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H Miyoshi

Publications and source records attributed to H Miyoshi.

At least 163 records · Page 9Linked to original sources
PubMed1996

Comparison of the structural features of ubiquinone reduction sites between glucose dehydrogenase in Escherichia coli and bovine heart mitochondrial complex I.

To characterize the structural features of the ubiquinone reduction site of glucose dehydrogenase (GlcDH) in Escherichia coli, we performed structure/activity studies of a systematic set of synthetic ubiquinone analogues and specific inhibitors (synthetic capsaicins) of this site. Considering the proposed similarity of the quinone binding domain motif between GlcDH and one subunit of mitochondrial complex I [Friedrich, T., Strohdeicher, M., Hofhaus, G., Preis, D., Sahm, H. & Weiss, H. (1990) FEBS Lett. 265, 37-40], we compared the structure/activity profiles of the substrates and inhibitors for GlcDH with those for bovine heart mitochondrial complex i. With respect to GlcDH, replacement of one or both methoxy groups in the 2 and 3 positions of benzoquinone ring by ethoxy group(s) resulted in a drastic decrease in the electron accepting activity. The presence of a 5-methyl group and the conformational property of the 6-alkyl side chain did not significantly contribute to the activity. These results suggested that only half of the benzoquinone ring (the moiety corresponding to the 2 and 3 positions) is recognized by the quinone reduction site in a strict sense. In contrast, quinone analogues with structural modifications at all positions in the benzoquinone ring retained the activity with mitochondrial complex I. This finding indicated that the catalytic site of complex I is spacious enough to accommodate a variety of structurally different quinone derivatives. The correlation of the inhibitory potencies of a series of synthetic capsaicins between the two enzymes was very poor. These findings indicated that the binding environment of ubiquinone in GlcDH is very specific and differs from that in mitochondrial complex I.

Animals↗
PubMed1996

Identification of a novel isoform of estrogen receptor, a potential inhibitor of estrogen action, in vascular smooth muscle cells.

Clinical and experimental studies showed that estrogen has antiatherogenic effects. We previously demonstrated that the estrogen receptor (ER) mRNA and protein are expressed in vascular smooth muscle cells (VSMC) derived from rat aorta. Here, the expression of isoforms of the ER was examined in VSMC. Reverse transcriptase-polymerase chain reaction using specific primers for rat ER cDNA was performed from RNA of rat VSMC. This revealed the existence of ER cDNA that is shorter than the wild-type ER cDNA. Sequencing of the amplified products identified three isoforms of the ER and the wild-type ER. These ER mRNA isoforms lacked the region corresponding to exon 4, exon 4 and 5, and exon 3 and 4. Therefore, they were designated as ERdelta4 isoform, ERdelta4/5 isoform and ERdelta3/4 isoform, respectively. Chloramphenicol acetyltransferase assay was performed with these ER isoforms constructed into the expression vector and the reporter plasmid containing the estrogen responsive element. The assay showed that these ER isoforms lost estrogen-dependent transactivation activities and that ERdelta4/5 isoform has a inhibitory effect on normal estrogen action when it was cotransfected with the wild-type ER. These ER isoforms might be involved in the regulation of VSMC by estrogen.

Amino Acid Sequence↗
PubMed1996

Comparison of the inhibitory action of synthetic capsaicin analogues with various NADH-ubiquinone oxidoreductases.

Capsaicin is a new naturally occurring inhibitor of proton-pumping NADH-ubiquinone oxidoreductase (NDH-1), that competitively acts against ubiquinone. A series of capsaicin analogues was synthesized to examine the structural factors required for the inhibitory action and to probe the structural property of the ubiquinone catalytic site of various NADH-ubiquinone reductases, including non-proton-pumping enzyme (NDH-2), from bovine heart mitochondria, potato tuber (Solanum tuberosum, L) mitochondria and Escherichia coli (GR 19N) plasma membranes. Some synthetic capsaicins were fairly potent inhibitors of each of the three NDH-1 compared with the potent rotenone and piericidin A. Synthetic capsaicin analogues inhibited all three NDH-1 activities in a competitive manner against an exogenous quinone. The modification both of the substitution pattern and of the number of methoxy groups on the benzene ring, which may be superimposable on the quinone ring of ubiquinone, did not drastically affect the inhibitory potency. In addition, alteration of the position of dipolar amide bond unit in the molecule and chemical modifications of this unit did not change the inhibitory potency, particularly with bovine heart and potato tuber NDH-1. These results might be explained assuming that the ubiquinone catalytic site of NDH-1 is spacious enough to accommodate a variety of structurally different capsaicin analogues in a dissimilar manner. Regarding the moiety corresponding to the alkyl side chain, a rigid diphenyl ether structure was more inhibitory than a flexible alkyl chain. Structure-activity studies and molecular orbital calculations suggested that a bent form is the active conformation of capsaicin analogues. On the other hand, poor correlations between the inhibitory potencies determined with the three NDH-1 suggested that the structural similarity of the ubiquinone catalytic sites of these enzymes is rather poor. The sensitivity to the inhibition by synthetic capsaicins remarkably differed between NDH-1 and NDH-2, supporting the notion that the sensitivity against capsaicin inhibition correlates well with the presence of an energy coupling site in the enzyme (Yagi, T. (1990) Arch. Biochem. Biophys. 281, 305-311). It is noteworthy that several synthetic capsaicins discriminated between NDH-1 and NDH-2 much better than natural capsaicin.

Animals↗
PubMed1996

Seroepidemiological study of human herpesvirus-6 and -7 in children of different ages and detection of these two viruses in throat swabs by polymerase chain reaction.

The presence of human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7) in throat swabs of 62 children of different age groups (group I, ages 0-5 month; group II, ages 6-11 months, group III, ages 12-23 months, group IV, age 2-8 years) and 28 adults was detected by polymerase chain reaction. The detection rate of HHV-6 DNA was the highest (87%) in children aged 1-year-old and decreased with age, whereas the detection rate of HHV-7 increased with age and reached a maximum in adults. HHV-6B was detected in almost all samples except for two children who secreted only HHV-6A. When the antibody prevalence was determined in the four groups of children, HHV-6 antibody was detected in 8/12 (66.7%), 10/12 (83.3%), 15/16 (93.8%), and 13/14 (92.9%), respectively. Antibody to HHV-7 in these groups was detected in 6/12 (50.0%), 4/12 (33.3%), 12/16 (75.0%), and 13/14 (92.9%), respectively. Detection of HHV-6 DNA in throat swabs of triplets who had the sequential onset of exanthem subitum was attempted by using samples sequentially collected from these children after the onset of the disease in the first patient. HHV-6 DNA with high copy numbers was detectable during the acute and convalescent phases of the disease in all patients, but no DNA was detected in samples collected before the onset of disease.

Adult↗
PubMed1996

Intracellular and extracellular Ca2+ regulate histamine-induced release of nitric oxide in vascular endothelial cells as shown with sensitive and selective nitric oxide electrodes.

The effects of various agents that alter the intracellular Ca2+ concentration and the extracellular Ca2+ concentration on histamine-induced release of nitric oxides (NO) from porcine aortic endothelial cells were studied using NO electrodes. The NO release induced by application of histamine (200 microM) in Ca(2+)-free solution was similar to that in solution with a normal Ca2+ concentration on its first application, but reduced on its second application. NO release was also suppressed by 1,2-bis (o-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA), which lowers the intracellular Ca2+ concentration, and was completely inhibited by Co2+, but it was not affected by verapamil, a voltage-dependent Ca2+ channel blocker. These results suggest that Ca2+ release from intracellular stores might play an important role in NO production, that influx of extracellular Ca2+ is required to refill the stores, and that this Ca2+ influx is not through voltage-dependent Ca2+ channels.

Animals↗
PubMed1996

Therapeutic efficacy of a case of pyruvate dehydrogenase complex deficiency monitored by localized proton magnetic resonance spectroscopy.

We experienced a case of pyruvate dehydrogenase deficiency observed by proton magnetic resonance spectroscopy (1H MRS). This case was diagnosed as West syndrome by characteristic convulsion and the periodic hypsarrhythmia pattern of EEG. At the age of 11 months, the first examination of 1H MRS revealed a high peak of lactate, and the high concentration of lactate and pyruvate was confirmed in sampled cerebrospinal fluid (CSF). Deficiency of pyruvate dehydrogenase complex was finally diagnosed by genetic examination. Dichloroacetate was administered to the patient as therapy. Decrease of lactate in the brain was found by 1H MRS. Lactate and pyruvate in the CSF was also decreased. In accordance with the suspension of dichloroacetate, increase of lactate in the brain was detected and the convulsions reappeared. After readministration of dichloroacetate, the patient was almost symptom free and lactate in the brain and CSF had decreased to the normal extent. We considered that 1H MRS provides useful information for screening metabolic disorders of infants and assessing the efficacy of therapy.

Brain↗
PubMed1996

Voltage-clamp studies of gap junctions between uterine muscle cells during term and preterm labor.

Gap junctions between myometrial cells increase dramatically during the final stages of pregnancy. To study the functional consequences, we have applied the double-whole-cell voltage-clamp technique to freshly isolated pairs of cells from rat circular and longitudinal myometrium. Junctional conductance was greater between circular muscle-cell pairs from rats delivering either at term (32 +/- 16 nS, mean +/- SD, n = 128) or preterm (26 +/- 17 nS, n = 33) compared with normal preterm (4.7 +/- 7.6 nS, n = 114) and postpartum (6.5 +/- 10 nS, n = 16); cell pairs from the longitudinal layer showed similar differences. The macroscopic gap junction currents decayed slowly from an instantaneous, constant-conductance level to a steady-state level described by quasisymmetrical Boltzmann functions of transjunctional voltage. In half of circular-layer cell pairs, the voltage dependence of myometrial gap junction conductance is more apparent at smaller transjunctional voltages (< 30 mV) than for other tissues expressing mainly connexin-43. This unusual degree of voltage dependence, although slow, operates over time intervals that are physiologically relevant for uterine muscle. Using weakly coupled pairs, we observed two unitary conductance states: 85 pS (85-90% of events) and 25 pS. These measurements of junctional conductance support the hypothesis that heightened electrical coupling between the smooth muscle cells of the uterine wall emerges late in pregnancy, in preparation for the massive, coordinate contractions of labor.

Adrenergic beta-Agonists↗
PubMed1996

Calpain activation in plasma membrane bleb formation during tert-butyl hydroperoxide-induced rat hepatocyte injury.

BACKGROUND & AIMS: The mechanism of plasma membrane blebbing (dissociation of the lipid bilayer from the membrane cytoskeleton) in hepatocyte injury is not known. The aim of this study was to investigate the role of calpain, a calcium-dependent cytosolic protease, in bleb formation induced by oxidative stress. METHODS: Hepatocytes from Wistar rats were injured with tertbutyl hydroperoxide in the presence or absence of ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) or a specific calpain inhibitor, calpeptin (Z-Leu-nLeu-H). Bleb formation was examined by phase-contrast and transmission electron microscopies. Intracellular calcium concentration was measured using Fura-2. Western blot analyses were performed for cytoskeletal proteins (talin, alpha-actinin, and vinculin) and the intermediate (activated) and proactivated forms of calpain mu. RESULTS: tert-Butyl hydroperoxide induced a sustained increase in intracellular calciu, bleb formation, and, ultimately, hepatocyte death. Talin and alpha-actinin were degraded in a time-dependent manner, although no apparent changes of actin filament were observed. Before the cytoskeletal protein degradation, the intermediate form of calpain mu appeared as its proactivated form decreased. In addition, calpeptin or EGTA inhibited not only calpain mu activation but also cytoskeletal protein degradation and bleb formation. CONCLUSIONS: In tert-butyl hydroperoxide-treated hepatocytes, the activation of calpain promotes membrane blebbing via degradation of cytoskeletal proteins.

Animals↗
PubMed1996

Uncoupling mechanism of glycoside antibiotic aculeximycin in isolated rat-liver mitochondria.

Effects of basic glycoside antibiotic aculeximycin (ACM) on the oxidative phosphorylation of rat-liver mitochondria were examined. ACM was shown to be a potent uncoupler of the oxidative phosphorylation. To cause the same extent of respiration release, higher concentration of ACM was required in phosphate (Pi)-free medium than in Pi medium. During the uncoupling caused by ACM in Pi medium, large amplitude swelling and oxidation of intramitochondrial NAD(P)H occurred, indicating that ACM remarkably enhances permeability of the inner mitochondrial membrane. The Pi uptake via Pi/H+ symporter was shown to play an important, but not essential, role in the uncoupling by ACM, indicating the increase in membrane permeability is mostly due to acceleration of Pi/H+ influx through Pi/H+ symporter activated by ACM. ACM is the first naturally occurring antibiotic, to our knowledge, which activates Pi/H+ symporter. However, since the inhibition of Pi/H+ symporter by N-ethylmaleimide did not completely abolish the uncoupling activity of ACM, and ACM induced the uncoupling even in Pi-free medium, an increase in the membrane permeability for other ions, such as Na+ and K+, due to a different action mechanism has also to be considered. On the other hand, positively charged amine local anesthetics, like dibucaine, prevented the uncoupling activity by ACM in both Pi and Pi-free medium. The uncoupling activity of N-diacetylated ACM lacking free amino groups was ca. 1/120th that of ACM, indicating that positively charged amino groups are important for the uncoupling activity. It is suggested that some specific interactions between positively charged amino groups of ACM and the binding site, which is probably negatively charged, are triggers that affect the permeability of the inner mitochondrial membrane. Amine local anesthetics may mask the negative charge of the binding site, thereby interfering with ACM binding.

Animals↗
PubMed1996

Direct interaction between mitochondrial succinate-ubiquinone and ubiquinol-cytochrome c oxidoreductases probed by sensitivity to quinone-related inhibitors.

The electron-transfer activities of bovine heart mitochondrial complexes I, II, and III, but not complex IV, were simultaneously inhibited by 2-alkyl-4,6-dinitrophenols to a different extent. The extent of inhibition of NADH and succinate oxidase activities by dinitrophenols was compared with that of individual complex activities using submitochondrial particles. The extent of inhibition of succinate oxidase activity by 1-methylpropyl and 1-methylbutyl derivatives was much larger than that of NADH oxidase activity. This large inhibition of succinate oxidase activity seemed not to be explainable by the extent of inhibition of individual complex activities (i.e., complexes II and III activities), based upon the homogeneous ubiquinone pool model. On the other hand, other dinitrophenols (n-propyl, 1-methylpentyl, 1-methylhexyl, and tert-butyl derivatives) very similar to the above compounds did not elicit such anomalous inhibitory action, indicating that the action of 1-methylpropyl and 1-methylbutyl derivatives is highly specific to their structure. The anomalous inhibition by these two compounds was also observed with the isolated succinate-cytochrome c oxidoreductase, in which there is no ubiquinone pool behavior [Rich, P.R. (1984) Biochim. Biophys. Acta 768, 53-79]. However, when the succinate-cytochrome c reductase of which the activity had been partially restored by adding phospholipid and exogenous quinone to the phospholipid- and ubiquinone-depleted succinate-cytochrome c reductase was assayed, the anomalous inhibitory action of interest was undetectable. These results indicated that electron-transfer between complexes II and III, which is mediated not only by free-form, but also by protein-bound ubiquinone, occurs in the mitochondrial membrane. The fact that the anomalous inhibition of succinate oxidase activity of submitochondrial particles was sensitive to changes in the external osmotic pressure which affected the total area of the particle supports this notion.

Animals↗
PubMed1996

Endotoxin-induced L-arginine pathway produces nitric oxide and modulates the Ca2+-activated K+ channel in cultured human dermal papilla cells.

Endotoxin includes an enzyme that synthesizes nitric oxide (NO) from l-arginine (NO synthase) in vascular smooth muscles cells, macrophages, and fibroblasts, leading to the release of NO. We evaluated the release of NO and its intracellular action on the Ca2+-activated K+ channel (KCa channel) in cultured human dermal papilla cells by use of the electron paramagnetic response (EPR) spin trapping method and the patch clamp technique. In dermal papilla cells pretreated for 24 h with endotoxin (1 microgram/microliter), application of 1 microM L-arginine generated NO, although no measurable release of NO was observed in cells without endotoxin pretreatment, as determined by the EPR spin trapping method. With the patch clamp technique, we found that the KCa channel of dermal papilla cells had high conductance and was voltage dependent. In addition, after endotoxin pretreatment, the extracellular application of 100 microM l-arginine modulated the KCa channel in the cell-attached patch configurations. In inside-out patch configuration, however, NO produced by L-arginine itself did not modulate the Kca channel. The modulation of the KCa channel was suppressed by pretreatment with 100 microM N omega-nitro-L arginine methyl ester, and inhibitor of inducible and constitutive NO synthases. Methylene blue, a blocker of guanylate cyclase, inhibited the L-arginine-induced activation of the Kca channel. Theses results indicate that the endotoxin-induced L-arginine pathway cell generates No, which consequently modulated the KCa channel in cultured human dermal papilla cells by increasing of cyclic GMP-dependent phosphorylation.

Arginine↗
PubMed1996

Long-term performance of albumin secretion of hepatocytes cultured in a packed-bed reactor utilizing porous resin.

A packed-bed reactor utilizing the porous polymer, polyvinyl formal (PVF) resin, as a supporting material was applied for the long-term culture of rat hepatocytes. Perfusion cultures using this reactor, as well as monolayer cultures, were performed for up to 1 week under 3 different media conditions. When serum-free or epidermal growth factor-containing medium was used, albumin secretion rates showed constant decreases in both the perfusion and the monolayer cultures, and approximately 10% of the activity exhibited on Day 1 was preserved at the end of the cultures. In contrast, the hepatocytes supplemented with serum-containing medium exhibited stable ability of albumin secretion throughout the culture period under the perfusion culture condition using PVF resin. From the microscopic observation, the immobilized hepatocytes in this medium revealed round shapes, and a cluster of cell aggregates was scarcely seen.

Albumins↗
PubMed1996

In vitro and in vivo antibacterial activities of CS-940, a new 6-fluoro-8-difluoromethoxy quinolone.

The in vitro and in vivo activities of CS-940, a new 6-fluoro-8-difluoromethoxy quinolone, were compared with those of ciprofloxacin, tosufloxacin, sparfloxacin, and levofloxacin. The in vitro activity of CS-940 against gram-positive bacteria was nearly equal to or greater than those of the other quinolones tested. In particular, CS-940 was two to eight times more active against methicillin-resistant Staphylococcus aureus than the other quinolones, at the MIC at which 90% of the clinical isolates are inhibited. Against gram-negative bacteria, the activity of CS-940 was comparable to or greater than those of tosufloxacin, sparfloxacin, and levofloxacin, while it was lower than that of ciprofloxacin. The activity of CS-940 was largely unaffected by medium, inoculum size, or the addition of horse serum, but it was decreased under acidic conditions, as was also seen with the other quinolones tested. CS-940 showed potent bactericidal activity against S. aureus, Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. In oral treatment of mouse systemic infections caused by S. aureus, Streptococcus pneumoniae, Streptococcus pyogenes, E. coli, K. pneumoniae, Serratia marcescens, and P. aeruginosa, CS-940 was more effective than ciprofloxacin, sparfloxacin, and levofloxacin against all strains tested. Against experimental pneumonia with K. pneumoniae in mice, CS-940 was the most effective of all the quinolones tested. These results suggest that CS-940 may be effective in the therapy of various bacterial infections.

Animals↗
PubMed1996

Effects of estrogen on atherosclerosis formation and serum nitrite/nitrate concentrations in cholesterol-fed ovariectomized rabbits.

This study was to examine the effects of estrogen replacement on atherosclerosis formation in ovariectomized cholesterol-fed rabbits. We also examined serum levels of nitrite/nitrate, stable metabolites of nitric oxide, to investigate the involvement of nitric oxide. Female New Zealand White rabbits were ovariectomized and divided into 3 groups; 1) fed a normal diet (ND group, n=5), 2) fed a 1% cholesterol diet (CD group, n=6), or 3) fed a 1% cholesterol diet and received estrogen replacement (CD+E group, n=7). After 3 months, the rabbits were sacrificed to examine atherosclerosis formation. Atherosclerosis was not observed in ND. The oil red 0 positive area in the aorta was significantly greater in CD than in CD+E (CD, 17.3+/-2.2; CD+E, 9.3+/-0.8%, p<0.05). Stenosis of the coronary artery was also significantly greater in CD than in CD+E (CD, 30.6+/-9.7; CD+E, 6.7+/-2.9%, p <0.05). There was no significant difference in serum lipids between CD and CD+E. Serum nitrite/nitrate levels were significantly lower in CD than in ND (ND, 37.6+/-3.6; CD, 25.3+/-3.1 microM, p<0.05). There was a non-significant trend towards higher nitrite/nitrate levels after estrogen replacement (CD+E, 34.4+/-3.8 microM, p=0.08 vs. CD). These results suggest that direct actions on vascular wall including nitric oxide production contribute to the anti-atherogenic effects of estrogen.

Animals↗
PubMed1996

[The indications for postoperative adjuvant therapy in node-negative breast cancer patients].

A clinicopathological study on prognosis has been carried out in 233 breast cancer patients with more than twenty inspected nodes and without lymph node metastasis. Multivariate analysis of clinicopathological findings in node-negative breast cancers showed that the best combination of clinical features predicting prognosis were age, menstruation status, tumor location and tumor diameter; and from 11 pathological features, three factors (histological type, histological grade and the grade of tumor infiltrating lymphocytes) were selected under condition of p<0.05. With solid-tubular carcinoma, histological grade III and negative tumor infiltrating lymphocytes as the 3 poor prognostic factors, node-negative breast cancers were classified. The recurrence rates of subgroups with the number of 3, 2, 1 and 0 were 28%, 16%, 7% and 0%, respectively. We conclude that the grade of tumor infiltrating lymphocytes is especially significant as the prognostic factor in node-negative breast cancers, and that postoperative adjuvant chemoendocrine therapy must be performed for patients with node-negative breast cancer with more than two poor prognostic pathological factors.

Adult↗
PubMed1996

[A patient with hepatocellular carcinoma originating from the caudate lobe, who underwent hepatectomy following microwave coagulation therapy].

Hepatocellular carcinoma (HCC) originating from the caudate lobe is rare (about 2%), and prognosis is poor due to difficulties in early diagnosis, transarterial chemoembolization (TAE), and surgical resection. Distant metastasis is also likely to develop. To prevent metastasis during surgical manipulations, we performed microwave coagulation therapy before caudate lobectomy in one patient. Although the efficacy of this procedure remains to be elucidated, the multidisciplinary treatment described in this study is essential for the improvement of prognosis in patients with HCC developing from caudate lobe.

Carcinoma, Hepatocellular↗
PubMed1995

Stabilization of a semiquinone radical at the high-affinity quinone-binding site (QH) of the Escherichia coli bo-type ubiquinol oxidase.

Reaction of ubiquinone in the high-affinity quinone-binding site (QH) in bo-type ubiquinol oxidase from Escherichia coli was revealed by EPR and optical studies. In the QH site, ubiquinol was shown to be oxidized to ubisemiquinone and to ubiquinone, while no semiquinone signal was detected in the oxidase isolated from mutant cells that cannot synthesize ubiquinone. The QH site highly stabilized ubisemiquinone radical with a stability constant of 1-4 at pH 8.5 and the stability became lower at the lower pH. Midpoint potential of QH2/Q couple was -2 mV at pH 8.5 and showed -60 mV/pH dependence indicative of 2H+/2e- reaction. The Em was more negative than that of low-spin heme b above pH 7.0. We conclude that the QH mediates intramolecular electron transfer from ubiquinol in the low-affinity quinol oxidation site (QL) to low-spin heme b. Unique roles of the quinone-binding sites in the bacterial ubiquinol oxidase are discussed.

Benzoquinones↗