Biomedical subjects

S Yamaguchi

Publications and source records attributed to S Yamaguchi.

At least 289 records · Page 16Linked to original sources

PubMed1999

Myrothecium verrucaria bilirubin oxidase and its mutants for potential copper ligands.

Bilirubin oxidase (EC:1.3.3.5) purified from a culture medium of Myrothecium verrucaria MT-1 (authentic enzyme) catalyzes the oxidation of bilirubin to biliverdin in vitro and recombinant enzyme (wild type) was obtained by using an overexpression system of the bilirubin oxidase gene with Aspergillus oryzae harboring an expression vector. The absorption and ESR spectra showed that both bilirubin oxidases are multicopper oxidases containing type 1, type 2, and type 3 coppers similar to laccase, ascorbate oxidase, and ceruloplasmin. Site-directed mutagenesis has been performed for the possible ligands of each type of copper. In some mutants, Cys457 --> Val, Ala, His94 --> Val, and His134.136 --> Val, type 1 and type 2 copper centers were perturbed completely and the enzyme activity was completely lost. Differing from the holoenzyme, these mutants showed type 3 copper signals. However, the optical and magnetic properties characteristic of type 1 copper were retained even by mutating one of the type 1 copper ligands, i.e., a mutant, Met467 --> Gly, showed a weak but apparent enzyme activity. A double mutant His456.458 --> Val had only type 1 Cu, showing a blue band at 600 nm (epsilon = 1.6 x 10(3)) and an ESR signal with very narrow hyperfine splitting (A parallel = 7.2 x 10(-)3 cm-1). Since the type 2 and type 3 coppers are not present, the mutant did not show enzyme activity. These results strongly imply that the peculiar sequence in bilirubin oxidase, His456-Cys457-His458, forms an intramolecular electron-transfer pathway between the type 1 copper site and the trinuclear center composed of the type 2 and type 3 copper sites.

Amino Acid Sequence↗

PubMed1999

Cleavage of RXRalpha by a lysosomal enzyme, cathepsin L-type protease.

In this study, we characterized a protease responsible for the cleavage of RXRalpha in two human derived cell lines, HepG2 and JEG-3 cells. Electrophoretic mobility shift assay (EMSA) combined with antibody supershift analysis suggested that contamination of cytoplasmic components during nuclear extract preparation could result in complete cleavage of RXRalpha at its N-terminus in JEG-3 cells, while such proteolytic activity was much less evident in HepG2. When the nuclei were purified in the presence of leupeptin, only full-length RXRalpha was found in the extracts prepared from both JEG-3 and HepG2 cells, suggesting a member of cysteine protease family is responsible for the cleavage. The presence of the protease in the cytoplasm, but not in the nucleus, was confirmed by incubating full-length 35S-labeled RXRalpha with each fraction. The cytoplasmic fraction from JEG-3 and HepG2 cells cleaved RXRalpha into smaller sizes with molecular mass of 45, 43, and 31 kD. Immunoprecipitation with antibodies recognizing distinct epitopes indicated that the cleaved RXRalpha with the size of 45 and 43 kD were truncated at N-terminus in which most of the A/B domain was absent. Using a series of protease inhibitors, the enzyme cleaving RXRalpha was characterized as cathepsin L-type protease. The enzyme activity in JEG-3 cells was much higher than that in HepG2 cells. This is the first demonstration that RXRalpha is cleaved by a lysosomal enzyme, cathepsin L-type protease.

Cathepsin L↗

PubMed1999

Cutaneous afferents producing a reflex pupil dilation in anesthetized rats.

Cutaneous afferents producing a reflex pupil dilation were examined using natural mechanical stimulation of the hindlimb skin and electrical stimulation of a sural nerve in anesthetized, artificially ventilated rats. Pupil diameter was continuously recorded after magnification using a microscope connected to a charge coupled device camera. Innocuous brushing, or weak pressing of the skin, did not have any effect on pupil diameter, while pressing the skin more than 720 g/cm2 produced a pressure-dependent pupil dilation. Pinching rather than pressing the skin induced the larger pupil dilation. Electrical stimulation of a sural afferent nerve with weak intensity, which was supra-threshold for Abeta-afferents and sub-threshold for Asigma-afferents, induced a reflex pupil dilation. This dilation continued to increase with further increases in stimulus intensity which involved excitation of Asigma afferents and C afferents. It is concluded that Abeta, Asigma and C afferents in the skin can work as afferents in eliciting reflex pupil dilation in anesthetized rats.

Afferent Pathways↗

PubMed1999

Clinicopathological variables and p53 overexpression as a combined prognosticator for hematogenic recurrence in colorectal cancer.

BACKGROUND AND OBJECTIVE: Precise evaluation of the prognostic factors for hematogenic recurrence after resection for colorectal cancer is important not only for the prediction of patient outcome but also for the determination of adjuvant therapy. The purpose of the current study was to elucidate the clinical significance of using clinicopathological variables in combination with p53 expression as a prognosticator for hematogenic recurrence. METHODS: One hundred forty-two patients with colorectal cancer were examined. The expression of p53 was determined by immunohistochemical staining. RESULTS: Eighteen (60%) of the 30 patients who were positive for both p53 overexpression and lymph node metastasis, 13 (41%) of the 32 patients who were positive for p53 and venous invasion, and 13 (39%) of the 33 patients who were positive for p53 and carcinoembryonic antigen (CEA) developed hematogenic recurrence. CONCLUSIONS: The combination of p53 overexpression and lymph node metastasis was an excellent prognostic indicator for hematogenic recurrence in colorectal cancer.

Adult↗

PubMed1999

Intrahepatic cholangiocarcinoma with extensive sarcomatous change: report of a case.

A 77-year-old woman was admitted to our hospital with severe upper abdominal pain. Ultrasonography showed a well-defined hypoechoic mass with heterogeneity in the left lobe of the liver, and computed tomography demonstrated a low-density mass with enhanced peripheral areas. Magnetic resonance imaging revealed a mass with iso- to low signal intensity on T1-weighted images (WI) and heterogeneous high and low signal intensity on T2 WI. The tumor was found to be hypovascular by angiography. During 5 months of observation, the tumor increased in size, which strongly suggested malignancy. A laparotomy was performed under the provisional diagnosis of a neoplasm other than hepatocellular carcinoma, revealing that the hepatic mass had invaded the gastric wall. Therefore, a left hepatic lobectomy with dissection of the lymph nodes and hemigastrectomy was carried out. Histologically, the tumor was found to be composed of a large amount of sarcomatous elements and a small amount of adenocarcinomatous elements, both of which were partly intermingled. Immunohistochemically, the sarcomatous element demonstrated the features of malignant fibrous histiocytoma (MFH). Thus, a diagnosis of intrahepatic cholangiocarcinoma with MFH-like sarcomatous change was confirmed.

Aged↗

PubMed1999

Propofol prevents delayed neuronal death following transient forebrain ischemia in gerbils.

PURPOSE: This study was conducted to ascertain whether propofol may protect against delayed neuronal death in the hippocampal CA1 subfield in gerbils. METHODS: Thirty-five gerbils were randomly assigned to five groups: Group I, the control group, a sham operation treated with physiological saline solution (PSS); Group II, ischemia/reperfusion treated with PSS; Group III, ischemia/reperfusion treated with 50 mg x kg(-1) propofol; Group IV, ischemia/reperfusion treated with 100 mg x kg(-1) propofol; Group V ischemia/reperfusion treated with 150 mg x kg(-1) propofol. Transient forebrain ischemia was induced by occluding the bilateral common carotid arteries for four minutes under N2O/O2/halothane anesthesia after administration of propofol or PSS. Five days later, histopathological changes in the hippocampal CA1 subfield were examined using a light microscope and degenerative ratio of the pyramidal cells were measured according to the following formula: (number of degenerative pyramidal cell/total number of pyramidal cells per 1 mm of hippocampal CA1 subfield) x 100. RESULTS: In group II, the pyramidal cells were atrophic and pycnotic; vacuolation and structural disruption of the radial striated zone was observed. In the other four groups, these changes were not observed. The degenerative ratios of pyramidal cells were as follows; group I: 5.9 +/- 1.9%, group II: 94.6 +/- 2.5% (P < 0.01), group III: 10.7 +/- 1.7%, group IV: 9.7 +/- 1.8%, group V: 9.2 +/- 1.9%. CONCLUSION: This study suggests that propofol may prevent delayed neuronal death in the hippocampal CA1 subfield after cerebral ischemia/reperfusion in gerbils.

Analysis of Variance↗

PubMed1999

Arterial revascularization. 18-year experience with coronary artery bypass grafting in familial hypercholesterolemia.

Familial hypercholesterolemia is characterized by a high plasma level of cholesterol and is frequently associated with rapidly progressing coronary heart disease. The internal thoracic artery is recognized as the conduit of choice for coronary artery bypass grafting. This study was performed to determine whether multiple arterial grafting was associated or not with additional benefits for patients with familial hypercholesterolemia. Between June 1980 and March 1998, 95 patients with familial hypercholesterolemia. underwent a total of 103 coronary artery bypass procedures with one hospital death. The patients were divided into 3 groups according to the type of bypass graft. Group 1 included 31 patients with only saphenous vein grafts; Group 2,48 patients with one arterial graft and supplemental vein grafts; and Group 3, 24 patients with multiple arterial grafts. The overall actuarial survival rate was 90.9% at 10 years and 81.8% at 18 years. The overall actuarial freedom from recurrent angina was 68.9% at 10 years and 55.8% at 16 years. The actuarial survival rate in group 2 was higher than that in Group 1 (p < 0.05). There was no difference in the actuarial survival or in the freedom from cardiac events between Group 2 and Group 3. Single arterial grafting improved the long-term survival in patients with familial hypercholesterolemia. However, no additional benefit from multiple arterial grafting was identified.

Coronary Artery Bypass↗

PubMed1999

Leiomyosarcoma of the thoracic aorta.

This case involves a patient with aortic leiomyosarcoma. A patient clinically suspected of type IIIa dissecting aortic aneurysm underwent surgery. The descending thoracic aorta was found to be filled with a soft, yellow tumor and was replaced with a woven Dacron graft. Microscopy of the surgical specimen revealed large, atypical spindle cells with numerous mitoses in bundles intersecting at 90 degrees, suggesting leiomyosarcoma. This diagnosis was confirmed immunohistochemical study. The postoperative course was uneventful and the patient was discharged on postoperative day 40; however, the patient developed bronchopneumonia due to malignant pleuritis 3 months postoperatively and died on postoperative day 103.

Aged↗

PubMed1999

Involvement of VEGF and its receptors in ascites tumor formation.

Vascular endothelial growth factor (VEGF) has potent endothelial cell mitotic and vascular permeability activity. Several reports have suggested that VEGF may be one of the major factors regulating ascites formation, although no quantitative and systematic analyses have been carried out. To determine the role of VEGF in ascites formation, we examined the expression of VEGF in 13 mouse ascites tumors (5 sarcomas, 3 carcinomas, and 5 hematopoietic malignancies). We found that significant amounts (6-850 ng/mL) of biologically active VEGF accumulated in the ascites fluid of all 13 tumors, particularly in tumors of sarcoma and carcinoma origin (430 +/- 234 ng/mL). The microvessel densities in the peritoneal walls of tumor-bearing mice, which are significantly higher than those in healthy mice, basically correlated with but did not parallel VEGF concentrations, suggesting the existence of an additional modulator(s) of the angiogenic process. Administration of anti-mouse VEGF-neutralizing antibody to mice bearing the carcinoma-derived ascites tumor MM2 suppressed ascites accumulation, tumor growth, and tendency to bleed. These results directly demonstrate the crucial role of VEGF in carcinoma-derived ascites tumor formation in vivo.

Animals↗

PubMed1999

Medication-induced hallucination and cerebral blood flow in Parkinson's disease.

Although hallucinations in Parkinson's disease (PD) are not unusual in the long-term treatment with anti-parkinsonian agents, their mechanism is not fully understood. We compared both the neuropsychiatric state and the results of 99mTc-labeled hexamethyl propyleneamine oxime single-photon emission computed tomography in 12 PD patients with medication-induced hallucinations and 21 PD patients without hallucinations. Hallucinatory patients showed significantly lower cerebral blood flow in left temporal regions than nonhallucinatory patients. The cerebral blood flow reduction in these regions may be related to the mechanism of medication-induced hallucinations in PD.

Aged↗

PubMed1999

In situ hybridization for matrix metalloproteinase-1 and cathepsin K in rat root-resorbing tissue induced by tooth movement.

The movement of teeth during orthodontic treatment occasionally induces undesirable root resorption. Although high collagenolytic activity has been detected in resorbing tissue of deciduous teeth, the cellular origin of collagenolytic enzymes in root-resorbing tissue caused by tooth movement has not been identified. Here, rats were subject to 7 days of experimental tooth movement to induce root resorption. In situ hybridization with digoxigenin-labelled RNA probes was performed on sections of the maxillary bone to detect the mRNAs that encode matrix metalloproteinase-1 (MMP-1) and cathepsin K in root-resorbing tissue. MMP-1 mRNA was detected in fibroblastic cells, cementoblasts and osteoblasts, but not in odontoclasts nor osteoclasts. Moreover, MMP-1 mRNA was highly expressed in some cementocytes located near odontoclasts and in many osteocytes. In contrast, cathepsin K mRNA was expressed only in odontoclasts and osteoclasts. These results suggest that MMP-1 and cathepsin K are important in root resorption during tooth movement in a mode similar to bone resorption.

Animals↗

PubMed1999

Conformational changes of bacteriorhodopsin along the proton-conduction chain as studied with (13)C NMR of [3-(13)C]Ala-labeled protein: arg(82) may function as an information mediator.

We have recorded (13)C NMR spectra of [3-(13)C]Ala-labeled wild-type bacteriorhodopsin (bR) and its mutants at Arg(82), Asp(85), Glu(194), and Glu(204) along the extracellular proton transfer chain. The upfield and downfield displacements of the single carbon signals of Ala(196) (in the F-G loop) and Ala(126) (at the extracellular end of helix D), respectively, revealed conformational differences in E194D, E194Q, and E204Q from the wild type. The same kind of conformational change at Ala(126) was noted also in the Y83F mutant, which lacks the van der Waals contact between Tyr(83) and Ala(126) present in the wild type. The absence of a negative charge at Asp(85) in the site-directed mutant D85N induced global conformational changes, as manifested in displacements or suppression of peaks from the transmembrane helices, cytoplasmic loops, etc., as well as the local changes at Ala(126) and Ala(196) seen in the other mutants. Unexpectedly, no conformational change at Ala(126) was observed in R82Q (even though Asp(85) is protonated at pH 6) or in D85N/R82Q. The changes induced in the Ala(126) signal when Asp(85) is uncharged could be interpreted therefore in terms of displacement of the positive charge of Arg(82) toward Tyr(83), where Ala(126) is located. It is possible that disruption of the proton transfer chain after protonation of Asp(85) in the photocycle could cause the same kind of conformational change we detect at Ala(196) and Ala(126). If so, the latter change would be also the result of rearrangement of the side chain of Arg(82).

Alanine↗

PubMed1999

Location of a cation-binding site in the loop between helices F and G of bacteriorhodopsin as studied by 13C NMR.

The high-affinity cation-binding sites of bacteriorhodopsin (bR) were examined by solid-state 13C NMR of samples labeled with [3-13C]Ala and [1-13C]Val. We found that the 13C NMR spectra of two kinds of blue membranes, deionized (pH 4) and acid blue at pH 1.2, were very similar and different from that of the native purple membrane. This suggested that when the surface pH is lowered, either by removal of cations or by lowering the bulk pH, substantial change is induced in the secondary structure of the protein. Partial replacement of the bound cations with Na+, Ca2+, or Mn2+ produced additional spectral changes in the 13C NMR spectra. The following conclusions were made. First, there are high-affinity cation-binding sites in both the extracellular and the cytoplasmic regions, presumably near the surface, and one of the preferred cation-binding sites is located at the loop between the helix F and G (F-G loop) near Ala196, consistent with the 3D structure of bR from x-ray diffraction and cryoelectron microscopy. Second, the bound cations undergo rather rapid exchange (with a lifetime shorter than 3 ms) among various types of cation-binding sites. As expected from the location of one of the binding sites, cation binding induced conformational alteration of the F-G interhelical loop.

Bacteriorhodopsins↗

PubMed1999

Effects of Y-24180, a receptor antagonist to platelet-activating factor, on allergic cutaneous eosinophilia in mice.

We examined the effects of Y-24180, a potent and long-acting antagonist to platelet-activating factor (PAF), on allergic cutaneous eosinophilia and cytokine production in the skin of mice. Mice sensitized actively with ovalbumin (OA) were challenged by an intradermal injection of OA solution. The number of inflammatory cells, including eosinophils and eosinophil peroxidase (EPO) activity reflecting eosinophil infiltration into the tissue increased in OA-challenged skin 12 hr after the challenge. The levels of interleukin-4 (IL-4) and IL-5 also increased significantly in the challenged skin 12 hr and 3-24 hr, respectively, but that of interferon-gamma (IFN-gamma) did not change. Then, we evaluated the effects of Y-24180, ketotifen, suplatast and prednisolone on the increase in EPO activity, IL-4 and IL-5. These drugs were orally administered once a day for 5 days beginning 4 days before the challenge. Y-24180 (10 mg/kg) and prednisolone (5 mg/kg) significantly suppressed these parameters. Suplatast did not affect EPO activity, but significantly decreased the levels of IL-4 and IL-5. Ketotifen had no effect on them. These results indicate that the inhibition of IL-4, IL-5 and PAF are required to suppress the cutaneous eosinophilia and Y-24180 contributes to the treatment of allergic cutaneous eosinophilia.

Animals↗

PubMed1999

Inhibition of activation of human peripheral blood eosinophils by Y-24180, an antagonist to platelet-activating factor receptor.

We examined the effects of Y-24180, a potent and long-acting antagonist to platelet-activating factor (PAF) receptor, on the PAF- or leukotriene B4 (LTB4)-induced activation of eosinophils using human peripheral blood in vitro. As activation markers, CD11b expression level and soluble intercellular adhesion molecule-1 (sICAM-1)-binding activity were analyzed by flow cytometry. Y-24180 significantly inhibited PAF-induced increase in the ratio of strongly positive cells for CD11b expression and sICAM-1 binding at 0.01 microM or more. WEB 2086, another PAF receptor antagonist, also inhibited the increase significantly at 1 microM or more. LTB4-induced increases in the ratio of strongly positive cells for CD11b expression and sICAM-1 binding were inhibited by Y-24180 at 1 microM, but not WEB 2086 up to 10 microM. These results indicate that Y-24180 inhibits the PAF- or LTB4-induced activation of eosinophils in human peripheral blood more potently than WEB 2086.

Azepines↗

PubMed1999

Synthesis and structure-activity relationships of 6-phenylaliphatic-substituted C19 steroids having a 1,4-diene, 4,6-diene, or 1,4,6-triene structure as aromatase inhibitors.

A series of 6alpha- and 6beta-phenylaliphatic-substituted androsta-1,4-diene-3,17-diones [9b-f and 10b-f; (CH2)nPh, n = 1-5] and their 4,6-diene and 1,4,6-triene analogs (11b-f and 12b-f) along with their respective phenyl analogs 9a-12a were synthesized and tested as aromatase inhibitors. All of the steroids examined were very powerful competitive inhibitors of aromatase in human placental microsomes with apparent Ki values ranging from 8.5 to 80 nM. The inhibitory activities of the benzyl- and phenethyl-4,6-dienes 11b and 11c (Ki, 9.0 and 10 nM) as well as the 6-phenethyl-1,4,6-triene 12c (Ki, 8.5 nM) were extremely high among them. All of the phenylaliphatic steroids, except for the 6beta-phenethyl compound 10c, and the 6-phenyl-4,6-diene 11a had higher affinity for aromatase than the corresponding parent 1,4-diene, 4,6-diene, and 1,4,6-triene steroids 9g, 11g, and 12g. All of the 6alpha-substituted 1,4-dienes (9a-9g) and the 6-substituted 1,4,6-trienes (12a-12g) caused a time-dependent inactivation of aromatase. On the other hand, only the 6beta-substituted 1,4-dienes (10a-10d) having no or less than four carbon atoms between the steroid nucleus and the phenyl group also caused a time-dependent inactivation of aromatase. Their inactivation rates (k(inact) 0.076-0.156 min(-1)) were higher than the respective parent steroids, 9g and 12g. In contrast, in the 4,6-diene series, only the 6-phenpropyl steroids 11d inactivated aromatase in a time-dependent manner with 0.155 min(-1) of k(inact) value. The inactivation was prevented by the substrate androstenedione, and no significant effect of L-cysteine on the inactivation was observed in each case. These results indicate that length and/or stereochemistry of the C-6 substituent of steroids 9-12 as well as a terminal phenyl group incorporated in the C-6 substituent play a critical role not only in tight binding to the active site of aromatase but also in the cause of a time-dependent inactivation of the enzyme.

Aromatase Inhibitors↗

PubMed1999

Induction of seizures by the potent K+ channel-blocking scorpion venom peptide toxins tityustoxin-K(alpha) and pandinustoxin-K(alpha).

The scorpion venom peptide toxins tityustoxin-K(alpha) (TsTx-K(alpha)) and pandinustoxin-K(alpha) (PiTx-K(alpha)) are novel, highly potent and selective blockers of voltage-activated K+ channels. PiTx-K(alpha) preferentially blocks rapidly inactivating (A-type) K+ channels whereas TsTx-K(alpha) is selective for slowly inactivating (delayed rectifier-type) channels. K+ channel blockers are known to induce seizures, but the specific K channel types that can serve as convulsant targets are not well defined. To address this issue, we examined for convulsant activity the K+ channel type-specific scorpion toxins and the selective K+ channel antagonists 4-aminopyridine (4-AP), an inhibitor of A-type voltage-activated K+ channels, and paxilline, a selective blocker of large conductance (maxi K) Ca(2+)-activated K+ channels. Intracerebroventricular injection of recombinant TsTx-K(alpha) and PiTx-K(alpha) in mice produced limbic and clonic-tonic seizures. The severity of the seizures increased during the 60-min period following injection, culminating in continuous clonic seizure activity (status epilepticus), tonic hindlimb extension, and eventually in death. The estimated doses producing limbic and clonic seizures in 50% of animals (CD50) for TsTx-K(alpha) and PiTx-K(alpha) were 9 and 33 ng, respectively. 4-AP produced seizure activity similar to the toxins (CD50, 76 ng) whereas paxilline failed to induce seizures at doses up to 13.5 microg. Carbamazepine protected fully against the toxin- and 4-AP-induced seizures whereas phenytoin had variable activity against the clonic component although it was protective against tonic hindlimb extension. The AMPA receptor antagonist GYKI 52466 also conferred full protection against toxin-induced seizures, but the NMDA receptor antagonists (R)-CPP and dizocilpine failed to affect limbic and clonic seizures, although they protected against hindlimb extension. We conclude that selective blockade of delayed rectifier- or A-type voltage-activated K+ channels can produce limbic, clonic and tonic seizures, whereas blockade of maxi K-type Ca(2+)-activated K+ channels does not. The convulsant effects may be related to enhanced glutamate release and, in the case of the limbic and clonic convulsions, activation of AMPA receptors.

4-Aminopyridine↗

PubMed1999

Sulfated sialic acid-polymers inhibit the cytotoxic action of bee and snake venom.

Colominic acid is an alpha2,8-linked sialic acid polymer produced by Escherichia coli. We found that synthetic sulfated-colominic acids (SC) remarkably inhibited the cytotoxicity of bee and snake venom toward mouse fibroblast cells, but colominic acids showed no inhibition themselves, indicating the important role of sulfate groups in the inhibitory activity of SC. Other sulfated carbohydrates such as chondroitin sulfates, heparin and heparan sulfate showed no inhibition. SC also exhibited potent inhibition of melittin, a highly basic peptide, which is a major cytotoxic component of bee venom. SC did not inhibit phospholipase A2 activity in bee venom. This suggests that the inhibition of bee and snake venom by SC is due to inhibition of melittin and cardiotoxin, which is a cytolytic peptide in snake venom, respectively. SC with a higher sulfur content and a larger molecular mass showed more potent activity. The interaction between SC and melittin basically seems an ionic one, however, the conformation of SC is also likely important. For the binding of SC to melittin leading loss of its cytotoxic activity, the sulfate groups of SC must be properly arranged to interact with lysine and arginine residues of melittin molecules, which play an important role in the cytolytic activity. A higher molecular mass of SC substituted with more sulfate groups is required for more obvious inhibition of the cytotoxic activity.

Animals↗