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Biomedical subjects

M Hatano

Publications and source records attributed to M Hatano.

At least 163 records · Page 9Linked to original sources
PubMed1989

Effects of Ca2+ and Zn2+ on trifluoperazine-S100 proteins interactions: induced circular dichroism and fluorescence spectra.

Interactions of trifluoperazine (TFP) with S100 proteins, EF-hand type Ca2+-binding proteins, in the presence of Ca2+ and Zn2+ were studied with induced circular dichroism (CD) and fluorescence spectra. The positive CD bands of TFP were induced at around 265 nm by adding either S100a or S100a0 protein in the presence of Ca2+. No CD band of TFP was, however, induced by adding S100b protein in the presence of Ca2+. Addition of Zn2+ to the TFP/S100 protein solutions did not induce any CD band at all. The fluorescence intensity of 2-p-toluidinylnaphthalene 6-sulfonate (TNS) bound to S100a or S100a0 protein decreased by adding TFP in the presence of Ca2+, while that bound to S100b protein decreased by adding TFP in the presence of Zn2+, indicating that TFP binds to S100a protein and S100a0 protein in a Ca2+-dependent manner and to S100b protein in a Zn2+-dependent manner. From these results together with other experimental findings it was suggested that (1) TFP binds to S100a protein and S100a0 protein in the presence of Ca2+, with half-saturation points of 18 and 3 microM, respectively, (2) TFP binds to S100b protein only in the presence of Zn2+, (3) alpha-subunit of S100 protein binds to TFP specifically in a Ca2+-dependent manner and beta-subunit in a Zn2+-dependent manner.

Animals↗
PubMed1989

Site-directed mutageneses of rat liver cytochrome P-450d: catalytic activities toward benzphetamine and 7-ethoxycoumarin.

Catalytic activities toward benzphetamine and 7-ethoxycoumarin of 11 distal mutants, 9 proximal mutants, and 3 aromatic mutants of rat liver cytochrome P-450d were studied. A distal mutant Thr319Ala was not catalytically active toward benzphetamine, while this mutant retained activity toward 7-ethoxycoumarin. Distal mutants Gly316Glu, Thr319Ala, and Thr322Ala displayed higher activities (kcat/Km) toward 7-ethoxycoumarin that were 2.4-4.7-fold higher than that of the wild-type enzyme. Although kcat/Km values of four multiple distal mutants toward benzphetamine were less than half that of the wild type, activities of these mutants toward 7-ethoxycoumarin were almost the same as or higher than the wild-type activity toward this substrate. The distal double mutant Glu318Asp, Phe325Tyr showed 6-fold higher activity than the wild-type P-450d toward 7-ethoxycoumarin. Activities of the proximal mutants Lys453Glu and Arg455Gly toward both substrates were much lower (less than one-seventh) than the corresponding wild-type activities. Catalytic activities of three aromatic mutants, Phe425Leu, Pro427Leu, and Phe430Leu, toward benzphetamine were less than 7% of that of the wild type, while the activities of these aromatic mutants toward 7-ethoxycoumarin were more than 2.5 times higher than the wild-type activity toward this substrate. From these findings, in conjunction with a molecular model for P-450d, we suggest that (1) the relative importance to catalysis of various distal helix amino acids differs depending on the substrate and that these differences are associated with the size, shape, and flexibility of the substrate and (2) the proximal residue Lys453 appears to play a critical role in the catalytic activity of P-450d, perhaps by participating in forming an intermolecular electron-transfer complex.

Amino Acid Sequence↗
PubMed1989

Mutations at the distal and proximal sites of cytochrome P-450d changed regio-specificity of acetanilide hydroxylations.

Regio-specificities of acetanilide hydroxylations were studied for 11 distal, 9 proximal and 3 aromatic mutants of cytochrome P-450d. Ratios of turnover numbers among these products were remarkably changed depending on the mutants. For example, the ratio of turnover number, para:ortho:meta = 7:0.1:0.3 for the wild type changed to 11:4:3 for a distal mutant, Thr322Ala, or to 13:13:1 for a proximal mutant, Arg455Gly. It was suggested that regio-specificities of microsomal P-450 enzymes are controlled cooperatively by the whole structure of the protein molecules which influences the tertiary structure of the distal environment.

Acetanilides↗
PubMed1989

Bindings of axial ligands to cytochrome P-450d mutants: a difference absorption spectral study.

By site-directed mutagenesis, we made several cytochrome P-450d (P-450d) mutants as follows: Asn310Phe (D13), Ile312Leu (D14), Glu318Asp (D15), Val320Ile (D16), Phe325Thr (D19), Asn310Phe,Ile312Leu (M6), Glu318Asp,Val320Ile (M7), Phe325Thr, Glu318Asp (M3). This region (Asn-310-Phe-325) is supposed to be located in the distal helix above the heme plane in P-450d, being conjectured from the structure of P-450cam. We studied Soret spectral changes of those mutants by adding several axial ligands such as aniline, pyridine, metyrapone, 2-phenylimidazole and 4-phenylimidazole. Binding constants (Kb) of aniline and pyridine to the single and double mutants were higher than those to the wild type by 2-10-times. The double mutations did not additively increase the Kb values compared with those to the single mutants. In contrast, Kb value (1.0.10(5) M-1) of metyrapone to the double mutant M3 was much higher than that (2.0.10(3) M-1) of the wild type and those of the single mutants, D15 (4.5.10(4) M-1) and D19 (1.6.10(4) M-1). The increased affinity of metyrapone to the mutant M3 may be attributed to an interaction of the hydrophobic group of metyrapone with nearby hydrophobic group(s) produced cooperatively by the double mutation of P-450d. Kb values of 2-phenylimidazole and 4-phenylimidazole to the mutant M3 were also the highest among those of the mutants and the wild type. Therefore, it was suggested that this region (from Asn-310 to Phe-325) must be located at the distal region of the heme moiety and form, at least, a substrate-binding region of membrane-bound P-450d.

Amino Acid Sequence↗
PubMed1989

Solvent effect on MCD of Fe(III) heme complexes: magnetic circular dichroism spectra of five-coordinated high-spin iron(III) protoporphyrin-IX-dimethylester in the visible region and their environmental effect. A characterization of the visible electronic transitions in Fe(III) high-spin porphyrins.

Magnetic circular dichroism (MCD) spectra were observed to characterize the nature of the visible bands for high-spin Fe(III) protoheme derivatives with p-nitrothiophenolate, p-nitrophenolate, and methoxy anion as the fifth ligands in several solvents. The visible MCD bands for p-nitrophenolate heme were very sensitive to the solvent polarity, but that those for p-nitrothiophenolate heme and methoxy heme were not dependent on solvent polarity. Thus, both the two visible MCD band positions and the magnitudes were dependent on ET value (a solvent polarity parameter) in the former complex, but not in the latter two complexes. The results are consistent with previously proposed electronic structures of high-spin Fe(III) heme complexes.

Circular Dichroism↗
PubMed1989

An outbreak of gastroenteritis associated with acute rotaviral infection in schoolchildren.

In April 1988 a large outbreak of group C rotavirus infection associated with acute gastroenteritis occurred among schoolchildren and their teachers simultaneously at seven elementary schools in Fukui city, Japan. Of 3,102, 675 (21.8%) became ill. Clinical symptoms were mild, predominantly abdominal pain and vomiting, with diarrhea reported in only 27.6%. The outbreak subsided within 2 d. No pathogenic bacteria were found in fecal specimens; the virus particles detected by electron microscopy were morphologically indistinguishable from conventional infantile rotavirus. Immune electron microscopy showed that these virions formed large aggregates with convalescent serum and with the reference serum specific to group C rotavirus. Polyacrylamide gel electrophoresis showed similar RNA patterns for virus from this outbreak and typical group C rotavirus.

Acute Disease↗
PubMed1989

Specific inhibition of class II MHC gene expression by anti-sense RNA.

We have established an anti-sense RNA system which is capable of regulating expression of the class II (Ia) molecule coded for by the major histocompatibility complex in cultured mouse cells. Various areas of the I-A beta chain gene were subcloned in an anti-sense orientation to the 3' of the dihydrofolate reductase (DHFR) cDNA under the control of the human metallothionein IIa gene promoter. These anti-sense DNA constructs were transfected into M12.4 cells, a BALB/c B lymphoma cell line which expresses both I-A and I-E molecules on the cell surface. I-A expression of selected clones transfected with anti-sense DNA encompassing the 5' untranslated region (UT) (100 or 310 bp) including the translation start site or the poly(A) addition signalling sequence in the 3' UT (250 bp) of the I-A beta chain gene were specifically reduced to less than 5% of the control M12.4 cell surface I-A expression. These clones had normal levels of I-E expression. However, transfection of the anti-sense DNA to the beta 1 domain (510 bp) including the splicing donor and acceptor sequences did not affect the expression of I-A molecules. The same antisense DNA constructs (100 bp of the 5' UT or 250 bp of the 3' UT) without the DHFR cDNA (710 bp) did not down-regulate the expression of I-A molecules, indicating that either the physical length of the anti-sense RNA or specific DHFR cDNA sequences are also important.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Metastatic small cell carcinoma to the pineal body: case report.

A case of metastatic small cell carcinoma to the pineal body is reported and the clinical and radiological features of this rare tumor are discussed. A 66-year-old man presented with progressive dementia, gait disturbance, vertical gaze palsy, and convergence retraction nystagmus. Computed tomographic scan revealed a 2 X 3-cm high-density mass in the pineal region, which showed strong contrast enhancement. The tumor was resected through a right occipital transtentorial approach. The pathological diagnosis of the surgical specimens was small cell carcinoma. A systemic workup for the primary lesion subsequently revealed small cell carcinoma of the lung. Although occurring rarely, metastatic tumor should be considered in the differential diagnosis of pineal tumor in elderly patients.

Aged↗
PubMed1989

Epstein-Barr virus with transforming and early antigen-inducing ability originating from nasopharyngeal carcinoma: mapping of the viral genome.

Epstein-Barr virus (EBV) from a human hybrid epithelial cell line (NPC-KT), derived from the fusion of human adenoidal cells and EBV genome-containing primary nasopharyngeal carcinoma cells (NPC) has both transforming and early antigen (EA)-inducing abilities. EBV DNA from NPC-KT cells was partially digested with TaqI and ligated to the cloning vector pJB8 at the ClaI site. This cosmid library encompassed the whole genomic DNA of the virus except for several kb of the terminal fragments. The identification and location of each of the cloned DNA fragments have been defined by hybridization to blots prepared with B95-8 and NPC-KT virion DNA. Defective heterogeneous restriction enzyme fragments of the viral DNA were not identified in any of the cosmid clones nor detected in hybridizations to virion DNA, which indicates that a single virus population derived from the NPC tissue has both transforming and EA-inducing activities.

Antigens, Viral↗
PubMed1989

Immuno-affinity purification of specific antibodies against vasoactive intestinal polypeptide (VIP) on VIP (1-10)-linked polydimethylacrylamide resin.

A novel method for immuno-affinity purification of specific antibodies against amino-terminal (N-terminal) porcine vasoactive intestinal polypeptide (p-VIP) was developed. The antiserum VP (No. 6203) elicited by p-VIP-immunoglobulin G (IgG) conjugate was heterogenous and reacted not only with the N-terminal fragments of p-VIP but also with carboxy-terminal (C-terminal) fragments. To obtain specific antibodies against the N-terminal fragment of p-VIP, antiserum VP was purified by column chromatography on N-terminal decapeptide (p-VIP 1-10)-linked polydimethylacrylamide resin. The antibody thus obtained was highly specific to the N-terminal sequence of p-VIP and hardly reacted with the C-terminal fragments of VIP in enzyme-linked immunosorbent assay.

Antibodies↗
PubMed1989

Role of the calcium-calmodulin system in the adenylate cyclase activity of vascular smooth muscle.

In order to evaluate the relationship between the calcium-calmodulin system and the adenylate cyclase activity of vascular smooth muscle, we examined the effects of several calcium effectors on the basal and stimulated adenylate cyclase activity. Thoracic aortae were removed from Wistar rats and the tissues were homogenated with cold homogenizing buffer containing 1 nM EDTA. Membrane protein fraction of the smooth muscle was prepared by centrifugation at 37,000 g. In this procedure, endogenous guanine nucleotides and contractile proteins remained. The protein fraction was incubated with 2 mM EGTA, 50 microM trifluoperazine, 0.1 microM A23187 or 25 microM calmodulin under basal and stimulated (50 microM isoproterenol, 100 microM GTP and 50 microM forskolin) conditions. The adenylate cyclase activity was determined by a method modified in our laboratory using double isotope counting. Trifluoperazine reduced the basal adenylate cyclase activity significantly (p less than 0.01) as well as the stimulated enzyme activities. A23187 did not affect the basal enzyme activity, but elevated the isoproterenol stimulated enzyme activity significantly (p less than 0.05). EGTA did not affect the basal and stimulated adenylate cyclase activities. Calmodulin elevated the basal enzyme activity significantly (p less than 0.02), but did not affect the stimulated enzyme activities. These results suggest that the calcium-calmodulin system is necessary for maintenance of the adenylate cyclase activity of vascular smooth muscle cells. The calmodulin acting site is considered to be the catalytic subunit, and stimulation of the enzyme is accelerated by calcium ion.

Adenylyl Cyclases↗
PubMed1989

[Silent myocardial ischemia evaluated by ambulatory left ventricular function monitoring].

To determine whether left ventricular function is less disturbed in asymptomatic ischemia than in symptomatic ischemia, exercise-induced left ventricular function was measured in beat-to-beat using an ambulatory left ventricular function monitoring system. The study subjects were 22 patients with coronary artery disease. Supine and sitting ergometer exercise tests were performed. Of 44 exercise tests, 33 showed significant ST depression on electrocardiograms. Among these 33, 17 were asymptomatic: 16, symptomatic. The left ventricular functions during exercise of these 33 were analyzed and compared with those of five normal controls. The changes in end-diastolic volume were not so significant either in controls or in the asymptomatic and symptomatic groups, but the changes in end-systolic volume were reversed in the diseased groups, particularly in the symptomatic group. The changes in ejection fraction (EF), therefore, were significantly negative in the diseased group, particularly in the symptomatic group. The correlation between exercise-induced left ventricular dysfunction and symptoms was evaluated among the 33 patients. Symptoms were present in 35% (6/17) in less than 10% decrease in EF, 44% (4/9) in 10-15% decrease, and 85% (6/7) in greater than or equal to 15% decrease, respectively. Thus, asymptomatic ischemia represents a lesser degree of myocardial ischemia as indicated by mild left ventricular dysfunction, compared with symptomatic ischemia. However, some cases of severe myocardial ischemia did not develop symptoms. We concluded that silent myocardial ischemia is responsible for a lesser degree of myocardial ischemia and decreased pain perception.

Electrocardiography, Ambulatory↗
PubMed1989

[Studies on the new clinical marker of diabetes mellitus (5). Seasonal variation of insulin specific binding to erythrocyte insulin receptor].

Specific binding of insulin to receptors of healthy human erythrocytes was slightly increased in December (6.9 +/- 1.7%) compared with that in May (5.1 +/- 1.4%). Random screening of insulin specific binding to erythrocyte insulin receptors of various patients (including not only diabetes mellitus but also other diseases) suggested that insulin specific binding is slightly higher in winter than in summer. Insulin specific bindings of healthy subjects in Sapporo (January, -6.1 degrees C), Saitama (5.1 degrees C) and Okinawa (16.0 degrees C) were 9.1 +/- 1.6%, 6.9 +/- 1.7%, 6.2 +/- 1.7%, respectively. A week negative correlation was observed between atmospheric temperature and insulin specific binding. These results suggest that insulin specific binding to the erythrocyte insulin receptor is affected by not only the pathological situation of the patient, but also season and atmospheric temperature.

Diabetes Mellitus↗
PubMed1989

Monoclonal anti-human C3d antibodies: stabilization of the alternative pathway C3 convertase.

IgG mouse monoclonal antibody (mAb) was prepared by fusion of spleen cells from mice immunized with human C3d (mAb:C3d) using syngeneic thymocytes as feeder cells. mAb:C3d was assessed for its effect on the stabilization of the cell-bound alternative pathway C3 convertase EAC3bBb. It bound to cell-bound C3b and stabilized C3bBb at 30 degrees in the presence of EDTA-GVB. The plasma protein H reduced the stabilization effect of the stabilized C3 convertase. These results suggest that binding of antibody to C3d may stabilize C3bBb. It seems likely that such antibody induces in C3b conformational change, which increases the C3bBb complex stability.

Antibodies, Monoclonal↗