Search PubMed⌕ Search

Biomedical subjects

T Ozawa

Publications and source records attributed to T Ozawa.

At least 73 records · Page 4Linked to original sources
PubMed2001

Functional modification of cytochrome c by peroxynitrite in an electron transfer reaction.

The redox reaction of cytochrome c after modification with peroxynitrite under physiological conditions was investigated. Cytochrome c was treated with a bolus of synthetic peroxynitrite at a sub-millimolar concentration, and then subjected to reduction by superoxide and oxidation by hydrogen peroxide. The ability for the membrane potential formation in the mitochondrial respiratory chain was also evaluated. After the treatment with peroxynitrite, the cytochrome c molecule was mono-nitrated mainly at a tyrosine residue, using liquid chromatography-electrospray ionizing mass spectrometry (LC-ESI-MS) and HPLC. Although the redox capacity of cytochrome c was not affected by the peroxynitrite treatment, the oxidation of ferrocytochrome c to ferricytochrome c by hydrogen peroxide was accelerated. When cytochrome c was treated with peroxynitrite in the presence of 5-methoxytryptamine, an inhibitor for the tyrosine nitration by peroxynitrite, the acceleration of hydrogen peroxide-mediated oxidation was suppressed. It was also found that the formation of membrane potential in the rat liver mitochondria was suppressed when peroxynitrite-treated cytochrome c was used instead of the intact cytochrome c in vitro. From these results, we concluded that the peroxynitrite-treated cytochrome c was nitrated at a tyrosine residue and became more susceptible to oxidation by hydrogen peroxide, concomitantly losing the ability to transfer electrons in the mitochondrial respiratory chain. It is suggested that the peroxynitrite-induced modification of cytochrome c increases the susceptibility to non-physiological oxidants, and may cause dysfunction of mitochondria by suppressing of membrane potential.

Animals↗
PubMed2001

Relationship between effects of phenolic compounds on the generation of free radicals from lactoperoxidase-catalyzed oxidation of NAD(P)H or GSH and their DPPH scavenging ability.

The influence of various phenolic compounds on the lactoperoxidase (LPO)/hydrogen peroxide (H2O2)-catalyzed oxidation of biochemical reductants such as reduced beta-nicotinamide adenine dinucleotide (NADH), reduced beta-nicotinamide adenine dinucleotide phosphate (NADPH) or reduced glutathione (GSH) was investigated by electron spin resonance (ESR) spectroscopy. Micromolar quantities of phenolic compounds such as 17beta-estradiol, phenol, and p-chlorophenol enhanced the LPO/H2O2-catalyzed oxidation of NAD(P)H or GSH to generate a large amount of superoxide radical (O2*-) or glutathione thiyl radical (GS*), while, phenolic compounds such as quercetin and Trolox C greatly suppressed the generation of O2*- and GS*. In order to elucidate the effects of phenolic compounds on the generation of O2*- and GS*, their quenching activities for a stable radical, 1,1-diphenyl-2-picrylhydrazyl (DPPH), were investigated by ESR spectroscopy. 17beta-Estradiol, phenol, and p-chlorophenol showed very weak scavenging activities for DPPH, but quercetin and Trolox C showed strong activities. This suggests that the ability of phenolic compounds to enhance LPO/H2O2-catalyzed oxidation of NAD(P)H or GSH relates inversely to their ability to quench DPPH. That is, phenolic compounds having weak quenching activity against DPPH may enhance the LPO/H2O2-catalyzed oxidation of NAD(P)H or GSH to generate a large amount of O2*- or GS*.

Bepridil↗
PubMed2001

Basilar artery vasculitis secondary to sphenoid sinusitis--case report.

A 35-year-old male presented with basilar artery vasculitis secondary to sphenoid sinusitis manifesting as rapidly deteriorating symptoms including consciousness disturbance and right hemiparesis. Computed tomography (CT) on admission showed sphenoid sinusitis without intracranial lesion. Emergency angiography demonstrated basilar artery stenosis. The neurological deterioration was considered to be caused by ischemia of the perforating arteries branching from the stenotic portion of the basilar artery. The patient was treated with urokinase infusion through a microcatheter just proximal to the stenosis 3 hours after the onset of the symptoms. His consciousness level and right hemiparesis markedly improved immediately after the procedure. Magnetic resonance (MR) imaging on day 5 revealed that extension of the sphenoid sinusitis into the prepontine cistern had formed an abscess which was attached to the clivus. The basilar artery was embedded in the abscess at the angiographic stenosis. Cerebrospinal fluid (CSF) analysis showed white blood cell count of 601/mm3 with 82% neutrophils, 89.2 mg/dl protein, and 31 mg/dl glucose. No causative organism in the CSF could be identified by smear or culture. Early MR imaging and CSF examination are recommended when patients present with both ischemic symptoms involving the basilar artery and opacification of the sphenoid sinus on CT to identify basilar artery vasculitis secondary to sphenoid sinusitis.

Adult↗
PubMed2001

[Nitric oxide-dependent inhibition of platelet aggregation in elderly patients with hypercholesterolemia].

Hypercholesterolemia reduces production of nitric oxide (NO), a potent inhibitor of platelet aggregation, in endothelial cells. Recently platelet has been found to have NO synthase. Hypercholesterolemia may influence platelet NO production. We investigated NO-dependent inhibition of platelet aggregation in elderly hypercholesterolemic patients with total cholesterol (Tchol) of 240 mg/dl or more (n = 21). In elderly controls with Tchol less than 240 mg/dl (n = 61), L-arginine (5-50 mM) inhibited ADP-induced platelet aggregation in a dose-dependent manner (42.4% inhibition at 50 mM). However, L-arginine did not inhibit platelet aggregation in elderly hypercholesterolemic patients. L-arginine increased cyclic GMP production in elderly controls, but not in hypercholesterolemic patients (p < 0.02). Hypercholesterolemic patients showed increased platelet aggregation compared with elderly controls(p = 0.018). L-nitro-arginine methyl ester 12.5-50 uM increased platelet aggregation in both groups. Superoxide dismutase improved L-arginine inhibition of platelet aggregation in elderly hypercholesterolemic patients (p = 0.02). LDL cholesterol of 160 mg/dl or more was an independent predictor for loss of L-arginine inhibition of platelet aggregation (relative risk 3.9, p = 0.0098). This result suggests that hypercholesterolemia causes decreased NO-dependent inhibition of platelet aggregation due to reduced NO utilization. NO-dependent platelet aggregation may be a powerful tool for detection of vascular injury.

Aged↗
PubMed2001

Ryanodine-sensitive Ca2+ release mechanism in non-excitable cells (Review).

The properties of a ryanodine-sensitive Ca2+ release channel (receptor) in non-excitable cells like exocrine cells or epithelial cells are described in this review. The ryanodine-sensitive Ca2+ release from the microsomal vesicles is activated by Ca2+, caffeine, ryanodine or cyclic ADP-ribose (cADPR) and is inhibited by ruthenium red or higher concentrations (> or =100 microM) of ryanodine. The properties are similar to those of excitable cells such as muscle cells or neuronal tissues. In some non-excitable cells, the Ca2+ release induced by caffeine, ryanodine or cADPR is stimulated by calmodulin (CaM) or FK506. Kd values of [3H]ryanodine binding to the receptor protein range from 6 to 17 nM and are similar to those of a high-affinity binding site in skeletal or cardiac muscle. Maximum binding capacities (Bmax) range from 40 to 620 fmol/ mg protein and are 10 approximately 200-fold lower than those for a high-affinity binding site in skeletal muscle. Caffeine, adenine nucleotide AMP-PCP, Mg2+, ruthenium red or FK506 affects the binding. In some non-excitable cells, the ryanodine receptor (RyR) isoform RyR2 or RyR3 is expressed and has been identified. However, unlike for excitable cells, information concerning the RyR proteins, including binding sites for modulators like CaM and phosphorylation sites has not yet been obtained.

Animals↗
PubMed2001

[EEG findings in a case of acute necrotizing encephalopathy of childhood associated with influenza A virus infection].

We reported here a case of symptomatic partial epilepsy following acute necrotizing encephalopathy of childhood associated with influenza A virus infection. This 2-year-old boy underwent repeated EEG recordings, which at the acute stage was dominated by diffuse 1-2 Hz slow waves. The background activity was 5 Hz theta waves on the 49th day. Paroxysmal activities appeared after the 89th day of illness. On the 231st day, EEG showed spike-and-waves on the left and right frontal areas. Interestingly, paroxysmal activities preceded the onset of epileptic seizures by 7 months, and spike-and-waves by 2 months. After 10 months, he had generalized seizures with fever, and partial seizures on awakening without fever. Interictal EEG showed spike-and-waves on the bilateral frontal areas, and diffuse polyspikes and slow waves were occasionally seen. Though the background activity improved, his consciousness level did not recover probably because the thalamus, basal ganglia, brainstem were damaged more severely than the cerebral cortex.

Child, Preschool↗
PubMed2001

[The usefulness of intra-operative angiography during off-pump CABG].

BACKGROUND: Recently OPCAB has become more and more common in CABG, although one problem of OPCAB like the quality of anastomosis has not been solved yet. We discussed the usefulness of intra-operative angiography during OPCAB. METHODS: During March 1997-July 2000, 55 patients underwent OPCAB (including 35 MIDCAB cases) in our institute. Graft flow and anastomosis were examined by intra-operative or immediate post-operative angiography. RESULTS: Immediate postoperative angiography was performed in 22 MIDCAB cases. In 15 cases an excellent graft flow without stenosis could be confirmed, in 7 cases the native LAD was so small, which caused a poor flow of the LITA and three cases needed additional PTCA because of anastomosis stenosis. Intra-operative angiography was done in 9 MIDCAB and 17 OPCAB. As a result, re-anastomosis was performed in one case of MIDCAB because of severe anastomosis stenosis, one case of OPCAB had confirmed poor flow because of a small LAD, but in 24 cases an excellent graft flow could be seen on time. In 2 cases OPCAB was combined with PTCA to achieve complete revascularization. CONCLUSIONS: Intra-operative angiography is a useful strategy to confirm the surgical results quickly in OPCAB (included MIDCAB). Using such a strategy, combination of OPCAB and PTCA as a new approach for complete revascularization can be performed safe and smooth.

Aged↗
PubMed2001

[Laser pointer and laser safety standard].

BACKGROUND: In recent years, cheap laser pointers have been used as toys for children. The safety of laser pointers has become a public issue. CASES: Many cases of visual disturbance without retinal symptoms were reported, and those might be of a psychogenic nature or due to afterimage effects. However, four cases of macular retinal pigment epithelial disturbances due to radiation of a laser pointer beam were reported. One patient was affected in both eyes because of alternate laser beam radiation to both eyes. The exposure time to the laser beam exceeded 10 seconds. All cases except one were young patients. For the safety, it is important that handheld laser pointers be kept away from children. Toy type laser pointers should be treated as Class 1 laser products according to the Consumer Affairs Council, Ministry of Economy, Trade, and Industry.

Consumer Product Safety↗
PubMed2001

[Retinal hazard from blue light emitting diode].

PURPOSE: To compare the effect of exposure time from a blue(460 nm) light emitting diode(LED) on the morphology of the outer retina and determine conditions where damage occurs. MATERIALS AND METHODS: Young adult rhesus monkeys were anesthetized, and received blue LED exposure from a modified slit-lamp. A 3 mm beam of 0.85 mW was imaged onto the retina through a lens positioned before the cornea and exposure damage was determined at time intervals for 12 to 90 min. Fundus photography, fluorescein angiography(FAG), retinal tomography(HRT), and s-cone electororetinogram(S-ERG) were recorded at baseline, 2, and 30 days. RESULTS: Two days after 40 min exposure, there was a grey, discolored region, which was over-fluorescent in FAG, and an incresse in HRT and S-ERG corresponding to the site which was exposed to LED light. In histological examination at 30 days, the LED had caused produced a marked disruption of the disks of photoreceptor cells, damaged retinal pigment epithelium(RPE) apical villi, and a loss of RPE melanin after 90 min exposure. CONCLUSION: A threshold level was found around 40 min. This morphological damage may impair function and continuous exposure to blue light is potentially dangerous to vision.

Animals↗
PubMed2001

[A 50-year history of new drugs in Japan: the developments and trends of antihyperlipidemic drugs].

The developments and trends of antihyperlipidemic drugs and their effects on the mortality of coronary heart disease in Japan were investigated. The developed drugs available for hyperlipidemia were recorded with their approval dates by the Ministry of Health and Welfare (Table 1). 1. Antihyperlipidemic drugs have been developed since the late 1950s. Useful drugs among them include the fibrate series and the statin (HMG-CoA reductase inhibitor) series. Clofibrate, developed in 1965, was the first fibrate drug, and pravastatin sodium (Mevalotin(R) Sankyo Co.), developed in 1989, was the first statin drug. They have sure effectiveness for lowering serum cholesterol and triglyceride. But they induce an unfavorable side-effect, rhabdomyolisis, especially after the continuous or simultaneous use of both. The other drug classes using hyperlipidemia include various different types, e.g., probucol, nicotinates, anion exchange resins, ethyl icosapentate, and dextran sodium sulfate. Despite their mild activities, the low incidence of adverse effects make them suitable for supplementary use with fibrates or statins. 2. "Guideline for Diagnosis and Treatment of Hyperlipidemia in Adult" was presented by the Japan Atherosclersis Society in 1997. The standard criteria of serum cholesterol and triglyceride levels in Japanese adults were proposed. The hypercholesterolemia is the state of more than a 220 mg/dl level of serum cholesterol, and hypertriglyceridemia is of more than a 150 mg/dl level of serum triglyceride. The pharmacotherapy should be applied for a high serum level of cholesterol exceeding 240 mg/dl. But the standard routine formula of drug therapy were not indicated in the present guideline. 3. Epidemiological surveys show that hyperlipidemia induces coronary heart diseases in the United States, European countries, and Japan. The mortality of all heart disease patients in Japan increased rapidly from the late 1960s, but the mortality resulting from coronary heart disease was suppressed from 1968. This suppression continued throughout 1994 when artificial statistical changes occurred. It may be due to the newly developed antihyperlipidemic drugs, e.g., the clofibrate group, the statin series, and others (Fig.2).

Coronary Disease↗
PubMed2000

Fluorescent indicators for cyclic GMP based on cyclic GMP-dependent protein kinase Ialpha and green fluorescent proteins.

We describe herein fluorescent indicators for cyclic GMP (cGMP) in single living cells. cGMP-dependent protein kinase Ialpha (PKG Ialpha), a receptor for cGMP, was fused with blue- and red-shifted green fluorescent proteins (GFPs) to its N- and C-termini, respectively. Using PKG lalpha delta1-47, in which the dimerization domain was deleted, fluorescence resonance energy transfer between the GFPs was found to increase upon cGMP-induced conformational change in PKG Ialpha delta1-47. We demonstrated that thus-developed fluorescent indicators reversibly responded to cGMP that was produced in nitric oxide-stimulated HEK293 cells. The present genetically encoded fluorescent indicators open a way not only for understanding the dynamics of cGMP signaling in single cells and organisms but also for discovering pharmaceuticals such as isoform-specific inhibitors for phosphodiesterases.

Animals↗
PubMed2000

Inhibition of thrombin by sulfated polysaccharides isolated from green algae.

Eight different sulfated polysaccharides were isolated from Chlorophyta. All exhibited thrombin inhibition through a heparin cofactor II (HCII)-dependent pathway, and their effects on the inhibition of thrombin were more potent than those of heparin or dermatan sulfate. In particular, remarkably potent thrombin inhibition was found for the sulfated polysaccharides isolated from the Codiales. In the presence of these sulfated polysaccharides, both the recombinant HCII (rHCII) variants Lys(173)-->Leu and Arg(189)-->His, which are defective in interactions with heparin and dermatan sulfate, respectively, inhibited thrombin in a manner similar to native rHCII. This result indicates that the binding site of HCII for each of these eight sulfated polysaccharides is different from the heparin- or dermatan sulfate-binding site. All the sulfated polysaccharides but RS-2 significantly stimulated the inhibition of thrombin by an N-terminal deletion mutant of HCII (rHCII-Delta74). Furthermore, hirudin(54-65) decreased only 2-5-fold the rate of thrombin inhibition by HCII stimulated by the sulfated polysaccharides, while HD22, a single-stranded DNA aptamer that binds exosite II of thrombin, produced an approximately 10-fold reduction in this rate. These results suggest that, unlike heparin and dermatan sulfate, the sulfated polysaccharides isolated from Chlorophyta activate HCII primarily by an allosteric mechanism different from displacement and template mechanisms.

Antithrombins↗
PubMed2000

How can Ca2+ selectively activate recoverin in the presence of Mg2+? Surface plasmon resonance and FT-IR spectroscopic studies.

We investigated the relationship between metal ion selective conformational changes of recoverin and its metal-bound coordination structures. Recoverin is a 23 kDa heterogeneously myristoylated Ca(2+)-binding protein that inhibits rhodopsin kinase. Upon accommodating two Ca(2+) ions, recoverin extrudes a myristoyl group and associates with the lipid bilayer membrane, which was monitored by the surface plasmon resonance (SPR) technique. Large changes in SPR signals were observed for Sr(2+), Ba(2+), Cd(2+), and Mn(2+) as well as Ca(2+), indicating that upon binding to these ions, recoverin underwent a large conformational change to extrude the myristoyl group, and thereby interacted with lipid membranes. In contrast, no SPR signal was induced by Mg(2+), confirming that even though it accommodates two Mg(2+) ions, recoverin does not induce the large conformational change. To investigate the coordination structures of metal-bound Ca(2+) binding sites, FT-IR studies were performed. The EF-hands, Ca(2+)-binding regions each comprising 12 residues, arrange to coordinate Ca(2+) with seven oxygen ligands, two of which are provided by a conserved bidentate Glu at the 12th relative position in the EF-hand. FT-IR analysis confirmed that Sr(2+), Ba(2+), Cd(2+), and Mn(2+) were coordinated to COO(-) of Glu by a bidentate state as well as Ca(2+), while coordination of COO(-) with Mg(2+) was a pseudobridging state with six-coordinate geometry. These SPR and FT-IR results taken together reveal that metal ions with seven-coordinate geometry in the EF-hands induce a large conformational change in recoverin so that it extrudes the myristoyl group, while metal ions with six-coordinate geometry in the EF-hands such as Mg(2+) remain the myristoyl group sequestered in recoverin.

Animals↗
PubMed2000

Reaction mechanism of electron transfer from FeII(CN)6(4-) or W(IV)(CN)8(4-) to the cupric ions in human copper, zinc superoxide dismutase.

The electron transfer reactions from FeII(CN)6(4-) and W(IV)(CN)8(4-) to the cupric ions in human copper, zinc superoxide dismutase were followed by the micro-stopped-flow method. The kinetic rate data clearly indicate that FeII(CN)6(4-) or W(IV)(CN)8(4-) first forms an adduct with the enzyme through the interaction with Arg143 of the active cavity and then an electron from FeII(CN)6(4-) or W(IV)(CN)8(4-) of the adduct transfers to the cupric ion in the enzyme. The dissociation constants of the adducts of FeII(CN)6(4-) and W(IV)(CN)8(4-) were 4.0(+/-0.3) x 10(-3) and 2.2(+/-0.3) x 10(-3) M, respectively. In spite of the difference between the standard redox potentials of FeIII(CN)6(3-)/FeII(CN)6(4-) (468 mV) and W(V)(CN)8(3-)/W(IV)(CN)8(4-) (556 mV), the electron transfer rate constant (0.148(+/-0.005) s(-1) of FeII(CN)6(4-) at 25 degrees C is very similar to that of W(IV)(CN)8(4-) (0.072(+/-0.011) s(-1)). The entropy values of the adduct formations and the activation energies of the electron transfer rates were determined by the temperature dependence of the dissociation constants of the adducts and the electron transfer rates. The enthalpy values of the formation of adducts are almost zero, so that the driving forces to form the adducts are mainly derived from the entropy. The activation energy of the electron transfer rate of FeII(CN)6(4-) is very similar to that of W(IV)(CN)8(4-). The formation of the adduct between FeII(CN)6(4-) and the enzyme was inhibited by the presence of various anions (ClO4-, SO4(2-), SCN-, and N3-). The bulky anions SO4(2-) and ClO4- behave as competitive inhibitors for FeII(CN)6(4-); these anions should interact mainly with Arg143, as it has a positive charge at the entrance of the active cavity. The competitive inhibition constants of ClO4-, SO4(2-), and SCN- were 0.010, 0.012, and 0.008 M. The azide ion, which is smaller than SO4(2-) or ClO4-, shows mixed inhibition, because N3- can interact with Arg143 (competitive inhibition) and also directly binds to the cupric ion in h-SOD (noncompetitive inhibition). The competitive and noncompetitive inhibition constants of N3- were 0.004 and 0.016 M, respectively.

Anions↗
PubMed2000

A fluorescent indicator for detecting protein-protein interactions in vivo based on protein splicing.

We describe a new method with general applicability for monitoring any protein-protein interaction in vivo. The principle is based on a protein splicing system, which involves a self-catalyzed excision of protein splicing elements, or inteins, from flanking polypeptide sequences, or exteins, leading to formation of a new protein in which the exteins are linked directly by a peptide bond. As the exteins, split N- and C-terminal halves of enhanced green fluorescent protein (EGFP) were used. When a single peptide consisting of an intein derived from Saccharomyces cerevisiae intervening the split EGFP was expressed in Escherichia coli, the two external regions of EGFP were ligated, thereby forming the EGFP corresponding fluorophore. Genetic alteration of the intein, which involved large deletion of the central region encoding 104 amino acids, was performed. In the expression of the residual N- and C-terminal intein fragments each fused to the split EGFP exteins, the splicing in trans did not proceed. However, upon coexpression of calmodulin and its target peptide M13, each connected to the N- and C-terminal inteins, fluorescence of EGFP was observed. These results demonstrate that interaction of calmodulin and M13 triggers the refolding of intein, which induces the protein splicing, thereby folding the ligated extein correctly for yielding the EGFP fluorophore. This method opens a new way not only to screen protein-protein interactions but also to visualize the interaction in vivo in transgenic animals.

Calmodulin↗