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Biomedical subjects

H Hayashi

Publications and source records attributed to H Hayashi.

At least 667 records · Page 37Linked to original sources
PubMed1996

Prevention of reoxygenation-induced arrhythmias in guinea pig papillary muscles.

Effects of various agents on reoxygenation-induced arrhythmias, action potentials, and tension of guinea pig papillary muscles were recorded to investigate the site of action. Triggered activities due to delayed afterdepolarizations (DADs) and aftercontractions were elicited on reoxygenation after 60-min substrate-free hypoxia. Low extracellular Ca2+ (0.1 mM) abolished arrhythmias, and high Ca2+ (4.9 mM) increased the amplitudes of DADs and aftercontractions. D-600 at the high concentration (20 microM) decreased the incidence of arrhythmias (p < 0.05 vs. no drug) and decreased the recovery of developed tension after reoxygenation (p < 0.001). Ryanodine (1 microM) abolished aftercontractions and arrhythmias but did not affect the recovery of developed tension. Tetrodotoxin (TTX 3 microM) and nicorandil (100 microM) decreased the incidence of arrhythmias (p < 0.05), but did not affect the recovery of developed tension or the amplitudes of aftercontractions. TTX caused only a slight decrease in Ca2+ transients in a fluo-3-loaded guinea pig ventricular myocyte. The Ca2+ entry through the Ca2+ channels apparently synchronized Ca2+ release from the sarcoplasmic reticulum, and D-600 at the high concentration apparently decreased the incidence of arrhythmias. TTX and nicorandil decreased arrhythmias, probably by decreasing the Na+ current or by increasing the ATP-sensitive K+ current, respectively.

Action Potentials↗
PubMed1996

Characterization of a toxin-deficient Clostridium perfringens strain, KZ1340.

Clostridium perfringens KZ1340, previously classified as Clostridium plagarum, is an isolate from Antarctic soil, and was identified as an alpha-, theta-, and kappa-toxin non-producing variant. On Southern hybridization, the variant was found to be defective in the pfoA (theta-toxin) gene, but the plc (alpha-toxin) and colA (kappa-toxin) genes were present on the same EcoRI fragment as in the standard strain, NCTC8237. Northern analysis revealed that mature plc mRNA was transcribed in KZ1340 though less efficiently than in NCTC8237, while no mature colA mRNA was present in KZ1340. After transformation of the pfoA and plc genes into the KZ1340 via shuttle vector, pJIR418, the pfoA gene was successfully expressed but the plc gene was not efficiently expressed, suggesting that in KZ1340 there is negative regulation of plc gene expression. Toxin-deficient C. perfringens KZ1340 might be a suitable host for expression analysis of the pfoA gene and other clostridial virulence genes, if expressed efficiently, because it produces a small amount of extracellular toxins.

Bacterial Toxins↗
PubMed1996

Allergenic epitopes of ovalbumin (OVA) in patients with hen's egg allergy: inhibition of basophil histamine release by haptenic ovalbumin peptide.

We studied allergenic determinants that induce hypersensitivity to OVA, the major allergen in egg allergy, using immunoblot and histamine release assays. Immunoblot analysis demonstrated a part of the OVA epitope was in the C-terminal region comprising residues 347-385 (OVA347-385). Histamine was released from basophils of a patient with egg allergy upon stimulation with the OVA fragment corresponding to OVA347-385. Furthermore, detailed epitope mapping using overlapping peptides (residues 347-366, OVA-A; residues 357-376, OVA-B; and residues 367-385, OVA-C) in the OVA 347-385 region was carried out using the histamine release assay. In order for histamine release from basophils to occur, the allergen must possess two or more allergenic determinants located on the protein molecule at distances that would be equivalent to the distances between IgE molecules on the membrane surface. these results suggest that there are at least two epitopes that bind IgE antibodies on each OVA peptide. In addition, one epitope that binds IgE antibodies in two patients appears to reside in the haptenic peptide OVA357-366 (OVA-B1). The histamine release from basophils stimulated by OVA-B was completely inhibited by OVA-B1 in one of these patients. Similarly, OVA-B1 inhibited the histamine release produced by OVA-A in the other by more than 40%. These results suggest that haptenic synthetic peptides could regulate the allergic reaction in the effector phase if common epitope(s) recognized by IgE antibodies in the patients with egg allergy can be found. These are the first studies that provide an antigen-specific approach to inhibiting histamine release from basophils by a haptenic peptide recognized by IgE antibodies in an allergic disorder.

Allergens↗
PubMed1996

Structure-activity studies on C-terminal hirudin peptides containing sulfated tyrosine residues.

To clarify the role of the negative charge of the C-terminal region of hirudin, we chemically synthesized the C-terminal peptide of hirudin variant-1 (HV-1), HV-1-(54-65), and its analogs, [E61Y,E62Y]HV-1-(54-65) and [E62Y]HV-1-(54-65), and then sulfated the Tyr residue(s) in these peptides by both enzymic and chemical methods. Enzymic O-sulfation of Tyr residues in the peptides by use of sulfotransferase isolated from Eubacterium A-44 allowed us to produce four kinds of the sulfated peptide, whose C-terminal sequences were -PEY(SO3H)YLQ, -PYY(SO3H)YLQ, -PYYY(SO3H)LQ and -PYY(SO3H)Y(SO3H)LQ. On the other hand, all Tyr residues in the peptides were successfully sulfated by chemical reaction with N,N'-dicyclohexylcarbodiimide in the presence of sulfuric acid. Based on the analysis of structure-activity relationships of these sulfated peptides for thrombin inhibition, the Tyr62 and Tyr63 bisulfated peptide GDFEEIPEY(SO3H)Y(SO3H)LQ was found to be the most potent inhibitor of thrombin among the products tested. No increase in potency was observed by further substitution of Glu61 with Tyr(SO3H). The inhibitory activity by substitution with Tyr(SO3H) at position 63 was greater than that obtained by the substitution at position 62.

Amino Acid Sequence↗
PubMed1996

Biological and antigenic characterization of three BApNA-hydrolyzing proteases from the culture supernatant of Porphyromonas gingivalis.

Biological and antigenic distinction of 3-N-alpha-benzoyl-DL-arginine p-nitroanilide (BApNA)-hydrolyzing proteases (Pase-B, Pase-C and Pase-S) isolated from the culture supernatant of Porphyromonas gingivalis were determined. Immunoblotting analysis of these enzymes using a polyclonal antibody against Pase-S, which is a soluble, clostripain-like protease, revealed immunological distinction from Pase-C, a vesicle-associated thiol-protease. Pase-B, a vesicle-associated clostripain-like protease, reacted with the antibody and was also found to contain a considerable amount of carbohydrates in its structure, as compared with the others. Analysis of N-terminal amino acids of Pase-B provided a sequence not found in the SwissProt data bank or previously reported as N-terminal sequences of proteases from P. gingivalis. Pase-S, resembling Pase-B in its hydrolytic specificity, cleaved only arginine residues of peptides and degraded type IV and denatured type I collagen. Pase-C hydrolyzed N-alpha-benzoyl-DL-lysine p-nitroanilide and showed the strongest capacity of degrading native type I collagen. This enzyme was also the only one to possess hemagglutinating activity. Our findings suggest that Pase-S from P. gingivalis is less active than Pase-C and that the enzyme may be an isozyme of Pase-B.

Affinity Labels↗
PubMed1996

Mammary hamartoma: immunohistochemical study of two adenolipomas and one variant with cartilage, smooth muscle and myoepithelial proliferation.

Three cases of mammary hamartoma were investigated immunohistochemically and are described. Case 1 was a 42 year old woman with an elastic hard tumor, 1.5 cm in diameter, in her left breast. Case 2 was a 49 year old woman with a semisoft tumor, 5 x 2 cm, in her right breast. Case 3 was a 47 year old woman with a hard tumor, 5 cm in diameter, in her left breast. In each case, mammography and ultrasonography revealed a benign-looking, well-circumscribed mass without calcification. Histologically, the tumors were composed of adipose tissue, mammary glands, and fibrous and/or fibromuscular tissue. The tumor in case 3 also contained small islands of hyaline cartilage. Immunohistochemical analysis was performed, and epithelial and mesenchymal components were discretely and differentially immunostained except that the smooth muscle component seemed to be derived from myoepithelial cells. Cartilage formation might be the result of metaplasia, and 'metaplastic variant of the mammary hamartoma' or 'choristoma' may be an appropriate term for cartilage-containing mammary hamartoma. Using proliferating cell nuclear antigen (PCNA)-immunostaining, we observed that each component of the tumors had an individual growth rate. This finding may reflect one aspect of the biological characteristics of hamartoma.

Adenoma↗
PubMed1996

The virR/virS locus regulates the transcription of genes encoding extracellular toxin production in Clostridium perfringens.

Extracellular toxin production in Clostridium perfringens is positively regulated by the two-component regulatory genes virR and virS. Northern (RNA) blots carried out with RNA preparations from the wild-type strain 13 and the isogenic virR and virS mutants TS133 and JIR4000 showed that the virR and virS genes composed an operon and were transcribed as a single 2.1-kb mRNA molecule. Primer extension analysis led to the identification of two promoters upstream of virR. Hybridization analysis of the mutants and their complemented derivatives showed that the virR/virS system positively regulated the production of alpha-toxin (or phospholipase C, theta-toxin (perfringolysin O), and kappa-toxin (collagenase) at the transcriptional level. However, the modes of regulation of these genes were shown to differ. The theta-toxin structural gene, pfoA, had both a major and a very minor promoter, with the major promoter being virR/virS dependent. The colA gene, which encodes the kappa-toxin, had two major promoters, only one of which was virR/virS-dependent. In contrast, the alpha-toxin structural gene, p1c, had only one promoter, which was shown to be partially regulated by the virR and virS genes. Comparative analysis of the virR/virS-dependent promoters did not reveal any common sequence motifs that could represent VirR-binding sites. It was concluded that either the virR/virS system modulates its effects via secondary regulatory genes that are specific for each toxin structural gene or the VirR protein does not have a single consensus binding sequence.

Bacterial Proteins↗
PubMed1996

Transient giant negative T wave in acute anterior myocardial infarction predicts R wave recovery and preservation of left ventricular function.

OBJECTIVE: To investigate the value of a giant negative T wave (> or = 1.0 mV) in precordial leads of 12-lead electrocardiograms in the acute phase of Q wave myocardial infarction as a predictor of myocardial salvage. METHODS: Coronary angiographic and electrocardiographic findings, left ventricular ejection fraction in the chronic stage, and levels of cardiac enzymes were compared in patients with myocardial infarction with (group GNT, n = 31) and without (group N, n = 20) a giant negative T wave. GNT patients were divided into two subgroups according to the presence (GNT:R[+], n = 10) or absence (GNT: R[-], n = 21) of R wave recovery with an amplitude > or = 0.1 mV in at least one lead that had shown Q waves. RESULTS: The maximum level of creatine kinase and the total creatine kinase were lower in group GNT compared with group N (P < 0.05). The left ventricular ejection fraction was higher in group GNT than in group N (P < 0.05). The maximum creatine kinase and total creatine kinase were lower in GNT:R(+) than in GNT:R(-) (P < 0.01). The left ventricular ejection fraction was higher in GNT:R(+) than in GNT:R(-) (P < 0.01). The frequency of R wave recovery was significantly higher when giant negative T waves appeared within 100 h of myocardial infarction or when the maximum potential was > or = 1.4 mV. The appearance of a giant negative T wave > or = 1.4 mV had a sensitivity of 90%, a specificity of 71.4%, a diagnostic accuracy of 77.4%, a positive predictive value of 60%, and a negative predictive value of 93.8% for prediction of R wave recovery. CONCLUSIONS: The appearance of a giant negative T wave, especially within 100 h of the onset of myocardial infarction, with a maximum potential of > or = 1.4 mV, may predict a reappearance of the R wave and a better left ventricular function in patients in the chronic stage of anterior myocardial infarction.

Cardiac Catheterization↗
PubMed1996

Stapedial reflex in amyotrophic lateral sclerosis.

OBJECTIVE: To examine mechanisms controlling the stapedial reflex in patients with amyotrophic sclerosis (ALS). METHODS: The stapedial reflex was examined using impedance audiometry in 38 patients with sporadic ALS and in 25 age matched controls. RESULTS: All patients showed normal reflex decay test results. There were no significant differences between patients with ALS and control subjects in reflex threshold, latency, amplitude, or contraction time (C50). Although each reflex variable in the patients with classic or progressive muscular atrophy types of ALS showed no significant difference from that in control subjects, the patients with bulbar type ALS showed significantly longer latency, C50, and retraction time (D50), and significantly lower amplitude than control subjects. Three types of abnormal reflex waveforms (polyphasic, abnormally delayed retraction, and abnormally early retraction) were noted in six patients. CONCLUSION: The subclinical involvement of the stapedius motor neurons or of the supranuclear stapedius motor system might be responsible for the abnormalities of the stapedial reflex in ALS.

Acoustic Impedance Tests↗
PubMed1996

Effects of Mg2+ on Ca2+ waves and Ca2+ transients of rat ventricular myocytes.

It has been shown that the occurrence of the transient inward current, which is responsible for triggered activity, was often associated with propagating regions of increased intracellular Ca2+ concentration ([Ca2+]i), i.e., the "Ca2+ wave." To investigate the mechanism of antiarrhythmic action of Mg2+, we have studied effects of high concentrations of Mg2+ on Ca2+ waves in isolated rat ventricular myocytes. [Ca2+]i was estimated using the Ca(2+)-indicating probe indo 1. Ca2+ waves in myocytes, stimulated at 0.2 Hz, were induced by perfusion of isoproterenol (10(-7) M). High Mg2+ concentration suppressed Ca2+ waves in a concentration-dependent manner (36% at 4 mM, 70% at 8 mM, and 82% at 12 mM). The Ca2+ channel blocker verapamil also suppressed Ca2+ waves in a similar way. In contrast with marked depression of Ca2+ transients by verapamil, Ca2+ transients were not affected by high Mg2+ concentration (8 mM). High Mg2+ concentration also reduced frequencies of Ca2+ waves in the absence of electrical stimulation, whereas verapamil failed to reduce frequencies of Ca2+ waves. Reduction in frequency of Ca2+ waves by high Mg2+ concentration was associated with slowing of propagation velocity of Ca2+ waves. To examine whether suppressive effects of high Mg2+ concentration on Ca2+ waves were related to an increase in intracellular Mg2+ concentration ([Mg2+]i), the effect of high-Mg2+ solution on [Mg2+]i was examined in myocytes loaded with mag-fura 2. An increase in extracellular Mg2+ concentration from 1 to 12 mM increased [Mg2+]i from 1.06 +/- 0.16 to 1.87 +/- 0.22 mM (P < 0.01) in 30 min. To examine the effect of high Mg2+ concentration on amount of releasable Ca2+ in the sarcoplasmic reticulum, the effect of high Mg2+ concentration on the Ca2+ transient induced by a rapid application of caffeine was examined. High-Mg2+ solution increased the peak of the caffeine-induced Ca2+ transient. These results suggest that the inhibitory effect of Mg2+ on Ca2+ waves was not due to inhibition of the sarcolemmal Ca2+ channel but could be due to a decreased propensity for the sarcoplasmic reticulum to divest itself of excess Ca2+.

Animals↗
PubMed1996

Angiogenin levels in the vitreous from patients with proliferative diabetic retinopathy.

The vitreous levels of angiogenin, which is a potent blood vessel-inducing protein, were measured to determine their association with proliferative diabetic retinopathy (PDR). Undiluted vitreous fluid specimens were collected from 30 eyes with PDR at the time of vitrectomy. A sandwich enzyme-liked immunosorbent assay was then used to quantitate the levels of angiogenin. As a control we determined the levels in the specimens from 21 patients with proliferative vitreoretinopathy (PVR) and 4 patients with idiopathic macular epiretinal membrane (IERM). The average angiogenin level in the eyes with PDR was 43.7 ng/ml, and no significant difference was observed among PDR, PVR and their reoperation cases. In the category of IERM, the mean concentration of angiogenin was 2.1 ng/ml, which was significantly lower than that of the PDR and PVR cases. Our study thus demonstrated a significant increase in the vitreous angiogenin levels in eyes with PDR, PVR and those undergoing reoperation for these conditions in comparison to eyes with IERM. We therefore postulated that the elevated angiogenin levels thus reflected a breakdown of the blood-ocular barrier in eyes with PDR and PVR.

Adult↗
PubMed1996

Molecular cloning and functional expression of cDNAs for Glycyrrhiza glabra squalene synthase.

We have isolated two cDNAs (GgSQS1 and GgSQS2) encoding squalene synthase of Glycyrrhiza glabra L. by cross-hybridization with that of Arabidopsis thaliana squalene synthase under conditions of low stringency. Their nucleotide sequences contained an open reading frame for a polypeptide of 413 amino acids (GgSQS1) and 412 amino acids (GgSQS2), respectively. The deduced amino acid sequence of GgSQS1 exhibits 88%, 81%, 78%, 45-44%, and 45-41% identity to those of the GgSQS2, Nicotiana benthamiana. Arabidopsis thaliana, mammal, and yeast squalene synthases, respectively. To express the G. glabra enzymes in Escherichia coli, the entire coding region was subcloned into an expression vector. The cell-free extracts of E.coli transformed with the respective plasmid converted 3H-farnesyl diphosphate into squalene.

Amino Acid Sequence↗
PubMed1996

FR901724, a novel anti-human immunodeficiency virus (HIV) peptide produced by Streptomyces, shows synergistic antiviral activities with HIV protease inhibitor and 2',3'-dideoxynucleosides.

A novel tricyclic 21-amino-acid peptide, FR901724, was isolated from the cultured broth of Streptomyces sp. No. 73264. This peptide appears to possess potent anti-human immunodeficiency virus (HIV) activity in vitro and might represent a lead to a new class of anti-HIV agents; it qualifies as an HIV-cell fusion inhibitor because of its weak inhibition of virus-cell binding and strong inhibition of syncytium formation. From the time-of-addition experiments, the mode of action of FR901724 was found to definitely differ from that of KNI-272, a peptide mimetic allophenylnorstatine-derivative HIV protease inhibitor. FR901724 appears to interact with a stage of the virus replicative cycle that may well correspond to virus-cell fusion. We also found that FR901724 was synergistic or had a strong tendency toward synergism when combined with other antiviral drugs, such as KNI-272, AZT, ddI and dextran sulfate.

Amino Acid Sequence↗
PubMed1996

Anticoagulant peptides; synthesis, stability and antithrombin activity of hirudin C-terminal-related peptides and their disulfated analog.

We designed a unique anticoagulant decapeptide, which possesses two O-sulfated tyrosine residues, based on the structure of hirudin's C-terminal functional domain. We first prepared a series of octa-, nona- and decapeptides with no sulfation, Suc-Phe-Glu-Pro-Ile-Pro-Glu-Tyr-Tyr-X-OH [X = bond, Leu or Leu-Gln], by a solution phase method and measured their thrombin times (TT) using human thrombin and rabbit plasma. The shortest octapeptide (3a) showed full antithrombin activity comparable to that of the lead compound hirudin (54-65), and the longest decapeptide (3c) prolonged TT most potently with an IC50 value of 5.8 microM. We consequently converted 3c to a disulfated decapeptide (NF-22) with SO3.pyridine complex and compared its antithrombin activity with that of known hirudin-related peptides: hirugen, MDL28050 and hirulog-1. NF-22 showed potent antithrombin activity with an IC50 value of 0.3 microM, being more potent than hirugen and MDL28050 (IC50 values of 4.0 microM and 1.1 microM, respectively). NF-22 was as potent as hirulog-1. NF-22 showed no change in activity in aqueous solution for 10 d at 60 degrees C, and remained about 90% unchanged in rat plasma on incubation for 24h at 37 degrees C, whereas the corresponding unsulfated peptide (3c) was completely digested under the same condition. NF-22 appears to be one of the most potent and stable peptide anticoagulants among the hirudin analogs.

Amino Acid Sequence↗
PubMed1996

Transmural potential changes associated with the in vitro absorption of theanine in the guinea pig intestine.

Theanine, L-N-ethylglutamine, is one of the major components of amino acids in Japanese green tea. To characterize the mode for intestinal absorption of theanine, the ionic dependency and kinetic properties of the theanine- and glutamine-evoked transmural electrical potential difference changes (delta PD) were investigated in vitro by using everted sacs prepared from the guinea pig ileum. Both theanine and glutamine applied to the luminal side induced dose-dependent increases in delta PD (increase in serosal positive value). The theanine- and glutamine-evoked delta PD values conformed to the Michaelis-Menten relationship, with delta PDmax not being different, whereas the half-saturation concentration was lower for glutamine (3.1 +/- 0.2 mM) than for theanine (21.4 +/- 0.6 mM). The theanine-evoked delta PD value was much smaller when theanine was applied in the presence of glutamine than when applied alone. The theanine- and glutamine-evoked delta PD values were both inhibited by removing Na+ from the luminal solution. These results suggest that the intestinal absorption of theanine and glutamine is mediated by a common Na(+)-coupled co-transporter in the brush-border membrane, the affinity of which is lower for theanine than for glutamine.

Animals↗
PubMed1996

Structures and activity of cellulase inhibitors enzymatically synthesized from cellooligosaccharides and 1-deoxynojirimycin.

Cellulase inhibitors were synthesized from cellooligosaccharides and 1-deoxynojirimycin (DNJ) by transglycosylation, using a commercial cellulase. The structures of these cellulase inhibitors were proved to be 4-O-beta-cellobiosyl-DNJ, 4-O-beta-D-glucopyranosyl-DNJ and 6-O-beta-cellobiosyl-DNJ by the results of 13C- and 1H-NMR analyses. The inhibitory activity of each inhibitor was investigated against several carboxymethylcellulases.

1-Deoxynojirimycin↗