Biomedical subjects

K Hamada

Publications and source records attributed to K Hamada.

At least 127 records · Page 7Linked to original sources

PubMed1999

Effect of amino acids and glucose on exercise-induced gut and skeletal muscle proteolysis in dogs.

The effect of amino acid and/or glucose administration before and during exercise on protein metabolism in visceral tissues and skeletal muscle was examined in mongrel dogs. The dogs were subjected to treadmill running (150 minutes at 10 km/h and 12% incline) and intravenously infused with a solution containing amino acids and glucose (AAG), amino acids (AA), glucose (G) or saline (S) in randomized order. The infusion was started 60 minutes before exercise and continued until the end of the exercise period. An arteriovenous-difference technique was used to estimate both tissue protein degradation and synthesis. When S was infused, the release of leucine (Leu) from the gut and phenylalanine (Phe) from the hindlimb significantly increased during exercise, thus indicating that exercise augmented proteolysis in these tissues. The balance of Leu across the gut during exercise demonstrated a net uptake with both AAG and AA, whereas a net release was observed for G and S. In addition, Leu uptake in the gut during the last 90 minutes of the exercise period tended to be greater with AAG versus AA (P = .06). Phe balance across the hindlimb during the late exercise period showed a significant release with S, AA, and G, whereas the balance with AAG did not show a significant release. These results suggest that exercise-induced proteolysis in the gut may be reduced by supplementation with AA, and this effect may be enhanced by concomitant G administration. However, in skeletal muscle, both AA and G may be required to prevent net protein degradation during exercise. G provided without AA did not achieve net protein synthesis in either tissue.

Amino Acids↗

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↗

PubMed1999

Expression of mRNA for androgen receptor, 5alpha-reductase and 17beta-hydroxysteroid dehydrogenase in human dermal papilla cells.

In order to determine whether adrenal and gonadal weak androgens are utilized to form active androgens in human hair, we studied the expression of mRNA for androgen receptor (AR), 5alpha-reductase and 17beta-hydroxysteroid dehydrogenase (17beta-HSD) in cultured dermal papilla cells (DPCs) from various body sites. AR mRNA was expressed in beard (Be) and axillary hair (Ax) DPCs from both sexes, but only at a low level in DPCs from occipital scalp hair (OS). Type I 5alpha-reductase mRNA was expressed by all DPCs. Type II 5alpha-reductase mRNA was identified in beard DPCs, but was absent from OS and Ax DPCs. Type II 17beta-HSD mRNA was strongly expressed in outer root sheath cells (ORSCs), while DPCs except for male Ax expressed no type II 17beta-HSD mRNA. In contrast, type III 17beta-HSD mRNA was strongly expressed in Be DPCs and Ax DPCs from both sexes; ORSCs showed a low level of expression. Expression of type III 17beta-HSD mRNA was not regulated by androgen in DPCs. These results suggest that the sensitivity of hairs to androgen is partially controlled by the site-specific expression of AR, 5alpha-reductase and 17beta-HSD in DPCs.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗

PubMed1999

Triggering of neuronal cell death by accumulation of activated SEK1 on nuclear polyglutamine aggregations in PML bodies.

BACKGROUND: A novel class of inherited human neurodegenerations is now known to be caused by expanded CAG repeats encoding polyglutamines. Polyglutamine-containing protein fragments have been shown to accumulate as aggregates in the nucleus and in the cytoplasm, and to induce cell death when expressed in cultured cells, leading to the proposal that polyglutamine aggregation is an important step in the pathogenesis. Supporting this, nuclear inclusions containing expanded polyglutamines have been identified in neurones from the brains of patients and in neurones from transgenic mouse models of this class of neural disorders. RESULTS: We analysed the consequences of polyglutamine expression in PC12 neuronal cells. Activated SEK1 accumulated with nuclear but not cytoplasmic polyglutamine aggregations, which consequently triggers cell death. Cell death induced by polyglutamine expression was inhibited by a dominant-negative SEK1 (DN-SEK1), but not by DN-SEK1 tagged with a nuclear export signal. Steady state SEK1 expression itself was enhanced two to three-fold. Nuclearly aggregated polyglutamines, which were identified in PML bodies, co-localized with not only activated SEK1 but also activated c-Jun. We also observed that nuclear inclusion-positive neurones from brains with Huntington's disease expressed SEK1. CONCLUSIONS: This study provides molecular links between the neurodegeneration observed in polyglutamine diseases, cell death signalling kinase cascades and nuclear subdomains related to cell death. We propose that the nuclear PML bodies containing polyglutamine aggregates activate the SEK1-JNK kinase cascade, resulting in the transduction of a death signal.

Animals↗

PubMed1999

Efficient screening for catalytic antibodies using a short transition-state analog and detailed characterization of selected antibodies.

One of the major obstacles to acquiring catalytic antibodies is that it requires labor-intensive procedures to select catalytic antibodies from huge repertories of antibodies. Here, we selected potential catalytic Abs by utilizing their affinity towards a short transition-state analog which contained only the transition-state structural element, and evaluated in detail its efficiency to enrich catalytic Abs. Hybridoma supernatants elicited against a phosphonate derivative, the TSA1, were screened by a three-step screening process: step 1, ELISA for TSA1-BSA; step 2, ELISA for the short TSA4; and step 3, competitive-inhibition by the short TSA2. Only 22. 8% of positive mAbs from step 1 were found to be catalytic. The rate of catalytic Abs increased to 45.7% using screening steps 1 plus 2, and reached 83.3% using all three screening steps. This clearly suggests that our screening protocol is an efficient method to select potential catalytic Abs. Furthermore, we characterized the properties of both the catalytic Abs and the noncatalytic Abs in detail. The catalytic Abs tended to have lower Kd for TSA1 and the short TSA2 than noncatalytic Abs. It was also observed that catalytic Abs showed clear enantiospecificity toward substrate 6 containing d-phenylalanine while noncatalytic Abs did not. The detailed analysis of kinetic and binding parameters for these antibodies gives us further insight into catalytic antibodies.

Animals↗

PubMed1999

Contractile responses to alpha1-adrenoceptor agonists in isolated human male and female urethra.

OBJECTIVE: To investigate the contractile responses mediated through alpha1-adrenoceptors in human urethra and to evaluate the effectiveness of NS-49 [(R)-(-)-3'-(2-amino-1-hydroxyethyl)-4'-fluoromethanesulphonanilide++ + hydrochloride], a novel alpha1-adrenoceptor agonist, against contraction of the human urethra. MATERIALS AND METHODS: The contractile responses were assessed in 10 male prostatic urethrae and six female urethrae. Antagonism was evaluated in the urethra using phenylephrine, a nonselective alpha1-adrenoceptor agonist, cumulatively applied > 20 min after applying 0.1 micromol/L prazosin or 0.1 micromol/L 5-methylurapidil, a selective alpha1A-adrenoceptor antagonist. Agonism was determined in both male and female urethrae to obtain the concentration-response curve for the agonist. RESULTS: Phenylephrine caused both male and female urethrae to contract, and showed high potency and efficacy. Prazosin antagonized these contractions with low affinity (apparent pKB of 8.30 in male urethrae). 5-Methylurapidil, also antagonized the contractions with low affinity (apparent pKB of 7.88 in male urethrae). Noradrenaline and phenylephrine caused both male and female urethrae to contract, with high potency and efficacy. A novel and selective alpha1A-and alpha1L-adrenoceptor agonist, NS-49, induced contractile responses with high potency and moderate efficacy, whereas methoxamine induced contractions with low potency and moderate efficacy. Norephedrine was a very weak contractile agonist. CONCLUSION: In the human urethra, phenylephrine-induced contractions were mediated through alpha1L-adrenoceptors and not through alpha1A-adrenoceptors. Contractions of the human urethra induced by NS-49 were also mediated mainly through alpha1L-adrenoceptors, with high potency and moderate efficacy. NS-49 may therefore be useful for the treatment of urinary stress incontinence, with minimal side-effects because it has subtype selectivity.

Adrenergic alpha-Agonists↗

PubMed1999

Effects of environmental aerosols on airway hyperresponsiveness in a murine model of asthma.

Increased morbidity in persons suffering from inflammatory lung diseases, such as asthma and bronchitis, has been associated with air pollution particles. One hypothesis is that particles can cause an amplification of the pulmonary inflammation associated with these diseases, thus worsening affected individuals' symptoms. This hypothesis was tested in a murine model of asthma by inhalation exposure to (1) concentrated air particles (CAPs), (2) the leachate of residual oil fly ash (ROFA-S), and (3) lipopolysaccharide (LPS). Allergen-sensitized mice (ip ovalbumin, OVA) were 21 days old when challenged with an aerosol of 3% OVA in phosphate-buffered saline (PBS) for 10 min (controls were challenged with PBS only) for 3 days. On the same days, mice were further exposed to 1 of 3 additional agents: CAPs (or filtered air) for 6 h/day; LPS (5 microg/ml, or PBS) for 10 min/day; or ROFA-S (leachate of 50 mg/ml, or PBS) for 30 min on day 2 only. At 24 h later, mice challenged with OVA aerosol showed airway inflammation and airway hyperresponsiveness (AHR) to methacholine (Mch), features absent in mice challenged with PBS alone. Both OVA- and PBS-challenged mice subsequently exposed to ROFA-S showed increased AHR to Mch when compared to their respective controls (OVA only or PBS only). In contrast, when OVA-challenged mice were further exposed to CAPs or LPS, no changes in AHR were seen in comparison to mice challenged with OVA only. Bronchoalveolar lavage (BAL) analysis and histopathology 48 h postexposure showed OVA-induced allergic inflammation. No significant additional effects were caused by CAPs or ROFA-S. LPS, in contrast, caused significant increases in total cell, macrophage, and polymorphonuclear cell numbers. The data highlight discordance between airway inflammation and hyperresponsiveness.

Air Pollutants↗

PubMed1999

Clinical courses and prognoses of pulmonary sarcoidosis.

In this article, long-term prognoses and prognostic factors of patients with sarcoidosis are reviewed. In patients with intrathoracic sarcoidosis, functional impairments and parenchymal lesions at the time of initial examination strongly predicted an unfavorable prognosis. We also discussed the significance of extrathoracic lesions in terms of clinical outcomes of intrathoracic sarcoidosis. In addition, we focused on the genetic approach and the new insights being offering with respect to the disease susceptibility and the development of pulmonary lesions.

Disease Susceptibility↗

PubMed1999

Role of the midline brainstem in feline amygdaloid kindling.

PURPOSE: To study the role played by the midline brainstem in feline amygdaloid kindling in general and the formation of transhemispheric positive transfer effect (PTE) in particular. METHODS: Eight adult male cats with either anterior or posterior midline brainstem bisection underwent primary and secondary site kindling and primary site retest. RESULTS: Stage 4 seizure pattern was altered with either absence of contralateral circling in the anterior group or emergence of ipsilateral circling in the posterior group. Electroclinical manifestation of the tonic phase was absent in the posterior but not the anterior group. Kindled convulsive pattern remained asymmetrical with contralateral dominance in the anterior but not the posterior group. Intermittent truncation of the secondary clonic phase of the kindled seizure occurred in both groups. PTE was reduced whereas the negative transfer effect was preserved in both groups. CONCLUSIONS: The anterior half of the midline brainstem participates in stage 4 contralateral circling. Both the anterior and posterior halves of the midline brainstem participate in the mechanisms of repetitive circling during stage 4 and the stable maintenance of the kindled stage 6 seizure. The anterior and posterior halves of the midline brainstem participate in symmetrical patterning and electroclinical tonic expression of the kindled seizure, respectively. The midline brainstem, extending from the midbrain to the pons with some posterior emphasis, participates in the mechanisms of PTE formation. The findings indicate the importance of the brainstem for electroclinical ictal processes and a transhemispheric positive transfer effect associated with temporal limbic epileptogenesis.

Amygdala↗

PubMed1999

The clinical significance of core promoter and precore mutations during the natural course and interferon therapy in patients with chronic hepatitis B.

OBJECTIVE: We aimed to determine the clinical significance of mutations in core promoter and precore regions in chronic hepatitis B. We investigated changes in these mutations during the natural course and interferon therapy in patients with chronic hepatitis B. METHODS: A total of 93 patients with hepatitis B virus surface antigen were divided into four groups according to hepatitis B e antigen (HBeAg)/anti-HBe status and serum aminotransferase levels. Group I (n = 16) comprised HBeAg-positive patients with normal aminotransferase levels, group II (n = 31) HBeAg-positive patients with elevated aminotransferase levels, group III (n = 30) anti-HBe-positive patients with normal aminotransferase levels, and group IV (n = 16) anti-HBe-positive patients with elevated aminotransferase levels. All patients of group II and seven of group IV were treated with interferon. Three serial serum samples per untreated patient and eight samples per treated patient were tested for HBV DNA levels and core promoter and precore mutations by polymerase chain reaction combined with restriction fragment length polymorphism, and some were cloned and sequenced. RESULTS: Core promoter mutation was found in 38% of group I, 74% of group II, 97% of group III, and 100% of group IV. Precore mutation was found in 6% of group I, 90% of group II, and 100% of groups III and IV. The HBV DNA levels were significantly higher in groups I, II, IV, and III, in that order. Serial determination of these two mutations and viral levels showed that the core promoter mutation appeared to occur first, followed by a completion of the precore mutation along with a decrease in viral levels in patients who seroconverted to anti-HBe after interferon therapy. Interferon therapy suppressed both precore wild- and mutated-type viral levels equally. However, it did not induce any specific mutations. CONCLUSIONS: Core promoter mutation appeared to develop or complete first, followed by completion of the precore mutation, and the virus with these two mutations seemed to be the form to persist in the natural course of chronic hepatitis B. The clinical significance of these mutations appeared to be profoundly associated with the viral levels.

Adolescent↗

PubMed1999

Amino acid residues in the omega-minus region participate in cellular localization of yeast glycosylphosphatidylinositol-attached proteins.

The final destination of glycosylphosphatidylinositol (GPI)-attached proteins in Saccharomyces cerevisiae is the plasma membrane or the cell wall. Two kinds of signals have been proposed for their cellular localization: (i) the specific amino acid residues V, I, or L at the site 4 or 5 amino acids upstream of the GPI attachment site (the omega site) and Y or N at the site 2 amino acids upstream of the omega site for cell wall localization and (ii) dibasic residues in the region upstream of the omega site (the omega-minus region) for plasma membrane localization. The relationships between these amino acid residues and efficiencies of cell wall incorporation were examined by constructing fusion reporter proteins from open reading frames encoding putative GPI-attached proteins. The levels of incorporation were high in the constructs containing the specific amino acid residues and quite low in those containing two basic amino acid residues in the omega-minus region. With constructs that contained neither specific residues nor two basic residues, levels of incorporation were moderate. These correlations clearly suggest that GPI-attached proteins have two different signals which act positively or negatively in cell wall incorporation for their cellular localization.

Amino Acid Sequence↗

PubMed1999

Alveolar cell stretching in the presence of fibrous particles induces interleukin-8 responses.

Inhalation of fibrous particulates is strongly associated with lung injury, but the molecular and cellular mechanisms that could explain the fiber-induced pathogenesis are not fully understood. We hypothesized that the physical stress exerted on the alveolar epithelium by the deposited fibers is greatly enhanced by the tidal cyclic motion of the epithelial cells that is associated with breathing, and that this initial mechanical interaction triggers a subsequent cell response. To test this hypothesis, we developed a dynamic model of fiber-induced cell injury using a cell-stretcher device. We exposed a cyclically stretched monolayer of the human alveolar epithelial cell line A549 to glass or crocidolite asbestos fibers for 8 h and then measured the production of the proinflammatory cytokine interleukin (IL)-8 as a readout of fiber-induced cell injury. Cyclic stretching significantly increased IL-8 production in the fiber-treated cultures, suggesting that the physical stress on the cells caused by the fibers was indeed enhanced by the motion. Coating of the asbestos fibers with fibronectin, a glycoprotein abundant in the alveolar lining fluid, further increased the fiber-induced cell response when the cells were cyclically stretched. This response was, however, significantly reduced by introducing into the culture medium, before fiber treatment, soluble RGD (Arg-Gly-Asp)-containing peptides, which specifically block binding to integrin receptors upon RGD attachment. These results suggested that adhesive interactions between protein-coated fibers and cell surface molecules are involved in the fiber-induced pathogenic process. Our novel findings indicate the importance of physical insults in fiber-induced cell stress, and bring to the forefront the need to study the mechanisms involved in this process.

Asbestos, Crocidolite↗

PubMed1999

Detection of alternative splicing of fibronectin mRNA in a single cell.

Pre-fibronectin mRNA is subject to alternative splicing at three sites, EDA, EDB and IIICS. We analyzed the alternative splicing of fibronectin mRNA in a single cell. Reverse transcription-polymerase chain reaction analyses showed cells that produced a single form of mRNA at each one of these sites as well as cells that produced multiple forms at a given site: for example, some cells produced either the EDA(+) or EDA(-) form of the mRNA and other cells produced both forms. About 80% of the cells produced both (+) and (-) forms of the mRNA at the EDA and EDB sites, and the remaining cells contained either the (+) or (-) form. Five forms of fibronectin mRNA can result from alternative splicing at the IIICS site. Complex combinations of alternative splicing products were observed among the individual cells: there were ten different combinations of mRNA isoforms with respect to the IIICS site. Statistically significant changes in alternative splicing at the IIICS site were observed during cellular senescence.

Alternative Splicing↗

PubMed1999

Serum amyloid A, cytokines, and corticosterone responses in germfree and conventional mice after lipopolysaccharide injection.

To determine why germfree mice are less susceptible to lipopolysaccharide (LPS) than conventional mice, we studied serum levels of serum amyloid A (SAA), tumor necrosis factor (TNF), interleukin 1 (IL-1), IL-6, and corticosterone in mice after treatment with LPS. A single injection of LPS caused an elevation of SAA, an acute-phase protein in the mouse, in both conventional and germfree IQI mice, and the response was significantly less in germfree mice. LPS-induced elevations of serum TNF, IL-1, and IL-6 levels were also significantly less in germfree mice, while serum corticosterone levels were greater in germfree mice than in conventional mice. These results suggest that the lower susceptibility to LPS and a smaller response of SAA elevation by LPS in germfree mice may result from less elevation in serum of these cytokines in these mice, which are known to mediate the acute phase response of SAA. High levels of serum corticosterone in germfree mice may be partly responsible for the lower responsiveness of these inflammatory cytokines to LPS in these mice.

Animals↗

PubMed1999

The inferior capsular shift operation for instability of the shoulder. Long-term results in 34 shoulders.

We reviewed 26 patients with 34 shoulders treated by the inferior capsular shift operation for inferior and multidirectional instability. The mean follow-up was 8.3 years. In total, 12 shoulders showed voluntary subluxation. Eight operations used an anterior and posterior approach, 11 were by the posterior route, and 15 shoulders had an anterior approach. In 30 shoulders (85%) the outcome was satisfactory and 20 (59%) scored good or excellent results on the Rowe system. Instability had recurred in nine shoulders (26%) from three months to three years after the operation. Six of the 12 shoulders with voluntary subluxation (50%) had recurrence, as against three of the other 22 (14%), a statistically significant difference. The operation is therefore not indicated for voluntary subluxation. The 19 shoulders which had been assessed in 1987 at a mean of 3.5 years after surgery, were also reviewed in 1995 and found to have no significant changes in instability or Rowe score. This shows that the capsular shift appeared to have maintained its tension over an eight-year period. After the use of a posterior approach, 64% of the shoulders showed a posterolateral defect on radiographs of the humerus.

Adolescent↗

PubMed1999

Establishment of a new cell line (MTT-95) showing basophilic differentiation from the bone marrow of a patient with acute myelogenous leukemia (M7).

A new myeloid cell line, MTT-95, was established from the bone marrow of a patient with acute myelogenous leukemia (AML, M7). MTT-95 cells differentiate into mature basophilic cells in culture medium with no chemical component or cytokine. Surface phenotypes were as follows: CD11b 79.3%, CD13 92.4%, CD33 99.8%, CD34 87.9%, CD41a 77.6% and HLA-DR 0.3%. MTT-95 cells were strongly positive for glycoprotein IIb/IIIa by immunohistochemical staining and revealed metachromatic granules. MTT-95 cells seem to possess characteristics of both megakaryocytes and basophils. These findings suggest that MTT-95 cells are basophil progenitors. MTT-95 cells might be useful in the study not only of the biological aspects of basophils, but also of the diversities of AML (M7).

Antigens, CD↗

PubMed1999

A clinicopathological study of lung cancer patients with occupational exposure to chrysotile asbestos fibers.

OBJECTIVE: To summarize the features of asbestos-related lung cancer. PATIENTS: Thirty-one lung cancer patients with occupational exposure to chrysotile asbestos fibers. They worked or had worked in one asbestos factory or its subcontracters. RESULT: All patients were male with mean age of 60.6 when diagnosed, and all except one were current or ex-'heavy' smokers. Histological types were fairly evenly divided into adeno-, squamous and small cell carcinoma and 24 (78%) of patients showed 'peripheral type' lung cancers. Regarding clinical stages, 20 patients (65%) were classified as III or IV (advanced stage). Tumor shadow(s) was detected on chest X-ray in 22 patients (71%), and in 5 patients with 'negative' chest X-ray, chest CT was necessary to recognize a primary tumor. Seventeen patients (55%) did not undergo periodical check-ups. CONCLUSION: Occupational asbestos exposure is interpreted as one of the important risks for lung cancer and frequent and accurate observation is necessary.

Adult↗

PubMed1999

A substitutional mutation in the DNA binding domain of the androgen receptor causes complete androgen insensitivity syndrome.

DNA analysis of the androgen receptor gene in a patient with complete androgen insensitivity syndrome identified a substitutional mutation (tyrosine converted to cysteine at position 571) in the DNA binding domain. In vitro transfection experiments with the patients' androgen receptor gene, indicated normal expression of the androgen receptor in transfected COS-7 cells compared to the wild type gene. There was also no evidence of impaired thermal stability of the 5 alpha-dihydrotestosterone-androgen receptor complex. However, the capacity of the androgen receptor to activate target gene transcription was found to be completely disrupted in a luciferase assay. These results confirmed that only one substitutional mutation in the DNA binding domain was related to the pathogenesis of the complete androgen insensitivity syndrome.

Adult↗