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Biomedical subjects

Y Inoue

Publications and source records attributed to Y Inoue.

At least 595 records · Page 33Linked to original sources
PubMed1998

Rapid and discrete isolation of oxygen-evolving His-tagged photosystem II core complex from Chlamydomonas reinhardtii by Ni2+ affinity column chromatography.

We have developed a simple and rapid procedure to isolate an oxygen-evolving photosystem II (PS II) core complex from Chlamydomonas reinhardtii. A His-tag made of six consecutive histidine residues was genetically attached at the carboxy terminus of D2 protein to create a metal binding site on the PS II supramolecular complex. The recombinant cells producing the His-tagged variant of D2 protein grew photoautotrophically as well as the wild-type cells. Characterization of the oxygen evolution and the thermoluminescence properties revealed that the His-tagging did not affect the functional integrity of the PS II reaction center. A PS II core complex was isolated from the detergent-solubilized thylakoids of the recombinant cells in 4 h by a single one-step Ni2+ affinity column chromatography. This preparation consists of D1, D2, CP43, CP47, 33 kDa, and a few low molecular weight proteins, and retains a high rate of oxygen-evolving activity (= 1000 micromol/mg Chl/h).

Animals↗
PubMed1998

Ubiquitous presence of cellular proteins that specifically bind to the 3' terminal region of hepatitis C virus.

The 3' terminal region (3'-X tail) of hepatitis C virus (HCV) genomic RNA forms a stable stem-loop structure. The 3'-X tail consists of 98 nucleotides (nt) that are highly conserved among the HCV strains and supposed to function as a cis-acting region for replication of negative strand RNA and/or viral encapsidation. In the present study, by UV cross-linking assay we found two kinds of cellular proteins of approximately 87 and 130 kDa, which specifically bind to the full-length 3'-X tail (nt 1 to 98), but not the 3'- or 5'-truncated 3'-X tail, consisting of nt 1 to 50 or nt 51 to 98, respectively. These proteins were detected in human cell lines such as hepatic tumor cell lines and a T-lymphocyte cell line and also in a human embryonic lung fibroblast cell strain. In addition, human hepatocellular carcinoma tissues expressed these proteins regardless of infection or uninfection of HCV. Furthermore, these proteins were also detected in normal human tissues derived from the lung, heart, kidney, stomach, intestine, and colon. Thus, these cellular proteins, which are ubiquitously present in human tissues, might be involved in viral replication and/or encapsidation.

Binding Sites↗
PubMed1998

Importance of glucose-6-phosphate dehydrogenase in the adaptive response to hydrogen peroxide in Saccharomyces cerevisiae.

Glucose-6-phosphate dehydrogenase (G6PDH)-deficient cells of Saccharomyces cerevisiae showed increased susceptibility and were unable to induce adaptation to oxidative stress. Historically, mainly in human erythrocytes, it has been suggested and accepted that decreased cellular GSH, due to loss of the NADPH-dependent activity of glutathione reductase (GR), is responsible for the increased sensitivity to oxidative stress in G6PDH-deficient cells. In the present study we investigated whether the increased susceptibility and the inability to induce adaptation to H2O2 stress of G6PDH-deficient yeast is caused by incompleteness of glutathione recycling. We constructed G6PDH- and GR-deficient mutants and analysed their adaptive response to H2O2 stress. Although G6PDH-deficient cells contained comparable amounts of GSH and GR activity to wild-type cells, GSSG was not reduced efficiently, and intracellular GSSG levels and the ratio of GSSG to total glutathione (GSSG/tGSH) were higher in G6PDH-deficient cells than in wild-type. On the other hand, GR-deficient cells showed a susceptibility identical with that of wild-type cells and induced adaptation to H2O2 stress, even though the GSSG/tGSH ratio in GR-deficient cells was higher than in G6PDH-deficient cells. These results indicate that incompleteness of glutathione recycling alone is not sufficient to account for the increased sensitivity and inability to induce adaptation to H2O2 stress of G6PDH-deficient yeast cells. In S. cerevisiae, G6PDH appears to play other important roles in the adaptive response to H2O2 stress besides supplying NADPH to the GR reaction.

Adaptation, Physiological↗
PubMed1998

Molecular identification of glutathione synthetase (GSH2) gene from Saccharomyces cerevisiae.

The hypothetical protein YOL049w on the chromosome XV was identified to be the structural gene for glutathione synthetase (GSH2) of Saccharomyces cerevisiae. Translational initiation site was identified by making the GSH2-lacZ fusion. The GSH2 gene contained an open reading frame (1473 bp) with 491 amino acids, and molecular weight of the GSH2 gene product was calculated to be 55,812. Glutathione synthetase activity in transformant carrying the GSH2 gene with multicopy plasmid increased approximately 4-fold. The GSH2 gene was not essential for growth of yeast cell, and glutathione was not detected from the gsh2 disrupter.

Amino Acid Sequence↗
PubMed1998

Functional characterization of Synechocystis sp. PCC 6803 delta psbU and delta psbV mutants reveals important roles of cytochrome c-550 in cyanobacterial oxygen evolution.

The functions of cytochrome c-550 and a 12 kDa protein in cyanobacterial oxygen evolution were studied with directed deletion mutants delta psbV and delta psbU of Synechocystis sp. PCC 6803, and the following results were obtained. (1) In contrast to the delta psbU mutant which is capable of autotrophic growth in the absence of Ca2+ or Cl- at a reduced rate, the delta psbV mutant lacking cytochrome c-550 could not grow at all without Ca2+ or Cl-. (2) The delta psbV mutant had a significantly reduced thermoluminescence emission intensity and flash oxygen yield, whereas the delta psbU mutant showed slight decreases in thermoluminescence intensity and flash oxygen yield, indicating corresponding decreases in the concentrations of O2-evolving centers in these mutants. (3) The delta psbV and delta psbU mutants exhibited elevated peak temperature for the thermoluminescence B- and Q-bands indicative of more stable S2 states. (4) The rise time of the O2 signal during the S3-[S4]-S0 transition was increased slightly in the delta psbV mutant but not in the delta psbU mutant. (5) The oxygen evolution was inactivated in the dark rapidly in the delta psbV mutant with a half-time of 28 min, but this did not happen in the delta psbU mutant. (6) Photoactivation of the oxygen-evolving complex after removal of the manganese cluster by hydroxylamine showed a higher quantum yield in the delta psbV mutant than in the delta psbU mutant or wild type. Taken together, these results indicated that cytochrome c-550 plays a substantial role in maintaining the stability and function of the manganese cluster in algal photosystem II, whereas the 12 kDa protein plays primarily a regulatory role in maintaining normal S-state transitions. These functional features of cytochrome c-550 and the 12 kDa protein were compared with those of the 23 and 17 kDa proteins in higher plant photosystem II and of the 33 kDa protein in both algal and plant photosystem II.

Bacterial Proteins↗
PubMed1998

Identification of oligo-N-glycolylneuraminic acid residues in mammal-derived glycoproteins by a newly developed immunochemical reagent and biochemical methods.

The occurrence of the alpha2-->8-linked oligomeric form of N-glycolylneuraminic acid (oligo-Neu5Gc) residues in mammalian glycoproteins was unequivocally demonstrated using a newly developed anti-oligo/poly-Neu5Gc monoclonal antibody as well as by chemical and biochemical methods. First, the antibody, designated mAb.2-4B, which specifically recognized oligo/poly-Neu5Gc with a degree of polymerization of >2, was developed by establishing a hybridoma cell line from P3U1 myeloma cells fused with splenocytes from an MRL autoimmune mouse immunized with dipalmitoylphosphatidylethanolamine-conjugated oligo/poly-Neu5Gc. Second, oligo-Neu5Gc was shown to occur in glycoproteins derived from pig spleen by Western blot analysis using mAb.2-4B, which was also confirmed by fluorometric high performance liquid chromatographic analysis of the product of periodate oxidation/reduction/acid hydrolysis of the purified glycopeptide fractions and by TLC and 600-MHz 1H NMR spectroscopic analysis of their mild acid hydrolysates. Finally, the ubiquitous occurrence of oligo-Neu5Gc chains as glycoproteinaceous components in Wistar rat tissue was immunochemically indicated. This is the first example demonstrating the diversity in oligo/poly-Sia structure in mammalian glycoproteins, where only poly-N-acetylneuraminic acid is known to occur. Such diversity in oligo/poly-Sia structure also implicates a diverged array of biological functions of this glycan unit in glycoproteins.

Animals↗
PubMed1998

Expression of the glyoxalase I gene of Saccharomyces cerevisiae is regulated by high osmolarity glycerol mitogen-activated protein kinase pathway in osmotic stress response.

Methylglyoxal is a cytotoxic metabolite derived from dihydroxyacetone phosphate, an intermediate of glycolysis. Detoxification of methylglyoxal is performed by glyoxalase I. Expression of the structural gene of glyoxalase I (GLO1) of Saccharomyces cerevisiae under several stress conditions was investigated using the GLO1-lacZ fusion gene, and expression of the GLO1 gene was found to be specifically induced by osmotic stress. The Hog1p is one of the mitogen-activated protein kinases (MAPKs) in S. cerevisiae, and both Msn2p and Msn4p are the transcriptional regulators that are thought to be under the control of Hog1p-MAPK. Expression of the GLO1 gene under osmotic stress was completely repressed in hog1Delta disruptant and was repressed approximately 80 and 50% in msn2Delta and msn4Delta disruptants, respectively. A double mutant of the MSN2 and MSN4 gene was unable to induce expression of the GLO1 gene under highly osmotic conditions. Glucose consumption increased approximately 30% during the adaptive period in osmotic stress in the wild type strain. On the contrary, it was reduced by 15% in the hog1Delta mutant. When the yeast cell is exposed to highly osmotic conditions, glycerol is synthesized as a compatible solute. Glycerol is synthesized from glucose, and a rate-limiting enzyme in glycerol biosynthesis is glycerol-3-phosphate dehydrogenase (GPD1 gene product), which catalyzes reduction of dihydroxyacetone phosphate to glycerol 3-phosphate. Expression of the GPD1 gene is also under the control of Hog1p-MAPK. Methylglyoxal is also synthesized from dihydroxyacetone phosphate; therefore, induction of the GLO1 gene expression by osmotic stress was thought to scavenge methylglyoxal, which increased during glycerol production for adaptation to osmotic stress.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1998

Cyclin E overexpression responsible for growth of human hepatic tumors with p21WAF1/CIP1/SDI1.

We examined a relationship between p21WAF1/CIP1/SDI1 and cell-cycle-related proteins in 12 human liver tumor cell lines (JHH-1, -2, -4, -5, -6, -7; HLE; HuH-7; Hep3B; PLC/PRF/5; HuH-6; HepG2). Seven (JHH-1, -2, -5, -6, -7; Hep3B; HepG2) out of eight cell lines having p21WAF1/CIP1/SDI1 protein overexpressed cyclin E protein, although one of them (JHH-5) overexpressed a reduced size of cyclin E. The rest (HuH-6) of the 8 cell lines with p21WAF1/CIP1/SDI1 showed a decreased expression of cyclin E. Four cell lines (JHH-4; HLE; HuH-7; PLC/PRF/5) deficient of p21WAF1/CIP1/SDI1 protein did not overexpress cyclin E protein. As to expression of the other cell-cycle-related proteins, cyclin A, cyclin D1, CDK2 or CDK4, no significant difference was detected among the 12 cell lines. These findings indicate that the human liver tumor cell lines which have the p21WAF1/CIP1/SDI1-inducible barriers of the cell cycle progression can go through the G1/S checkpoint by overexpressing cyclin E.

Blotting, Northern↗
PubMed1998

Three-dimensional electron microscopy of the photosystem II core complex.

A three-dimensional image of the spinach photosystem II core complex composed of CP47, D1, D2, cytochrome b-559, and psbI gene product was reconstructed at 20-A resolution from the two-dimensional crystals negatively stained with phosphotungstate. Confirming the previous proposal, the crystal had a p22121 symmetry. One PSII core complex was measured to be 80 x 80 A in the membrane plane and 88 A normal to it. The mass distribution was asymmetric about the lipid bilayer, consistent with predictions from the amino acid sequences. The lumenal mass consisted of three domains forming a characteristic triangular platform with another domain on top of it. Three stromal domains were smaller and linearly arranged. Due to strong stain exclusion in the hydrophobic core part of the lipid bilayer, the transmembrane region appeared to be imaged with a reversed contrast. Inverting the contrast resulted in a reasonable density distribution for that part. Thus, though the information on the transmembrane region is limited, the domain structure of the PSII core complex was revealed and allowed us to propose a model for the arrangement of subunits in the PSII core complex.

Crystallization↗
PubMed1998

Hepatic falciform ligament artery: angiographic anatomy and clinical importance.

The hepatic falciform ligament artery (HFLA) was evaluated by angiography and also by dissections. Based on the findings, the mechanism of the post-chemoembolization skin rash was studied. A total of 340 liver cirrhosis patients who underwent hepatic artery chemoembolization for hepatocellular carcinoma were reviewed in terms of the angiographic incidence of the HFLA, variations in its origin, and the incidence of skin rash. The HFLA was demonstrated in 26 (7.6%) of the 340 patients on angiography. Two HFLAs were observed in one patient. The origin was the middle hepatic artery (A4) in 16 cases, the superior branch of the middle hepatic artery in three, the inferior branch of the middle hepatic artery in two, the inferior branch of the left hepatic artery (A3) in three, and the confluence of A3 and A4 in three cases. There were no patients who developed post-chemoembolization skin rash. Two cadavers were dissected to investigate the anastomosis between the HFLA and the subcutaneous artery. Two different anastomoses were found: (1) direct and (2) via the ensiform branch of the internal thoracic artery. These were located at the lower and upper part of the falciform ligament, respectively. The distribution of a chemotherapeutic agent through these anastomoses is the likely cause of post-chemoembolization skin rash. If prophylactic embolization of the proximal portion of the HFLA using a metallic coil is performed, the skin rash will be prevented.

Adult↗
PubMed1998

[The role of bidirectional cavopulmonary shunt on selection of Fontan patients].

There are no objection against that pulmonary vascular resistance index (PVRI) is one of the most important factors for completion of successful application of Fontan-type operation. However, calculated PVRI in single ventricle physiology in often unreliable because of difficulty in accurate measurement of pulmonary blood flow, especially in patients with decreased pulmonary blood flow. Although the role of bidirectional cavopulmonary shunt (BCPS) in such patients has been increasingly recognized, the impact of BCPS on PVRI has not been fully understood. Between November 1993 and November 1996, 24 patients, aged between 0.54 and 22.2 years, with a wide variety of cardiac malformations underwent BCPS, and were followed up for the mean of 15.1 months. There were four hospital deaths (16.7%) and three deaths in follow-up (12.5%). Serial catheterization revealed that significant increase in mean arterial oxygen saturation from 75.8% to 83.9% (p = 0.005), and decrease in mean Nakata's index from 433 to 311 (p < 0.0001). PVRI calculated by using formulas derived from Ohm's law before BCPS (Pulmonary flow was derived from Fick formula) was highly (greater than 10 u.m2) or moderately (between 4 and 10 u.m2) elevated in 6 and 7 patients, respectively. However, PVRI in these patients was normal after BCPS. Fourteen out of 24 patients underwent total cavopulmonary connection (TCPC) with 8 to 15 months of interval from BCPS, and have been currently surviving, and in NYHA functional class I or II, except 2 patients who underwent take-down. In conclusion, early and midterm outcome after staged operation appears to be excellent. BCPS is a good interim procedure, in part because one can more properly select patients undergoing Fontan operations from the PVRI point of view.

Adolescent↗
PubMed1998

Effects of various factors including the CYP2D6 genotype and coadministration of flunitrazepam on the steady-state plasma concentrations of bromperidol and its reduced metabolite.

The effects of various factors, including the cytochrome P450 (CYP) 2D6 genotype and the coadministration of flunitrazepam, on the steady-state plasma concentrations (Css) of bromperidol and its reduced metabolite were studied in 62 schizophrenic inpatients receiving bromperidol 12 mg/day. By use of allele-specific PCR analysis, the wild type allele (CYP2D6*1A) and four mutated alleles causing either absent (CYP2D6*3, CYP2D6*4 and CYP2D6*5) or decreased (CYP2D6*10) CYP2D6 activity were identified. The means (ranges) of the Css of bromperidol and reduced bromperidol corrected to the median body weight were 7.2 (1.3-17.4) and 2.2 (0.4-8.9) ng/ml, respectively. Neither the Css of bromperidol nor that of reduced bromperidol significantly differed among the patients with no (n = 28), one (n = 30) and two mutated alleles (n = 4). The patients coadministered with flunitrazepam (n = 52) had significantly (P < 0.05) higher Css of bromperidol, but not reduced bromperidol, than those not (n = 10). Age, sex and smoking had no significant effects on the Css of these compounds. The present study thus suggests that the polymorphic CYP2D6 is not involved in the metabolism of bromperidol and reduced bromperidol to a major extent. The coadministration of flunitrazepam inhibits the metabolism of bromperidol, but age, sex and smoking do not affect it.

Adolescent↗
PubMed1998

Dependence of short-latency ocular following and associated activity in the medial superior temporal area (MST) on ocular vergence.

Motion of a large-field pattern elicits short-latency ocular following responses (OFR) in the monkey, which are mediated at least in part by the medial superior temporal area of the cortex (MST). The magnitude of the OFR is known to be inversely related to viewing distance, and we investigated the dependence of OFR and the associated neuronal activity in the MST on a major cue to viewing distance, ocular vergence, in alert monkeys (Macaca fuscata). The vergence angle, expressed in terms of the apparent viewing distance, ranged from infinity to 16.6 cm (0-6 m(-1)). The magnitude of the initial OFR increased monotonically with increases in convergence at a mean (+/-SD) rate of 19.6+/-4.5%/m(-1) in four monkeys (over the range 0-4 m(-1)). In two monkeys, we recorded the single unit activity of 160 MST neurons that responded to motion of a large-field pattern with directional selectivity. The mean latency (+/-SD) of the MST discharges elicited by large-field motion was 50+/-7.5 ms (n=115), which preceded the onset of OFR by an average of 10+/-9.9 ms. The discharge modulation elicited by large-field motion showed a significant dependence on vergence in 91/160 neurons (57%), 72 of which (79%) increased their firing rate with increasing convergence ("near" neurons), and the remainder increasing their firing rate with decreasing convergence ("far" neurons). However, on average, the sensitivity of these MST neurons to vergence was only about 30% of that shown by the OFR. It could be that only those neurons that are very sensitive to vergence angle contribute to the OFR, but it is also possible that much of the modulation of OFR with vergence occurs downstream from the MST or in alternative pathways (yet to be discovered) that contribute to OFR.

Animals↗
PubMed1998

Lymphocytic adenohypophysitis: skull radiographs and MRI.

We report the skull radiograph, CT and MRI findings in three patients with lymphocytic adenohypophysitis mimicking pituitary adenoma. All cases were associated with pregnancy. CT demonstrated a pituitary mass but did not differentiate lymphocytic adenohypophysitis from pituitary adenoma. The skull radiographs showed either a normal sella turcica or minimal abnormalities; they did not show ballooning or destruction. The MRI appearances were distinctive: relatively low signal on T1-weighted images; preservation of the bright posterior pituitary lobe despite the presence of a relatively large pituitary mass, less common in macroadenomas; marked contrast enhancement compared with pituitary macroadenomas; and dural enhancement adjacent to a pituitary mass.

Adenoma↗
PubMed1998

Role of positron emission tomography using fluorine-18 fluoro-2-deoxyglucose in predicting improvement in left ventricular function in patients with idiopathic dilated cardiomyopathy.

Improvement in left ventricular (LV) function in patients with idiopathic dilated cardiomyopathy (DCM) by medical treatment has been suggested. Thus, it is important to evaluate which patients will respond to medical therapy. Positron emission tomography (PET) with fluorine-18 fluoro-2-deoxyglucose (FDG) and cardiac catheterization were performed in 20 patients with DCM before the initiation of medical therapy. The regional myocardial glucose utilization rate (rMGU) was measured with FDG PET. Subjects were divided into two groups, group 1 (event-free patients, n=10) and group 2 (clinical cardiac events, n=10). Haemodynamic and PET parameters before the initiation of medication were compared between the two groups and between patients with and patients without improvement in LV function. Ejection fraction (EF) was significantly higher in group 1 (35.8%+/-9.0%) than in group 2 (24.8%+/-7.0%) and LV end-diastolic pressure (LVEDP) was significantly lower in group 1 (8.4+/-1.7 mmHg) than in group 2 (11.6+/-3.5 mmHg). Average rMGU (mg min-1 100 g-1) was similar in group 1 (11.2+/-2.5 mg min-1 100 g-1) and group 2 (11.2+/-2.9 mg min-1 100 g-1), while %CV of rMGU was significantly lower in group 1 (11.1%+/-6.3%) than in group 2 (29. 9%+/-13.9%, P<0.01). Furthermore, LV function normalized in seven patients in group 1. In these seven patients, EF (35.1%+/-10.9%), LVEDP (8.2+/-2.0 mmHg) and average rMGU (11.8+/-2.7 mg min-1 100 g-1) were comparable with those in patients without LV functional improvement (EF: 31.6%+/-9.1%; LVEDP: 10.7+/-3.3 mmHg; average rMGU: 10.8+/- 2.7 mg min-1 100 g-1). However,% CV of rMGU in patients with LV functional improvement (9.6%+/-5.6%) was significantly lower than in those without such improvement (26.3%+/-14.1%, P<0.01). %CV of rMGU <13.6% predicted prognosis with a sensitivity of 80%, a specificity of 100% and an accuracy of 90%. %CV of rMGU <13.6% also predicted improvement in LV function, with a sensitivity of 75%, a specificity of 92% and an accuracy of 85%. However, EF failed to predict improvement of LV function. In is concluded that homogeneous myocardial glucose utilization rate can predict both prognosis and improvement in LV function achieved by medical therapy in patients with DCM.

Adult↗
PubMed1998

Estimation of deadtime in imaging human subjects.

Deadtime count loss may cause significant errors in quantitative measurements performed by scintigraphic techniques. Deadtime depends on the scattering condition, and may vary among patients. The aim of this study was to estimate deadtime in imaging a human subject. A cylindrical phantom filled with various concentrations of technetium-99m solution was imaged with a gamma camera, and deadtime was assessed assuming the gamma camera to be a non-paralysable system (multi-dose method). Deadtime for the cylindrical phantom was also estimated using a point source to monitor counting efficiency (reference source method). Radionuclide angiography of the chest was performed in 38 patients with a reference source in the field of view, and temporal changes in count loss were assessed. The deadtime for each patient was calculated by the reference source method. The deadtime for the cylindrical phantom was estimated as 6.96+/-0.09 micros by the reference source method and was almost identical to that obtained by the multi-dose method (6.93 micros). Deadtime ranged from 6.01 to 9.58 micros in radionuclide angiography, and was positively correlated with the ratio of body weight in kg to body height in cm (y=8.566 x+5.611, r=0.869). Count loss was successfully corrected using the deadtime predicted with the regression line. In summary, the deadtime in imaging a human subject has a wide range and is related to the constitution of the subject. Correction for count loss using a deadtime predicted for each patient may be helpful in improving the reliability of quantitative nuclear medicine.

Body Constitution↗
PubMed1998

Hearing in the MRL/lpr mouse as a possible model of immune-mediated sensorineural hearing loss.

In order to clarify the possible mechanism of hearing loss in immune-mediated sensorineural hearing loss, basic research needed includes animal model studies. In the present investigation, we examined hearing thresholds and cochlear histologies of the MRL/lpr mouse which is now well-known as a model for pathology consistent with systemic lupus erythematosis (SLE). Present findings demonstrated that there were no statistically significant differences in auditory brainstem response (ABR) thresholds between 4- to 6-week-old "young" and 20- to 25-week-old "old" MRL mice. These differences were not sex-dependent. Under light microscopy, there were no abnormal morphological findings in the cochleas of either young or old MRL mice. With immunohistochemistry, mouse IgG was detected around the capillary walls in the stria vascularis in both young and old MRL mice. Serum IgG level of the MRL mice significantly decreased after predonisolone (PSL) administration. However, expression of mouse IgG in the stria vascularis was not observed in the MRL mice after PSL administration. From these results, we speculate that the hearing of the MRL mouse does not always deteriorate, and the deposition of mouse IgG on the capillary wall in the stria vascularis is not a sufficient factor to induce hearing loss. At this point, we conclude that the MRL mouse should not be considered a useful model for immune-mediated sensorineural hearing loss.

Aging↗