) previously isolated in Japan were examined after restriction digestion and crossed-field gel electrophoresis. These profiles were essentially identical to each other and the various isolates and strains appeared to have a common genotype, designated 'genotype J', with respect to two restriction enzymes, ApaI and NotI. These results suggest a common source for all these isolates obtained over the course of more than 10 years in Japan.

Animals↗
PubMed1995

The function of dynamin in endocytosis.

Temperature-sensitive shibire mutants of Drosophila melanogaster become rapidly paralyzed upon a shift to the restrictive temperature, which is due to a block in synaptic vesicle endocytosis. The shibire gene encodes the GTPase dynamin. Recent studies have shown that dynamin forms rings at the neck of invaginated clathrin-coated pits, and have suggested that a conformational change in the ring, which correlates with GTP hydrolysis, plays an essential role in vesicle fission.

Animals↗
PubMed1995

rbSec1A and B colocalize with syntaxin 1 and SNAP-25 throughout the axon, but are not in a stable complex with syntaxin.

rbSec1 is a mammalian neuronal protein homologous to the yeast SEC1 gene product which is required for exocytosis. Mutations in Sec1 homologues in the nervous systems of C. elegans and D. melanogaster lead to defective neurotransmitter secretion. Biochemical studies have shown that recombinant rbSec1 binds syntaxin 1 but not SNAP-25 or synaptobrevin/VAMP, the two proteins which together with syntaxin 1 form the synaptic SNARE complex. In this study we have examined the subcellular localization of rbSec1 and the degree of interaction between rbSec1 and syntaxin 1 in situ. rbSec1, which we show here to be represented by two alternatively spliced isoforms, rbSec1A and B, has a widespread distribution in the axon and is not restricted to the nerve terminal. This distribution parallels the localization of syntaxin 1 and SNAP-25 along the entire axonal plasmalemma. rbSec1 is found in a soluble and a membrane-associated form. Although a pool of rbSec1 is present on the plasmalemma, the majority of membrane-bound rbSec1 is not associated with syntaxin 1. We also show that rbSec1 is not part of the synaptic SNARE complex or of the syntaxin 1/SNAP-25 complex we show to be present in non-synaptic regions of the axon. Thus, in spite of biochemical studies demonstrating the high affinity interaction of rbSec1 and syntaxin 1, our results indicate that rbSec1 and syntaxin 1 are not stably associated. They also suggest that the function of rbSec1, syntaxin 1, and SNAP-25 is not restricted to synaptic vesicle exocytosis at the synapse.

Alternative Splicing↗
PubMed1995

Cellubrevin and synaptobrevins: similar subcellular localization and biochemical properties in PC12 cells.

There is strong evidence to indicate that proteins of the synaptobrevin family play a key role in exocytosis. Synaptobrevin 1 and 2 are expressed at high concentration in brain where they are localized on synaptic vesicles. Cellubrevin, a very similar protein, has a widespread tissue distribution and in fibroblasts is localized on endosome-derived, transferin receptor-positive vesicles. Since brain cellubrevin is not detectable in synaptic vesicles, we investigated whether cellubrevin and the synaptobrevins are differentially targeted when co-expressed in the same cell. We report that in the nervous system cellubrevin is expressed at significant levels only by glia and vascular cells. However, cellubrevin is coexpressed with the two synaptobrevins in PC12 cells, a neuroendocrine cell line which contains synaptic vesicle-like microvesicles. In PC12 cells, cellubrevin has a distribution very similar to that of synaptobrevin 1 and 2. The three proteins are targeted to neurites which exclude the transferrin receptor and are enriched in synaptic-like microvesicles and dense-core granules. They are recovered in the synaptic-like microvesicle peak of glycerol velocity gradients, have a similar distribution in isopycnic fractionation and are coprecipitated by anti-synaptobrevin 2 immunobeads. Finally, cellubrevin, like the synaptobrevins, interact with the neuronal t-SNAREs syntaxin 1 and SNAP-25. These results suggest that cellubrevin and the synaptobrevins have similar function and do not play a specialized role in constitutive and regulated exocytosis, respectively.

Amino Acid Sequence↗