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Biomedical subjects

H Kanazawa

Publications and source records attributed to H Kanazawa.

At least 55 records · Page 3Linked to original sources
PubMed2000

Role of endogenous nitric oxide in exercise-induced airway narrowing in patients with bronchial asthma.

BACKGROUND: Increased amounts of nitric oxide (NO) in expired air and induced sputum have been found in asthmatic patients, and the role of excessively produced NO in the pathogenesis of bronchial asthma is under active investigation. OBJECTIVE: This study was designed to investigate the involvement of endogenous NO in exercise-induced bronchoconstriction (EIB) in asthmatic patients by using the sputum induction method. METHODS: The concentration of NO derivatives and inflammatory indices in induced sputum were examined in 18 asthmatic subjects and 10 normal control subjects. All asthmatic subjects performed an exercise test for 6 minutes. For 8 weeks after the first exercise testing, 400 microg of beclomethasone dipropionate twice daily was administered for asthmatic subjects with EIB, and the exercise testing and sputum induction were repeated in these patients. RESULTS: The concentration of NO derivatives in induced sputum was significantly higher in 9 asthmatic subjects with EIB (1580 +/- 280 micromol/L) than in 9 asthmatic subjects without EIB (1130 +/- 210 micromol/L) and normal control subjects (510 +/- 150 micromol/L). Moreover, there was a significant correlation between the concentration of NO derivatives and the percentage of maximal fall in FEV(1) (r = 0.569, P =.019). The concentration of NO derivatives was also more closely correlated with the area under the curve of the percentage fall in FEV(1) plotted against time for 30 minutes (AUC(0-30); r = 0.812, P <.001). After treatment with inhaled beclomethasone dipropionate in asthmatic subjects with EIB, there was a significant decrease in the concentration of NO derivatives in induced sputum. The change in the concentration of NO derivatives was significantly correlated with the change in the AUC(0-30) (r = 0.896, P =.0114) but not with the change in the percentage of maximal fall in FEV(1). CONCLUSION: These findings suggest that excessive production of NO is associated with EIB in patients with asthma and contributes to the prolonged airway narrowing phase rather than to the maximal airway narrowing evoked by exercise.

Adult↗
PubMed2000

Monoclonal antibodies recognizing surface residues of the beta subunit of Escherichia coli F(1) ATPase: functional importance of the epitope residues.

Two monoclonal antibodies, beta 208 and beta 210, against the beta subunit of the F(1) ATPase from Escherichia coli reacted with an intact beta subunit and also a peptide corresponding to a portion of beta between residues 1 and 145. Mutations at Ala-1, Val-15, Glu-16, Phe-17, Leu-29, Gly-65, or Leu-66, and His-110 or Arg-111 for beta 210 and beta 208, respectively, caused decreased antibody binding to beta, suggesting that these residues form the epitopes and are thought to lie close together on the surface of the beta subunit. The topological locations of the corresponding residues in the atomic structure of the bovine beta subunit agree well with these expectations, except for Ala-1 and Leu-29. beta 210 binds to two beta strands including the epitope residues that are 50 residues apart, indicating that this antibody recognizes the tertiary structure of the N-terminal end region. Mutations in the epitope residues of beta 210 do not affect the F(1) ATPase activity, suggesting that surfaces of the two beta strands in the amino-terminal end region are not functionally essential. To analyze the functional importance around His-110 recognized by beta 208 we introduced site specific mutations at residues His-110 and Ile-109. Ile-109 to Ala or Arg, and His-110 to Ala or Asp caused defective assembly of F(1). However, the His-110 to Arg mutation had no effect on molecular assembly, suggesting that Ile-109 and His-110, especially the positive charge of His-110 are essential for the assembly of F(1). The His-110 to Arg mutation caused a large decrease in F(1)-ATPase activity, suggesting that a subtle change in the topological arrangement of the positive charge of His-110 located on the surface of beta plays an important role in the catalytic mechanism of the F(1)-ATPase.

Amino Acid Sequence↗
PubMed2000

Clinical investigation of pulmonary Mycobacterium avium complex infection in human T lymphotrophic virus type I carriers.

BACKGROUND: Little is known about pulmonary Mycobacterium avium complex (MAC) infection in human T lymphotrophic virus type I (HTLV-I) carriers. A study was undertaken to investigate and clarify the characteristics of pulmonary MAC infection in these subjects. METHODS: Twenty nine patients with pulmonary MAC infection without any underlying pulmonary disorder were investigated. The clinical features and radiographic appearance of HTLV-I carriers and non-carriers were compared and the bronchoalveolar lavage (BAL) fluid of these 29 patients and eight normal female control subjects was analysed. RESULTS: The prevalence of the HTLV-I carrier state in patients with pulmonary MAC infection was 34.5% (10/29) compared with 16.7% (529/3169) among all patients admitted to our department between 1994 and 1998 (odds ratio (OR) 2.63, 95% confidence interval (CI) 1.21 to 5.68). The HTLV-I carriers were all women and all had clinical symptoms, but they did not show systemic dissemination. Peripheral multifocal bronchiectasis with nodular shadowing was seen frequently on the chest computed tomographic (CT) scans of HTLV-I carriers. The area of the pulmonary lesions was more extensive than in non-carriers (p<0.05). White blood cell (WBC) counts and C reactive protein (CRP) levels on admission were significantly lower in HTLV-I carriers than in non-carriers (WBC: difference (D) = 1565/microl, 95% CI -68.9 to 3198.4/microl; CRP: D = 1.8 mg/dl, 95% CI -0.35 to 3.89 mg/dl). The concentrations of neutrophil elastase (NE) and interleukin (IL)-8 in BAL fluid were significantly higher in HTLV-I carriers than in non-carriers (NE: D = 1342 microg/l, 95% CI 704 to 1980.3 microg/l; IL-8: D = 304.5 pg/ml, 95% CI 89.7 to 519. 4 pg/ml). CONCLUSIONS: Pulmonary MAC infection causes more diffuse and widespread lesions in HTLV-I carriers than in non-carriers.

Aged↗
PubMed2000

Deletion polymorphisms in the angiotensin converting enzyme gene are associated with pulmonary hypertension evoked by exercise challenge in patients with chronic obstructive pulmonary disease.

Angiotensin converting enzyme (ACE) plays an important role in the pathogenesis of pulmonary hypertension. In this study we determined whether the deletion (D)/insertion (I) polymorphism in the ACE gene may be associated with pulmonary hypertension evoked by exercise challenge in patients with chronic obstructive pulmonary disease (COPD). ACE genotypes were determined in 19 patients with COPD. All patients underwent right heart catheterization followed by a constant-load exercise test while breathing room air or oxygen. Subgroups were created of seven patients with the II genotype, six with the ID genotype, and six with the DD genotype who were well-matched with respect to age, blood gas data at rest or after exercise, baseline lung function, results of incremental exercise testing, and hemodynamic data at rest. The mean pulmonary arterial pressure (Ppa) and pulmonary vascular resistance (Rpv) at rest in the three subgrpoups did not differ significantly during breathing of either room air or oxygen. However, the Ppa after exercise challenge in patients with the DD genotype (55.7 +/- 4.9 mm Hg [mean +/- SD]) was significantly higher than in patients with the II genotype (42.6 +/- 7.1 mm Hg, p = 0.008). The Rpv after exercise in patients with the DD genotype was also significantly higher than in patients with the ID and II genotypes. During breathing of oxygen to diminish acute hypoxic pulmonary vasoconstriction, the Ppa in patients with the DD genotype (52.3 +/- 3.1 mm Hg) was higher than in patients with the ID genotype (40.5 +/- 5.9 mm Hg, p = 0.0049) or the II genotype (37.7 +/- 5.9 mm Hg, p = 0.0027). In addition, the Rpv in patients with the DD genotype was higher than in patients with the ID and II genotypes. These results suggest that D-I polymorphism in the ACE gene may be associated with pulmonary hypertension evoked by exercise challenge in patients with COPD. However, the number of patients in this study was very small for a genetic association study, and our results should be examined in larger studies.

Aged↗
PubMed2000

Involvement of Iba1 in membrane ruffling and phagocytosis of macrophages/microglia.

Ionized calcium binding adaptor molecule 1, Iba1, is an EF hand calcium binding protein whose expression is restricted to macrophages/microglia. In this study, Iba1 was shown to colocalize with F-actin in membrane ruffles induced by macrophage colony-stimulating factor and in phagocytic cups formed during zymosan phagocytosis. Expression of mutant Iba1 carrying either N- or C-terminal deletions or carrying a substitution in the calcium binding domain, suppressed the membrane ruffling and the phagocytosis. These results indicate that Iba1 is a key molecule in membrane ruffling and the phagocytosis of macrophages/microglia. Furthermore, Iba1 colocalized with a small GTPase Rac in the membrane ruffles and the phagocytic cups. The Iba1 mutants also suppressed membrane ruffling induced by dominant active Rac1V12, but do not affect microspikes by Cdc42V12 and stress fibers by RhoAV14. These observations suggest that Iba1 is involved in Rac and calcium signaling pathways.

Actins↗
PubMed2000

Luminal acidification of diverse organelles by V-ATPase in animal cells.

Eukaryotic cells contain organelles bounded by a single membrane in the cytoplasm. These organelles have differentiated to carry out various functions in the pathways of endocytosis and exocytosis. Their lumina are acidic, with pH ranging from 4.5 to 6.5. This article describes recent studies on these animal cell organelles focusing on (1) the primary proton pump (vacuolar-type H(+)-ATPase) and (2) the functions of the organelle luminal acidity. We also discuss similarities and differences between vacuolar-type H(+)-ATPase and F-type ATPase. Our own studies and interests are emphasized.

Amino Acid Sequence↗
PubMed2000

Determination of alpha-tocopherol and alpha-tocopherylquinone in rat tissues and plasma by high-performance liquid chromatography with electrochemical detection.

Alpha-tocopherol and alpha-tocopherylquinone in rat tissues and plasma were determined simultaneously by using high-performance liquid chromatography-electrochemical detection (HPLC-ED) with dual electrodes in the series mode. Biological samples were saponified in the presence of a mixture of butylated hydroxytoluene, ascorbic acid, and pyrogallol and then extracted with hexane. The compounds were separated on a C18 column using a mobile phase containing 95% methanol and 0.05 M sodium perchlorate as the supporting electrolyte. After HPLC separation, alpha-tocopherylquinone was first reduced at an upstream electrode at -500 mV Both alpha-tocopherol and the reduction product of alpha-tocopherylquinone were then oxidized downstream at +600 mV. Only the downstream electrode current was monitored for the determination. Linearity of the standard curves was obtained over the range 5-30 pmol for alpha-tocopherol and alpha-tocopherylquinone. Minimum detectable quantities (S/N of 3) were 0.25 pmol for alpha-tocopherol and 0.31 pmol for alpha-tocopherylquinone. The method was applied to analysis of the contents of alpha-tocopherol and alpha-tocopherylquinone in rat tissues and plasma. By hyperoxia, the content of alpha-tocopherol was decreased remarkably in lung, and in contrast, the contents of alpha-tocopherylquinone were increased in all tissues studied with the exception of plasma, though the content of alpha-tocopherylquinone in normal rats is quite small. The technique is particularly useful in the quantitation of the oxidation of alpha-tocopherol in biological samples.

Animals↗
PubMed2000

[Simultaneous determination of theophylline and its metabolites by HPLC].

A high performance liquid chromatography (HPLC) method has been developed for the simultaneous determination of theophylline and its metabolites, with caffeine and its metabolites. The method is simple and applicable to planning the administration of theophylline. We used a column-switching system in combination with atmospheric pressure chemical ionization liquid chromatography-mass spectrometry (LC-APCI-MS) for the determination of theophylline and its metabolites in biological samples. In the mass spectrum, the molecular ions of these drugs and metabolites were clearly observed as base peaks. The method is sufficiently sensitive and accurate for the pharmacokinetic studies of these drugs.

Caffeine↗
PubMed2000

The effect of calcium-sensing receptor gene polymorphisms on serum calcium levels: a familial hypocalciuric hypercalcemia family without mutation in the calcium-sensing receptor gene.

Familial hypocalciuric hypercalcemia (FHH) is a benign syndrome with elevated levels of serum calcium, relative hypocalciuria, and non-suppressed serum parathyroid hormone (PTH) levels. FHH usually occurs by a heterozygous mutation of the calcium sensing receptor (Casr), but some FHH patients show no mutations of the Casr. We encountered a unique FHH family in which the proband and her mother had many calcium deposits on their skin. The proband was medicated with Levothyroxine for hypothyroidism due to an iodine transport defect (ITD). We searched for mutation of the Casr, but found none. The only change distinguishing the proband and her mother from her father was at codon 990, reported to be a polymorphic site. We investigated the frequency of polymorphism at codon 990, but a significant relationship between the three genotypes and the serum calcium concentration was excluded. At the other polymorphic sites at codon 536, 926, 986, and 1011, polymorphisms were rare in Japanese, and we could not confirm a significant relationship. In conclusion, mutation in the Casr gene alone does not explain all cases of FHH. The other mechanisms should be identified.

Base Sequence↗
PubMed1999

Targeted disruption of the gene encoding the proteolipid subunit of mouse vacuolar H(+)-ATPase leads to early embryonic lethality.

Vacuolar H(+)-ATPase (V-ATPase) is responsible for acidification of intracellular compartments in eukaryotic cells. Its 16-kDa subunit (proteolipid, PL16) plays a central role in V-ATPase function, forming the principal channel via which protons are translocated. To elucidate physiological roles of V-ATPase in mammalian cell function and embryogenesis, we attempted to generate a PL16 null mutant mouse by gene-targeting. Mice heterozygous (PL16(+/-)) for the proteolipid mutation were intercrossed and their offspring were classified according to genotype. There were no homozygous (PL16(-/-)) pups among 69 neonates examined, but a few PL16(-/-) embryos were found during the pre-implantation stages of embryonic development, up to day 3.5 post-coitum. These results suggested that PL16 (and hence V-ATPase) may play an essential role in cell proliferation and viability during early embryogenesis. PL16(+/-) mice were indistinguishable from their wild-type littermates and displayed no discernible abnormalities, although the PL16 mRNA level in PL16(+/-) mice decreased to about one-half of wild-type levels.

Animals↗
PubMed1999

Expression of functional Na+/H+ antiporters of Helicobacter pylori in antiporter-deficient Escherichia coli mutants.

An open reading frame with a sequence homologous to Escherichia coli Na+/H+ antiporter A (ENhaA) was found in the total genomic sequence of Helicobacter pylori, a pathogenic bacterium of gastric inflammation, and was named HNhaA. The primary sequences and the hydropathy profiles of ENhaA and HNhaA were very homologous except for one additional region found in HNhaA. This sequence has about 40 hydrophilic amino acid residues inserted at the position next to residue 235 of ENhaA which corresponds to residue 245 of HNhaA. HNhaA was expressed in E. coli mutants deficient in Na+/H+ antiporters and complemented the salt-sensitive phenotype of the mutants. Membrane vesicles prepared from these transformants of HNhaA using mutants deficient in the antiporters had the antiporter activities. Surprisingly, the antiporter activity in the transformant membranes was high at acidic and neutral pH, while ENhaA did not function at these pHs. A hydrophilic region around residue 235 in ENhaA and the additional hydrophilic region of about 40 residues in the same region found in HNhaA might be responsible for this difference in activity by acting as putative pH sensors.

Amino Acid Sequence↗
PubMed1999

Chiral separation and quantitation of pentazocine enantiomers in pharmaceuticals by capillary zone electrophoresis using maltodextrins.

The chiral separation of pentazocine was achieved by capillary electrophoresis using oligosaccharides. Enantiomers were separated on 100 mM Tris/H3PO4 buffer (pH 2.5) with 5% maltodextrin as a chiral selector, and migration behavior was monitored at 200 nm. Under these conditions, (-)- and (+)-pentazocine and dextromethorphan (internal standard) migrated within 9 min, and the resolution of pentazocine enantiomers was 2.54. Linear calibration curves were obtained in the range 5-50 microg/ml(-1) for each enantiomer. The detection limit of pentazocine enantiomers was 29 pg, and the recoveries of(-)- and (+)-pentazocine were 98.9 (R.S.D., 3.4%) and 101.4% (R.S.D., 4.3%) with 10 microg/ml(-1), respectively.

Analgesics, Opioid↗
PubMed1999

Three-dimensional appearance of Bowman's layer after radial keratotomy.

PURPOSE: To examine the 3-dimensional collagen fibrillar architecture of Bowman's layer after radial keratotomy (RK). SETTING: Department of Ophthalmology, Niigata University School of Medicine, Niigata, Japan. METHODS: This study used monkey eyes in which 0.3 mm deep radial incisions were made on the cornea 2 weeks and 1, 6, and 12 months before the animals were killed. Corneal buttons were immersed in a fixative and the cells macerated with sodium hydroxide 10%. Scanning electron microscopy (SEM) was performed according to standard procedures. A part of the specimens was embedded in epoxy resin for light microscopic (LM) observation for comparison. RESULTS: The 3-dimensional collagen fibrillar architecture of Bowman's layer was revealed by SEM. The rupture of Bowman's layer could be seen 12 months after surgery and there was no continuity of collagen fibrils in the ruptured area. In LM observations, the width of the stromal incisions gradually became narrower near 12 months after surgery. CONCLUSION: Our cell-maceration/SEM method showed that the rupture of Bowman's layer remained up to 12 months after RK. This suggests that discontinuity of Bowman's layer may be responsible for globe rupture after RK.

Animals↗
PubMed1999

Administration of SIN-1 induces guinea pig airway hyperresponsiveness through inactivation of airway neutral endopeptidase.

BACKGROUND: Peroxynitrite plays an important role in the pathogenesis of airway inflammation. We have already found that peroxynitrite may contribute to decreased beta(2)-adrenoceptor responses in airway smooth muscle. However, it is not known whether peroxynitrite can alter neutral endopeptidase (EC 3.4.24.11; NEP) activity in the airways. This study was designed to determine whether peroxynitrite induces airway hyperresponsiveness to substance P (SP) and endothelin-1 (ET-1) through the inactivation of airway NEP. METHODS: We examined whether the administration of S-morpholinosydnonimine (SIN-1), a compound that releases peroxynitrite, increased bronchoconstrictor responses to SP and ET-1 in anesthetized guinea pigs. In addition, we assayed NEP activity in the airways of SIN-1-exposed guinea pigs. RESULTS: Though SIN-1 (10(-7) M) alone had no effect on pulmonary resistance, pretreatment with SIN-1 significantly enhanced SP- and ET-1-induced bronchoconstriction. Pretreatment with phosphoramidon, an NEP inhibitor, also enhanced SP- and ET-1-induced bronchoconstriction. However, simultaneous administration of phosphoramidon and SIN-1 had no additive effect on SP- and ET-1-induced bronchoconstriction. Peroxynitrite formation by SIN-1 was completely inhibited by N-acetylcysteine (NAC) and glutathione (GSH) in vitro, and pretreatment with NAC and GSH significantly reversed the potentiation by SIN-1 of SP-induced bronchoconstriction. In addition, the NEP activity of the trachea after SIN-1 exposure was significantly reduced compared to the level in control guinea pigs (solvent for SIN-1: 30.0+/-4.2 fmol.min(-1).mg tissue(-1); 10(-7) M SIN-1; 15.5+/-4.5 fmol.min(-1).mg tissue(-1), p<0.05). CONCLUSIONS: These findings suggest that peroxynitrite induces airway hyperresponsiveness to SP and ET-1 through the inactivation of airway NEP, and that peroxynitrite is an important mediator of the alterations in airway functions.

Animals↗