Biomedical subjects

A Hirata

Publications and source records attributed to A Hirata.

At least 55 records · Page 3Linked to original sources

PubMed2001

A mutation in SPC42, which encodes a component of the spindle pole body, results in production of two-spored asci in Saccharomyces cerevisiae.

In the yeast Saccharomyces cerevisiae, SPC42 is an essential gene, which encodes one of the major components of the spindle pole body (SPB). We report on a mutation in the SPC42 gene (spc42-102) that results in a sporulation-specific defect. Mitotic growth of haploid and diploid spc42-102 strains is normal and both exhibit the same growth rates as the isogenic wild-type strains. Many diploid spc42-102/spc42-102 cells undergo normal meiotic nuclear divisions, producing four haploid nuclei. However, a significant fraction of meiotic spc42-102/spc42-102 cells contain two immature SPBs and aberrant nuclei that are not surrounded by a prospore membrane. Some 40% of the resultant asci contain only two spores, while wild-type diploid cells almost always produce four-spored asci. Segregation of auxotrophic markers that are tightly linked to the centromere reveals that two-spore asci formed from spc42-102/spc42-102 diploid cells exclusively contain nonsister haploid spores. Western analysis and measurements of the fluorescent signal from an Spc42p-GFP (green fluorescent protein) fusion reveal that the mutant strain fails to accumulate Spc42p at meiosis. Thus, our results suggest that insufficiency of Spc42p during meiosis results in a pair of immature nonsister SPBs that are not enclosed by prospore membrane.

Blotting, Western↗

PubMed2001

Fundus changes corresponding to visual field defects after vitrectomy for macular hole.

OBJECTIVE: Visual field defects are one of the complications of macular hole surgery, and mechanical retinal damage induced by infusion air is a proposed causative factor of this complication. In this study, we examined the fundus to see whether the changes observed corresponded to postoperative visual field defects. DESIGN: Observational case series. PARTICIPANTS: Seventeen eyes of 17 patients who had postoperative visual field defects after vitrectomy for idiopathic macular hole were examined. METHODS: The fundus was examined by ophthalmoscopy and by fluorescein and indocyanine green angiography. MAIN OUTCOME MEASURES: Fundus changes corresponding to postoperative visual field defects. RESULTS: In eight eyes, detectable fundus changes were observed, including regional mottling and degeneration of the retinal pigment epithelium, filling delay of the choroidal circulation, subretinal fibrosis, and epiretinal membrane formation. These findings corresponded exactly to the visual field defects observed. Although the visual field defects had been detected shortly after surgery, fundus changes were detected, on average, more than 8 months after surgery. CONCLUSIONS: Fundus changes become apparent after surgery, and they are progressive. Therefore, it is important to examine eyes with visual field defects for a follow-up period of several years.

Aged↗

PubMed2001

Soft particle analysis of bacterial cells and its interpretation of cell adhesion behaviors in terms of DLVO theory.

The electrokinetic properties of two nitrifying strains, Nitrosomonas europaea and Nitrobacter winogradskyi, and three heterotrophic bacteria, Escherichia coli, Pseudomonas putida and Pseudomonas aeruginosa, were examined by electrophoretic mobility measurement and analyzed using the soft particle electrophoresis theory that is suitable for biological particles. The bacterial adhesion characteristics onto glass bead substratum were also evaluated by packed bed method. The mobility of the bacterial cells employed converged to a non-zero value as the ionic concentration increased, suggesting that the bacterial cells exhibited typical soft particle characteristics. Moreover, cell surface potentials based on the soft particle theory were lower than those estimated by the conventional Smoluchowski formula, i.e. zeta potential. Cell collision efficiencies onto glass beads (alpha(0)) were largely dependent on interfacial interaction, although almost electrically neutral P. aeruginosa did not follow that trend. From a comparison of alpha(0) with DLVO interaction energy maximum (V(max)), it was assumed that heterocoagulation between cell and substratum at primary minimum potential took place under V(max) of 24-34 kT based on soft particle analysis. On the other hand, V(max) predictions using the Smoluchowski theory gave 81-223 kT, which indicated the possibility of overestimating electrostatic repulsive forces by the conventional Smoluchowski theory. Thus, the application of this new electrophoresis theory to several kinds of bacterial cells has led to the revision of the interpretation of bacterial mobility data and provided a more detailed understanding of the bacterial adhesion phenomenon.

Journal Article↗

PubMed2001

Characterization of platelet-derived growth factor-induced p38 mitogen-activated protein kinase activation in vascular smooth muscle cells.

BACKGROUND: The mitogen-activated protein (MAP) kinase super-family plays a crucial role in cell growth and differentiation and even in programmed cell death in response to diverse extracellular stimuli. The platelet-derived growth factor (PDGF)-BB is well known to promote the proliferation of vascular smooth muscle cells (VSMC) via extracellular-regulated protein kinases (ERKs), leading to the development of cardiovascular diseases. However, it has not yet been clarified whether PDGFs that include other isoforms can activate the other parallel signal transduction pathways, c-jun NH2-terminal protein kinase (JNK) and p38 MAP kinase (p38), in VSMC. In this study, we investigated the effect of PDGFs on p38 activation in cultured rat VSMC. MATERIALS AND METHODS: After stimulation by PDGFs with SB-203580 or PD-98059, the cells were solubilized, and the expressions of MAP kinases, MAP kinase kinases (MKKs), phosphorylated DNA-binding proteins, and cyclooxigenases (COXs) were examined by immunoblot analysis. RESULTS: PDGFs activated p38 phosphorylation dose-dependently, and the phosphorylations were specifically inhibited by SB-203580 but not by PD-98059. PDGFs also activated the phosphorylation of MKK 3/MKK 6 but not that of either stress-activated protein kinase/ERK kinase or JNK. PDGFs affected the activation of a cyclic AMP response-element binding protein, which was inhibited by SB-203580. However, the activating transcription factor-2 was not activated by PDGFs. Interestingly, the stimulation of PDGFs for 72 h enhanced the level of COX-2, and these levels were decreased by SB-203580. CONCLUSION: These results have clarified that PDGFs activate the p38 cascade via an MKK 3/6 pathway, independently of the ERK cascade, and subsequently regulate the level of COX-2 in rat VSMC, providing that PDGFs influence the inflammatory process in the vascular wall.

Activating Transcription Factor 2↗

PubMed2001

Characterization of an inhibitory effect of pioglitazone on balloon-injured vascular smooth muscle cell growth.

This study investigates whether pioglitazone could suppress an atherogenic process such as balloon-injured carotid intimal thickening and the proliferation of vascular smooth muscle cells (VSMC). We first examined the effect of pioglitazone to determine whether it could suppress intimal thickening induced by balloon catheterization in Sprague-Dawley rats. After 14 days postcatheterization in the left common carotid artery, the neointimal layers were completely occupied by proliferated VSMC, and the area ratio of neointima to media treated with 10 mg/kg/d of pioglitazone was significantly decreased to 57%. Next, we evaluated the effect of pioglitazone on the proliferation of rat cultured VSMC. Piogliotazone dose-dependently decreased the values of DNA synthesis, total cellular protein content, phosphorylations of extracellular signal-regulated protein kinase 1/2 and mitogen-activated protein kinase kinase 1/2, and proliferative cell nuclear antigen in VSMC. Pioglitazone also inhibited the phosphorylation of Pyk2. We conclude that pioglitazone itself could be effective for suppressing the growth of VSMC and consequent carotid intimal thickening.

Animals↗

PubMed2001

Identification of sinus node dysfunction by use of P-wave signal-averaged electrocardiograms in paroxysmal atrial fibrillation: a prospective study.

BACKGROUND: In patients with paroxysmal atrial fibrillation (Paf), the identification of the coexistence of sinus node dysfunction (SND) has therapeutic implications. This study sought to prospectively determine whether SND in patients with Paf would be identified by use of atrial early potential (EP), low-amplitude potentials early in signal-averaged P wave. METHODS: The study population consisted of 149 patients with Paf. Signal-averaged electrocardiography was recorded with the P-wave-triggering technique. The root mean square voltage for the initial 30 MS and the duration of initial low-amplitude signals < 4 microV of signal-averaged P wave were measured in the vector magnitude. The criteria of EP were defined as "the root mean square voltage for the initial 30 MS < 3.0 microV and the duration of initial low-amplitude signals < 4 microV >22 MS." SND was diagnosed by use of the conventional 12-lead electrocardiography, 24-hour Holter monitoring, and bedside electrocardiographic monitoring. RESULTS: Thirty-eight of 149 patients with Paf had EP. Eighteen (47%) of 38 patients with Paf and EP had SND, whereas SND was found in only 5 (5%) of the other 111 patients with Paf without EP (P <.0001). EP gave a sensitivity of 78% and a specificity of 84% for the detection of SND in patients with Paf. CONCLUSION: EP would be useful for the identification of SND in patients with Paf.

Aged↗

PubMed2001

The Schizosaccharomyces pombe spo3+ gene is required for assembly of the forespore membrane and genetically interacts with psy1(+)-encoding syntaxin-like protein.

Formation of the forespore membrane, which becomes the plasma membrane of spores, is an intriguing step in the sporulation of the fission yeast Schizosaccharomyces pombe. Here we report two novel proteins that localize to the forespore membrane. spo3(+) encodes a potential membrane protein, which was expressed only during sporulation. Green fluorescent protein (GFP) fusion revealed that Spo3 localized to the forespore membrane. The spo3 disruptant was viable and executed meiotic nuclear divisions as efficiently as the wild type but did not form spores. One of the spo3 alleles, spo3-KC51, was dose-dependently suppressed by psy1(+), which encodes a protein similar to mammalian syntaxin-1A, a component of the plasma membrane docking/fusion complex. psy1(+) was essential for vegetative growth, and its transcription was enhanced during sporulation. As expected, Psy1 localized to the plasma membrane during vegetative growth. Interestingly, Psy1 on the plasma membrane disappeared immediately after first meiotic division and relocalized to the forespore membrane as the second division initiated. In the spo3 null mutant, the forespore membrane was initiated but failed to develop a normal morphology. Electron microscopy revealed that membrane vesicles were accumulated in the cytoplasm of immature spo3Delta asci. These results suggest that Spo3 is a key component of the forespore membrane and is essential for its assembly acting in collaboration with the syntaxin-like protein.

Amino Acid Sequence↗

PubMed2001

The Schizosaccharomyces pombe spo20(+) gene encoding a homologue of Saccharomyces cerevisiae Sec14 plays an important role in forespore membrane formation.

The Schizosaccharomyces pombe spo20-KC104 mutation was originally isolated in a screen for sporulation-deficient mutants, and the spo20-KC104 mutant exhibits temperature-sensitive growth. Herein, we report that S. pombe, spo20(+) is essential for fission yeast cell viability and is constitutively expressed throughout the life cycle. We also demonstrate that the spo20(+) gene product is structurally homologous to Saccharomyces cerevisiae Sec14, the major phosphatidylinositol transfer protein of budding yeast. This structural homology translates to a significant degree of functional relatedness because reciprocal complementation experiments demonstrate that each protein is able to fulfill the essential function of the other. Moreover, biochemical experiments show that, like Sec14, Spo20 is a phosphatidylinositol/phosphatidylcholine-transfer protein. That Spo20 is required for Golgi secretory function in vegetative cells is indicated by our demonstration that the spo20-KC104 mutant accumulates aberrant Golgi cisternae at restrictive temperatures. However, a second phenotype observed in Spo20-deficient fission yeast is arrest of cell division before completion of cell separation. Consistent with a direct role for Spo20 in controlling cell septation in vegetatively growing cells, localization experiments reveal that Spo20 preferentially localizes to the cell poles and to sites of septation of fission yeast cells. We also report that, when fission yeasts are challenged with nitrogen starvation, Spo20 translocates to the nucleus. This nuclear localization persists during conjugation and meiosis. On completion of meiosis, Spo20 translocates to forespore membranes, and it is the assembly of forespore membranes that is abnormal in spo20-KC104 cells. In such mutants, a considerable fraction of forming prespores fail to encapsulate the haploid nucleus. Our results indicate that Spo20 regulates the formation of specialized membrane structures in addition to its recognized role in regulating Golgi secretory function.

Amino Acid Sequence↗

PubMed2001

Efficient liquid-phase synthesis of short-length nucleotides via the phosphoramidite method using stoichiometric amounts of reactants.

The condensation of a nucleoside phosphoramidite and a nucleoside by the aid of a suitable promoter in stoichiometric use is achieved in acetonitrile in the presence of molecular sieves 3A or 4A to give a desired coupling product in an excellent yield. This strategy is particularly useful for the large-scale synthesis of short nucleotides.

Chemistry, Organic↗

PubMed2001

Renal vascular damage in Japanese patients with systemic sclerosis.

OBJECTIVE: To evaluate renal vascular damage in Japanese patients with systemic sclerosis (SSc) by colour-flow Doppler ultrasonography. METHODS: The pulsatility index (PI) was measured in renal interlobar and segmental arteries by colour-flow Doppler ultrasonography. RESULTS: PI values of interlobar arteries were increased in SSc patients (n=53) with normal renal function compared with healthy persons (n=16), systemic lupus erythematosus patients (n=12) and dermatomyositis patients (n=3). SSc patients with elevated PI levels had digital pitting scar, short sublingual frenulum, contracture of phalanges, pulmonary fibrosis, decreased per cent vital capacity, heart involvement, positivity for anti-topoisomerase I antibody, and elevated C-reactive protein more frequently than those with normal PI levels. CONCLUSION: Although renal crisis is rare in Japanese SSc patients, our study suggests that latent and subclinical renal damage exists in these patients.

Adult↗

PubMed2001

Dosimetry in models of child and adult for low-frequency electric field.

Induced electric field and current density in a child's body exposed to a 60-Hz electric field are calculated and compared with those for an adult's body. Because of the different proportions of the child body relative to those of the adult body, differences in the induced electric field and current density values in various organs are observed. These results are interpreted in terms of international guideline limits, and hypotheses regarding plausible interactions.

Adult↗

PubMed2001

Direct detection by in situ PCR of the amoA gene in biofilm resulting from a nitrogen removal process.

Ammonia oxidation is a rate-limiting step in the biological removal of nitrogen from wastewater. Analysis of microbial communities possessing the amoA gene, which is a small subunit of the gene encoding ammonia monooxygenase, is important for controlling nitrogen removal. In this study, the amoA gene present in Nitrosomonas europaea cells in a pure culture and biofilms in a nitrifying reactor was amplified by in situ PCR. In this procedure, fixed cells were permeabilized with lysozyme and subjected to seminested PCR with a digoxigenin-labeled primer. Then, the amplicon was detected with an alkaline phosphatase-labeled antidigoxigenin antibody and HNPP (2-hydroxy-3-naphthoic acid-2'-phenylanilide phosphate), which was combined with Fast Red TR, and with an Alexa Fluor 488-labeled antidigoxigenin antibody. The amoA gene in the biofilms was detected with an unavoidable nonspecific signal when the former method was used for detection. On the other hand, the amoA gene in the biofilms was detected without a nonspecific signal, and the cells possessing the amoA gene were clearly observed near the surface of the biofilm when Alexa Fluor 488-labeled antidigoxigenin antibody was used for detection. Although functional gene expression was not detected in this study, detection of cells in a biofilm based on their function was demonstrated.

Biofilms↗

PubMed2001

Prognostic significance of cardiac (123)I metaiodobenzylguanidine imaging for mortality and morbidity in patients with chronic heart failure: a prospective study.

OBJECTIVE: To determine whether cardiac iodine-123 metaiodobenzylguanidine ((123)I MIBG) imaging is useful in predicting the prognosis of patients with chronic heart failure. DESIGN: Cardiac (123)I MIBG imaging was done on entry to the study. The cardiac MIBG washout rate was calculated from anterior chest view images obtained 20 and 200 minutes after injection of the isotope. Study patients were divided into two groups with washout rates above and below 27% (the mean value + 2 SD obtained in 20 normal subjects), and were then followed up. SETTING: Tertiary referral centre. PATIENTS: 79 patients with chronic heart failure in whom the left ventricular ejection fraction was less than 40%. RESULTS: There were 37 patients in group 1 (washout rate of >/= 27%) and 42 in group 2 (< 27%). During a follow up period of between 1 and 52 months, eight patients died suddenly and five died of worsening heart failure in group 1, while none died in group 2; 13 patients in group 1 and four in group 2 were admitted to hospital for progressive heart failure. Kaplan-Meier analysis showed that group 1 had a significantly higher mortality and morbidity (p = 0.001 and p < 0.001, respectively) than group 2. CONCLUSIONS: Cardiac (123)I MIBG washout rate seems to be a good predictor of prognosis in patients with chronic heart failure.

3-Iodobenzylguanidine↗

PubMed2001

Loss of the mitochondrial Hsp70 functions causes aggregation of mitochondria in yeast cells.

Ssc1p, a member of the Hsp70 family in the mitochondrial matrix of budding yeast, mediates protein import into mitochondria and prevents irreversible aggregation of proteins in the mitochondrial matrix during folding/assembly or at elevated temperature. Here, we show that functional inactivation of the mitochondrial Hsp70 system causes aggregation of mitochondria. When temperature-sensitive mitochondrial Hsp70 mutant cells were incubated at restrictive temperature, a tubular network of mitochondria was collapsed to form aggregates. Inhibition of protein synthesis in the cytosol did not suppress the mitochondrial aggregation and functional impairment of Tim23, a subunit of mitochondrial protein translocator in the inner membrane, did not cause mitochondrial aggregation. Therefore defects of the Hsp70 function in protein import into mitochondria or resulting accumulation of precursor forms of mitochondrial proteins outside the mitochondria are not the causal reason for the aberrant mitochondrial morphology. By contrast, deletion of Mdj1p, a functional partner for mitochondrial Hsp70 in prevention of irreversible protein aggregation in the matrix, but not in protein import into mitochondria, caused aggregation of mitochondria, which was enhanced at elevated temperature (37 degrees C). The aggregation of mitochondria at 37 degrees C was reversed when the temperature was lowered to 23 degrees C unless protein synthesis was blocked. On the basis of these results, we propose that the mitochondrial matrix contains a protein that is responsible for the maintenance of mitochondrial morphology and requires mitochondrial Hsp70 for its function.

Actins↗

PubMed2001

Microbial community analysis in the denitrification process of saline-wastewater by denaturing gradient gel electrophoresis of PCR-amplified 16S rDNA and the cultivation method.

The metallurgic wastewater generated from the processes of recovering precious metals from industrial wastes contains high concentrations of nitrogen compounds and salts. Biological nitrogen removal from this wastewater was attempted using a circulating bioreactor system equipped with an anaerobic packed bed or an anaerobic fluidized bed. The denitrification capability of the system with the anaerobic packed bed was more stable than that of the system with the anaerobic fluidized bed. The NOx removal rate of the anaerobic packed bed was as high as 97%. Microbial community analysis by denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S ribosomal DNA (rDNA) fragments and the cultivation method revealed that the community diversity varied in accordance with wastewater composition such as the level of salinity and so on. Phylogenetic analysis suggested that the taxonomic affiliation of the dominant species in the anaerobic reactors was to the gamma-Proteobacteria including Halomonadaceae species. The PCR-DGGE method as a non-cultivation method was found to be a powerful tool for analysis of the microbial community, because the cultivation method could detect only a fraction of the microbial species present in these systems. The genetic diversity of the isolated bacteria belonging to the gamma-Proteobacteria which reduced both nitrate and nitrite in the anaerobic packed bed was higher than that of the bacteria in the anaerobic fluidized bed. This suggested that a genetically diverse microbial community stabilized the denitrifying performance in the anaerobic packed bed.

Journal Article↗

PubMed2001

Metabolic behavior of denitrifying phosphate-accumulating organisms under nitrate and nitrite electron acceptor conditions.

The effects of various types of electron acceptors on anoxic phosphorus uptake were investigated in detail to obtain a better insight into the metabolic behavior of denitrifying phosphate-accumulating organisms. Batch experimental tests under three different electron acceptor conditions, i.e., nitrate, nitrite and mixtures of nitrate and nitrite, were carried out using activated sludge cultivated in a sequencing batch reactor. The experimental results confirmed no inhibition of the utilization of nitrate or nitrite as an electron acceptor for anoxic phosphorus uptake. Anoxic phosphorus uptake occurred provided there was an electron acceptor present regardless of whether it was nitrate or nitrite. However, for nitrite a relatively small amount of anoxic phosphorus was taken up per nitrogen denitrified compared to nitrate. On the other hand, the amount of anoxic phosphorus taken up per nitrogen denitrified increased with an increase in the initial loading amount of electron acceptor in the case of nitrate, whereas it slightly decreased nitrite. Moreover, the amount of phosphorus taken up per nitrogen denitrified decreased with increasing mixed liquor suspended solid (MLSS) concentration in the case of nitrate, while it slightly increased for nitrite. From these results, it was confirmed that the activity of anoxic phosphorus uptake is strongly associated with the type and the initial loading amount of electron acceptor and the MLSS concentration under anoxic conditions.

Journal Article↗

PubMed2001

Platelet-derived growth factor BB-induced p38 mitogen-activated protein kinase activation causes cell growth, but not apoptosis, in vascular smooth muscle cells.

The aim of this experiment was to examine the regulation of p38 mitogen-activated protein (MAP) kinase by platelet-derived growth factor (PDGF)-BB and its biological effects on rat cultured vascular smooth muscle cells (VSMCs). VSMCs were obtained from aortae of male Wistar rats by the media explant technique. After being stimulated by PDGF-BB with or without the p38 MAP kinase-specific inhibitor, SB-203580, the cells were solubilized, and the levels of phosphorylated p38 MAP kinase were examined by immunoblot analysis. The amounts of DNA synthesis and content were measured by using [3H]-thymidine and Hoechst-33258 dye, respectively. The detection of apoptotic cells was evaluated by the TUNEL method. PDGF-BB could phosphorylate p38 MAP kinase dose-dependently, and the phosphorylation was specifically inhibited by SB-203580 in a dose-dependent manner. However, PDGF-BB did not affect the protein level of p38 MAP kinase. Both [3H]-thymidine incorporation and total cellular DNA content were increased by PDGF-BB, and these elevations were prevented by SB-203580. In contrast, PDGF-BB-stimulated VSMCs did not show apoptotic change in spite of the presence or absence of SB-203580. These results established that PDGF-BB activated p38 MAP kinase and subsequently regulated cell growth in VSMCs, providing a molecular mechanism by which p38 MAP kinase can cause the development of cardiovascular diseases, including atherosclerosis.

Animals↗

PubMed2001

Immunolocalization of keratan sulfate proteoglycan in rat calvaria.

We investigate, by the immunogold method, the localization of keratan sulfate (KS) proteoglycan in rat calvaria in order to clarify the detailed process of intramembranous ossification. KS was localized in bone nodules corresponding to calcified nodules, close to the saggital suture of calvaria. The immunoreactivity decreased in fully calcified regions distant from the suture. Electron microscopic observation revealed that KS was distributed in and around matrix vesicles, among collagen fibrils at the initial crystal deposition stage, and then concentrated in bone nodules. According to the progress of mineralization, KS tended to be localized in the peripheral region of the nodules. In addition, these nodules came in contact with collagen fibrils which also showed KS-positive reactivity. In cell organelles of osteoblasts, KS was detected in the Golgi apparatus. These findings suggest that osteoblasts in intramembranous ossification sites actively synthesize KS. KS in the calcified nodules, as well as other glycosaminoglycans in osteoid, may play an important role in additional and/or collagenous calcification by trapping calcium ions through its negative charge.

Animals↗