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Biomedical subjects

Y Nagano

Publications and source records attributed to Y Nagano.

At least 37 records · Page 2Linked to original sources
PubMed2000

Recombinant carboxyltransferase responsive to redox of pea plastidic acetyl-CoA carboxylase.

Acetyl-CoA carboxylase regulates the rate of fatty acid synthesis. This enzyme in plants is localized in plastids and is believed to be composed of biotin carboxyl carrier protein, biotin carboxylase, and carboxyltransferase made up of alpha and beta polypeptides, although the enzyme has not been purified yet. Accumulated evidence shows that pea plastidic acetyl-CoA carboxylase is activated by light and the activation is caused by light-dependent reduction of carboxyltransferase, but not of biotin carboxylase, via a redox cascade. To understand the reductive activation of carboxyltransferase at the molecular level here, we obtained the active enzyme composed of decahistidine-tagged (His tag) alpha and beta polypeptides through the expression of the pea plastidic carboxyltransferase gene in Escherichia coli. Gel filtration showed that the molecular size of the recombinant carboxyltransferase is in agreement with that of partially purified carboxyltransferase from pea chloroplasts. The catalytic activity of the recombinant enzyme was similar to that of native carboxyltransferase. These results indicate that the molecular structure and conformation of recombinant carboxyltransferase resemble those of its native counterpart and that native carboxyltransferase is indeed composed of alpha and beta polypeptides. This recombinant enzyme was activated by dithiothreitol, a known reductant of S-S bonds, with a profile similar to that of its native counterpart. The recombinant enzyme was activated by reduced thioredoxin-f, a signal transducer of redox potential in chloroplasts under irradiation. Thus, this enzyme was redox-regulated, like that of the native carboxyltransferase.

Acetyl-CoA Carboxylase↗
PubMed2000

Altered gene expression and secretion of interleukin-6 in stromal cells derived from endometriotic tissues.

OBJECTIVE: To compare the expression of interleukin-6 (IL-6) in endometrial and endometriotic cells. DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology, Tottori University Hospital, Yonago, Japan. PATIENT(S): Twenty patients who underwent either hysterectomy or laparoscopic surgery. INTERVENTION(S): Endometrial and endometriotic stromal cells were obtained from normal endometrium and from chocolate cyst linings of the ovary. Peritoneal macrophages were isolated from peritoneal fluids. Cells were cultured in the presence or absence of tumor necrosis factor-alpha. MAIN OUTCOME MEASURE(S): Gene expression of IL-6 was examined by Northern blot analysis. Interleukin-6 protein production was examined by immunocytochemical staining and ELISA. RESULT(S): A single IL-6 messenger RNA band of approximately 1.3 kilobases was detected in endometriotic stromal cells. Tumor necrosis factor-alpha increased the expression of IL-6 messenger RNA in endometriotic cells in a dose-dependent manner. In endometrial stromal cells, IL-6 messenger RNA signals were much weaker. Endometriotic stromal cells produced significantly larger amounts of IL-6 compared with endometrial stromal cells under basal conditions and after stimulation with tumor necrosis factor-alpha. Interleukin-6 protein was detected in cells isolated from endometriotic tissues by immunocytochemical staining. Interleukin-6 production by cultured macrophages from patients with endometriosis and endometriotic stromal cells was comparable. CONCLUSION(S): Altered gene expression and protein secretion of IL-6 in patients with endometriosis may contribute to the pathogenesis of the disease and/or to endometriosis-associated infertility.

Adult↗
PubMed2000

Tumor necrosis factor-alpha promotes proliferation of endometriotic stromal cells by inducing interleukin-8 gene and protein expression.

Endometriosis, a common disease among women of reproductive age, is characterized by the presence of endometrial-like tissue outside the uterus. We and others showed that several cytokine levels, including interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNFalpha), are elevated in the peritoneal fluid of women with endometriosis compared with those in women without endometriosis. We also demonstrated that the addition of IL-8 to the culture medium stimulated the proliferation of cultured endometriotic stromal cells. TNFalpha is a multipotent cytokine that induces IL-8 production in various cell types. Therefore, we hypothesized that TNFalpha may also contribute to the pathogenesis of endometriosis by inducing the production of IL-8. To test this hypothesis, we analyzed the peritoneal fluid concentrations of IL-8 and TNFalpha using enzyme-linked immunosorbent assay (ELISA). We observed a significant correlation between the levels of TNFalpha and IL-8 in the peritoneal fluid of endometriosis patients. We also obtained the endometriotic stromal cells from chocolate cyst linings of the ovary. The expression of the receptors for TNFalpha (TNFR) was examined by RT-PCR. We observed the expression of both TNFR-I and TNFR-II genes in endometriotic stromal cells. The expression of IL-8 gene and protein was analyzed by Northern blot hybridization and enzyme-linked immunosorbent assay, respectively. TNFalpha induced the gene and protein expression of IL-8 in endometriotic stromal cells in a dose-dependent fashion. The addition of TNFalpha promoted the proliferation of the endometriotic stromal cells, and the stimulatory effects of TNFalpha were abolished by adding anti-IL-8 antibody. We demonstrated for the first time that TNFalpha stimulated proliferation of endometriotic stromal cells through induction of IL-8 gene and protein expression. We concluded that the TNFalpha may be one of the essential factors for the pathogenesis of endometriosis.

Adult↗
PubMed2000

Effect of a thromboxane A(2) antagonist on sputum production and its physicochemical properties in patients with mild to moderate asthma.

STUDY OBJECTIVE: To determine the effects of a specific thromboxane A(2) (TxA(2)) receptor antagonist, seratrodast, on asthma control and airway secretions. DESIGN: Multicenter, double-blind, randomized, placebo-controlled study. PATIENTS: Forty-five patients with mild to moderate asthma who had been continuously expectorating sputum of > 20 g/d. Patients with a current pulmonary infection or taking oral corticosteroids, antibiotics, or mucolytic agents were excluded from the trial. INTERVENTIONS: Following a 2-week run-in period, while pulmonary function, sputum production, and mucociliary function were assessed, patients were assigned to receive seratrodast, 40 mg/d, or placebo for 6 weeks. MEASUREMENTS AND RESULTS: During the treatment period, the changes in FEV(1) and peak expiratory flow (PEF) were not different between the two patient groups, but there were significant reductions in diurnal variation of PEF (p = 0.034), frequency of daytime asthma symptoms (p = 0.030), and daytime supplemental use of beta(2)-agonist (p = 0.032) in the seratrodast group. For sputum analysis, seratrodast treatment decreased the amount of sputum (p = 0.005), dynamic viscosity (p = 0. 007), and albumin concentration (p = 0.028), whereas it had no effect on elastic modulus or fucose concentration. Nasal clearance time of a saccharin particle was shortened in the seratrodast group at week 4 (p = 0.031) and week 6 (p = 0.025), compared with the placebo group. CONCLUSION: Blockade of TxA(2) receptor has minimal effects on pulmonary function, but may cause an improvement in mucociliary clearance by decreasing the viscosity of airway secretions.

Adult↗
PubMed2000

[Presacral malignant peripheral nerve sheath tumor in a patient with von Recklinghausen disease: case report].

A 32-year-old woman was admitted to our institution with dysuria and pain in the left foot. She was diagnosed with von Recklinghausen disease based on the physical findings, and an elastic, firm mass was palpable with the fingertip on the posterior wall of the rectum. Ultrasonography, computed tomography, and magnetic resonance imaging revealed a solid tumor, which measured 8 cm in diameter, in the presacral space. The tumor was diagnosed as a presacral solid tumor and the tumor was extirpated. The specimen was a 10 x 8 x 8 cm yellowish solid tumor weighing 250 g. The histopathologic diagnosis was malignant peripheral nerve sheath tumor (MPNST). Presacral MPNST is rarely diagnosed in the early stage, and the prognosis for MPNST patients with von Recklinghausen disease is relatively poor. Tumor resection with wide surgical margins is necessary and close follow-up after surgery is mandatory.

Adult↗
PubMed2000

[Anesthetic management of tracheal stent insertion under total intravenous anesthesia--a report of two cases].

Tracheal stent insertion is a useful method for patients with malignant tracheal stenosis. Expandable metal stents were inserted in two patients with severe dyspnea due to tracheal stenosis caused by lung cancer and esophageal cancer. The tracheas were intubated after spraying the pharynx with 4% lidocaine solution. The respiration was assisted. Anesthesia was maintained by intravenous propofol. There were no episodes of coughing during and after the procedure.

Aged↗
PubMed1999

Overexpression of salt-tolerant glutaminase from Micrococcus luteus K-3 in Escherichia coli and its purification.

A high-expression plasmid, pKSGHE3-1, containing the salt-tolerant glutaminase (EC 3.5.1.2) from marine bacterium Micrococcus luteus K-3 was constructed. pKSGHE3-1 was made by inserting the DNA fragment (1.43 kb) containing the structural gene synthesized by polymerase chain reaction into the downstream region of the tac promoter of expression vector pKK223-3. The translational start codon was located 10 bases downstream of the Shine-Dalgarno sequence (AGGA) of pKK223-3. Escherichia coli JM109 transformed with pKSGHE3-1 exhibited more than 190-fold higher glutaminase activity than M. luteus K-3 under optimal culture conditions. The enzyme was purified to homogeneity through three column chromatography steps with a final yield of 17.1%. The recombinant enzyme showed the same enzymatic properties, including salt tolerance, as those of M. luteus K-3. This glutaminase expression system allows the production of sufficient quantities of glutaminase for basic structure-function studies including chemical modification and future X-ray crystallization analysis.

Bacterial Proteins↗
PubMed1999

Menstrual cycle-specific inhibition of the proliferation of endometrial stromal cells by interleukin 6 and its soluble receptor.

OBJECTIVE: This study investigated the possible roles of interleukin 6 and soluble interleukin 6 receptor in the growth of endometrial and endometriotic cells. STUDY DESIGN: Endometrial and endometriotic stromal cells were collected from the uterus or from ovarian chocolate cysts. We examined the effects of interleukin 6, soluble interleukin 6 receptor, and a combination of both factors on the proliferation of endometrial and endometriotic stromal cells. The action of sex steroids on the interleukin 6 regulation of the growth of stromal cells was also evaluated. The gene expressions of interleukin 6 receptor and glycoprotein 130 were examined in endometrial and endometriotic cells by reverse transcription-polymerase chain reaction. RESULTS: Interleukin 6 had no effect on the growth of stromal cells in tissue from the proliferative phase. In contrast, the addition of concentrations of >/=100 pg/mL interleukin 6 induced significant inhibition of stromal cell proliferation in tissue from the secretory phase. Similarly, the addition of soluble interleukin 6 receptor caused significant suppression in the growth of endometrial stromal cells in tissue from the secretory phase but not the proliferative phase. On the other hand, stromal cells of endometriotic tissues were resistant to interleukin 6, showing no inhibitory response. Although the combination treatment did not affect the proliferation of stromal cells of the proliferative phase and of endometriotic tissues, 10 pg/mL interleukin 6 inhibited proliferation of stromal cells of the secretory phase in the presence of 1 ng/mL soluble interleukin 6 receptor. Treatment with estradiol and progesterone for 10 days newly induced the inhibitory response to interleukin 6 in the endometrial cells from the proliferative phase. Expressions of transcripts of interleukin 6 receptor and glycoprotein 130 were observed in the endometrial cells from the proliferative and secretory phases and in endometriotic cells. CONCLUSIONS: Interleukin 6 may play a central role in regulation of the growth of endometrial cells as a mediator of endocrine action. Endometriotic cells may behave differently from their normal counterparts in terms of the inhibitory regulation exerted by interleukin 6.

Antigens, CD↗
PubMed1999

Moyamoya disease presenting as cerebral infarction after cesarean.

BACKGROUND: Moyamoya disease with pregnancy is rare and might present with cerebral hemorrhage. CASE: A 22-year-old primigravida suddenly developed muscular weakness in the right arm and facial discomfort 3 days after cesarean. Computed tomography and cerebrovascular angiography found cerebral infarction attributable to moyamoya disease. Bilateral anastomosis of superficial temporal and middle cerebral arteries was done. CONCLUSION: Moyamoya disease with pregnancy might present as cerebral infarction after cesarean.

Adult↗
PubMed1999

Identification of a cis-regulatory element involved in phytochrome down-regulated expression of the pea small GTPase gene pra2.

The pra2 gene encodes a pea (Pisum sativum) small GTPase belonging to the YPT/rab family, and its expression is down-regulated by light, mediated by phytochrome. We have isolated and characterized a genomic clone of this gene and constructed a fusion DNA of its 5'-upstream region in front of the gene for firefly luciferase. Using this construct in a transient assay, we determined a pra2 cis-regulatory region sufficient to direct the light down-regulation of the luciferase reporter gene. Both 5'- and internal deletion analyses revealed that the 93-bp sequence between -734 and -642 from the transcriptional start site was important for phytochrome down-regulation. Gain-of-function analysis showed that this 93-bp region could confer light down-regulation when fused to the cauliflower mosaic virus 35S promoter. Furthermore, linker-scanning analysis showed that a 12-bp sequence within the 93-bp region mediated phytochrome down-regulation. Gel-retardation analysis showed the presence of a nuclear factor that was specifically bound to the 12-bp sequence in vitro. These results indicate that this element is a cis-regulatory element involved in phytochrome down-regulated expression.

Amino Acid Sequence↗
PubMed1999

Role of the oxygen uptake efficiency slope in evaluating exercise tolerance.

OBJECTIVE: To investigate the interprotocol agreement of oxygen uptake efficiency slope (OUES). METHODS: 16 Japanese children and adolescents (10 boys and six girls) underwent two sessions of maximal exercise testing according to the following two treadmill protocols: the standard Bruce protocol and the rapidly increasing staged (RIS) protocol. Maximal oxygen uptake (VO2max), the ventilatory anaerobic threshold (VAT), and the OUES were obtained from the gas analysis data. Agreement between the protocols was tested by means of the Bland-Altman method. RESULTS: Interprotocol agreement was excellent for the OUES (limit of agreement, -18% to 17% of the mean value), slightly less good for VO2max (limit of agreement, -20% to 24% of the mean value), and poor for the VAT (limit of agreement, -31% to 31% of the mean value). CONCLUSION: These results confirm the clinical usefulness of the OUES as a measure of evaluating exercise tolerance in the paediatric population.

Adolescent↗
PubMed1999

Role of cytokines in progression of endometriosis.

Peritoneal fluid in women with endometriosis contains an increased number of activated macrophages that secrete a variety of cytokines, including interleukin (IL)-6, IL-8, vascular endothelial growth factor, and tumor necrosis factor-alpha (TNF-alpha). Cytokines may be involved in the control of implantation and the growth of endometrial cells outside the uterus. In addition, several cytokines have been implicated in or directly associated with angiogenic activity in endometriosis. There could be a relationship between the levels of cytokines in the peritoneal fluid of patients with endometriosis and the status of the lesions in such patients. Peritoneal endometriosis can be classified as having red, black, or white lesions. Red lesions are known to be an active form of early endometriosis, because vascularization and mitotic activity are shown to be most prominent in these lesions. We found that the peritoneal fluid levels of TNF-alpha and IL-8 were significantly higher in patients with endometriosis, and correlated with the size and number of active lesions. In addition, TNF-alpha and IL-8 stimulated the growth of ectopic endometrial stromal cells. These cytokines with angiogenic activity may therefore have significant roles in the pathogenesis of endometriosis.

Cell Division↗
PubMed1999

Effect of cromolyn on adenosine-induced airway microvascular leakage in sensitized rats.

Inhalation of adenosine causes bronchoconstriction in asthmatic subjects, but the effect of this purine nucleotide on airway vascular permeability is unknown. In order to determine whether adenosine produces airway microvascular leakage and, if so, to examine the effect of cromolyn (sodium cromoglycate (SCG)) on this extravasation of Evans blue was measured in the airways of ovalbumin-sensitized Brown Norway rats. Inhaled adenosine caused microvascular leakage in sensitized but not in non-sensitized rats, and the response was abolished by capsaicin pretreatment or the tachykinin neurokinin-1 receptor antagonist FK888. Adenosine-induced vascular leakage became apparent in nonsensitized rats when treated with phosphoramidon, and airway neutral endopeptidase activity was lower in sensitized than in non-sensitized animals. The extravasation induced by adenosine in sensitized rats was dose dependently inhibited by SCG aerosols, SCG likewise inhibited microvascular responses to substance P, but had no effect on those to platelet-activating factor. These results suggest that: 1) adenosine induces airway microvascular leakage in sensitized rats through stimulation of neurokinin-1 receptors; 2) this effect is associated with a sensitization-induced decrease in neutral endopeptidase activity; and 3) sodium cromoglycate inhibits adenosine-induced extravasation, presumably via functional antagonism of tachykinins.

Adenosine↗
PubMed1999

[Endothelial injury by oxidized LDL].

Recent progress in the research of oxidized LDL has revealed that this lipoprotein causes not only foam cell transformation of macrophages but also several endothelial dysfunction, and the effects on endothelial cells are also involved with the process of atherogenesis. Receptors for oxidized LDL on endothelial cells, such as LOX-1 and SREC, have been cloned and their characteristics are now under investigation. In addition to lowering plasma cholesterol level, it is expected that new strategies to prevent atherosclerosis is established by focusing on the endothelial injury caused by oxidized LDL.

Arteriosclerosis↗
PubMed1998

Cholesterol efflux effect of high density lipoprotein is impaired by whole cigarette smoke extracts through lipid peroxidation.

It has been reported that high density lipoprotein (HDL) plays an anti-atherogenic role by stimulating cholesterol efflux from the foam cells in the atheromatous lesion. In this study, we prepared a novel modified form of HDL (CS-HDL) by incubating HDL with whole cigarette smoke (CS) extracts containing both particulate matter and gas-phase smoke, and examined its effect on cholesterol efflux. CS-HDL showed a marked increase of conjugated dienes and denaturation of apoA-I, a major protein component of HDL. The cholesterol efflux effect of CS-HDL was remarkably reduced to the same level as that of oxidatively modified HDL induced by copper ion (Ox-HDL). Addition of 20 microg/ml superoxide dismutase (SOD) during the CS-modification of HDL caused retrieval of cholesterol efflux activity by 53% and a remarkable decrease in the conjugated dienes level. SOD, however, had no ameliorative effect on apoA-I denaturation. When HDL was incubated only with gas-phase smoke (gasCS-HDL), neither increase of conjugated dienes nor impairment of the cholesterol efflux effect was observed, whereas apoA-I was denaturated to the same extent as seen in CS-HDL. These results indicate that whole CS-extracts, but not gas-phase smoke, reduces cholesterol efflux effect of HDL and that lipid peroxidation associated with superoxide anion is involved in this functional impairment.

Animals↗
PubMed1998

Purification and primary structure of pituitary adenylate cyclase activating polypeptide (PACAP) from the brain of an elasmobranch, stingray, Dasyatis akajei.

Pituitary adenylate cyclase activating polypeptide (PACAP) was isolated from ovine hypothalami and found to exist as two amidated forms with 38 (PACAP 38) and 27 (PACAP 27) residues. The amino acid sequences of PACAPs isolated from the vertebrates, such as a bird, a frog and teleost fish, appear to be well conserved. In the present study, we attempted to isolate PACAP from the brain of an elasmobranch fish, Dasyatis akajei (stingray), which belongs to the Chondrichthyes (cartilaginous fish), by extraction of the acetone-dried powder with acetic acid, followed by successive high-performance liquid chromatography (HPLC) on a gel-filtration, a cation-exchange and two reverse-phase columns. Purification was monitored by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blotting analysis using an anti-PACAP 27 serum. The PACAP thus obtained consisted of 44 residues. The amino acid sequence of the comparable portion of its N-terminal 38 residues showed 92%, 89%, 89%, and 82% identity with those of mammalian, chicken, frog and teleost PACAPs with 38 residues, respectively. The extra six C-terminal residues of the stingray resembled those of tetrapod and teleost PACAP precursors which were deduced from the respective cDNAs. These results indicate that PACAP, which has an amino acid sequence showing high similarity with those of tetrapod and teleost PACAPs, is present in the elasmobranch brain.

Amino Acid Sequence↗