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Biomedical subjects

S Shinoda

Publications and source records attributed to S Shinoda.

At least 19 recordsLinked to original sources
PubMed2003

Enhancement of thymidine kinase-mediated killing of malignant glioma by BimS, a BH3-only cell death activator.

Herpes simplex virus thymidine kinase (HSV-tk)/gancyclovir (GCV) therapy has the ability to inhibit tumor formation in animal models but the results of clinical trials have been disappointing. To improve the performance of tk/GCV therapy, we tried combination therapy designed to enhance its cytotoxic effects by introducing genes that induce apoptosis of the tumor cells through different pathways. We concentrated our efforts on the use of Bim, a BH3-only member of death activators in the Bcl-2 superfamily, because Bim is not involved in the pathways through which HSV-tk/GCV therapy induces apoptosis in malignant glioma cells. Among three alternative splicing variants, BimEL, BimL, and BimS, BimS lacks the binding domain for the dynein light chain LC8, which negatively regulates the proapoptotic function of BimEL and BimL. All four malignant glioma cell lines, U251, A172, T-430, and U373 underwent cell death after transfer of BimS using an adenovirus vector (AVC2). Intriguingly, combination of AVC2-BimS with AVC2-tk markedly increased the sensitivity of U251 cells to GCV both in vitro and in vivo. In contrast, AVC2-BimL did not induce significant cell death. These results indicated that BimS had the ability to improve the efficiency of HSV-tk/GCV therapy in the treatment of malignant glioma and suggested that the targeting of different proapoptotic pathways may be a useful strategy for the development of an effective gene therapy approach to treatment.

Adenoviridae↗
PubMed2002

Predicting pesticide concentrations in river water with a hydrologically calibrated basin-scale runoff model.

Hydrological diffuse pollution models require calibration before they can be used to make accurate long-term predictions for a range of hydrological and meteorological conditions. As such, the applicability of the models to the dispersion of new pesticides is limited due to the lack of calibration data. In this study, the performance of a GIS-based basin-scale runoff model for predicting the concentrations of paddy-farming pesticides in river water was examined when calibrated using hydrological data alone, without optimization based on empirical pesticide concentration data. The prediction accuracy on a daily or hourly scale was somewhat unsatisfactory due to inevitable compromises concerning rice farming schedules. However, the month-averaged pesticide concentrations were satisfactorily accurate; more than 50% of predicted values were between half and twice the observed values, considering the deficiencies of the input data, particularly for pesticide usage, which may include up to 50% error.

Calibration↗
PubMed2001

Ester-armed cyclens having quadruplicated helical geometry: remarkably stable and selective encapsulation of Na+ ion.

A new series of ester-armed cyclens nicely accommodated a Na+ ion in their quadruplicated helical binding spheres and effectively discriminated the cation from Li+ and K+ ions. Crystallographic studies revealed that four ester-functionalized sidearms provided effective coordination with the Na+ ion trapped in the 12-membered cyclen ring. Log K values for their Na+ complexes were estimated as 9-11 in CD3CN or C2D5OD, which were comparable to those of common bicyclic cryptands. FAB-MS, liquid-liquid extraction, and NMR binding experiments demonstrated that the cooperative action of the parent cyclen ring and ester-functionalized sidearms offered stable and selective encapsulation of the Na+ ion based on unique quadruplicated helical geometry.

Journal Article↗
PubMed2001

Specific recognition of chiral amino alcohols via lanthanide coordination chemistry: structural optimization of lanthanide tris(beta-diketonates) toward effective circular dichroism/fluorescence probing.

Lanthanide tris(beta-diketonates) formed stable, 1:1 highly coordinated complexes with amino alcohols, and the resulting complexes exhibited large enhanced fluorescence and intense induced circular dichroism (CD) signals. The stability constants of the highly coordinated complexes were determined for various combinations of lanthanide centers, beta-diketonate ligands, and organic substrates. These revealed that amino alcohol coordinated with the lanthanide center much more strongly than monoamine, monoalcohol, or diol derivative. On the basis of the highly coordinated complexation, several lanthanide tris(beta-diketonates) acted as CD/fluorescence probes specific for amino alcohols. Tris(6,6,7,7,8,8,8-heptafluoro-2,2-dimethyl-3,5-octadionato)europium(III) showed enhanced fluorescence in the presence of amino alcohols, while the corresponding ytterbium complex exhibited chirality-dependent CD signals for amino alcohols. In particular, the observed CD spectral profiles related well with the absolute configuration and optical purity of the bound amino alcohol, indicating that the structural optimization of lanthanide tris(beta-diketonates) offered specific sensing of amino alcohols and precise determination of their enantiomer excess percentages.

Journal Article↗
PubMed2001

The C-terminal domain promotes the hemorrhagic damage caused by Vibrio vulnificus metalloprotease.

Vibrio vulnificus, an opportunistic human pathogen, produces a 45-kDa zinc metalloprotease (V. vulnificus protease; VVP) as an important virulence determinant. VVP injected intradermally into the dorsal skin causes the hemorrhagic damage through specific degradation of type IV collage in the vascular basement membrane. The N-terminal 35-kDa polypeptide (VVP-N), the catalytic domain, also evoked the hemorrhagic skin reaction within minutes. However, the hemorrhagic activity of VVP-N was one-third of that of VVP. Besides, the proteolytic activity of VVP-N toward the reconstituted basement membrane or type IV collagen was found to be about 50 % of VVP. VVP-N, like VVP, was quickly inactivated by an equimolar amount of alpha(2)-macroglobulin, a broad-spectrum plasma protease inhibitor. These findings indicate that the C-terminal 10-kDa polypeptide, the substrate-binding domain mediating the effective binding to protein substrates, functions to augment the hemorrhagic reaction of VVP.

Amino Acid Sequence↗
PubMed2001

Identification of beta-lactoglobulin-derived peptides and class II HLA molecules recognized by T cells from patients with milk allergy.

BACKGROUND: Cow's milk allergy impairs the health and development of many infants since it deprives them of adequate nutrition. Cow's milk fractions contain many allergens, and beta-lactoglobulin (BLG) is one of the major allergens. OBJECTIVE: The purpose of this study was to determine T cell epitopes, antigen-presenting molecules and cytokine production by T cells in relation to BLG. The results can provide new therapeutic possibilities of using analogue peptides of BLG for infants with cow's milk allergy. METHODS: Using a mixture of a panel of overlapping synthetic peptides that cover the entire BLG molecule, we established polyclonal BLG-specific short-term T cell lines and clones from peripheral blood mononuclear cells of four patients with allergy to cow's milk carrying most of the common human leucocyte antigen (HLA) haplotypes seen in the Japanese population. We then identified the T cell epitopes and antigen-presenting molecules, and measured the production of cytokines interleukin (IL)-4, IL-5 and interferon-gamma in the culture supernatants. RESULTS: The T cell lines established from the four patients responded to seven different peptides. Three of the peptides stimulated the T cells of two donors, regardless of the HLA types. The patterns of inhibition of the proliferative responses of the cell lines by anti-HLA class II antibodies were heterogeneous; three were mainly inhibited by anti-HLA-DR mAbs, and the other was inhibited by anti-HLA-DQ mAbs. High levels of IL-5 were produced by these T cell lines. CONCLUSIONS: Patients' T cells recognized BLG in association with a variety of HLA-DR or -DQ as antigen-presenting molecules. Although some peptides did have a more potent T cell stimulatory activity than others, the T cell receptor ligands formed with the BLG molecule are heterogeneous. Peptides for the desensitization of T cells of the patients with cow's milk allergy need to be designed keeping in mind the different requirements in different ethnic groups.

Amino Acid Sequence↗
PubMed2001

Functional evaluation of cytochrome P450 2D6 with Gly42Arg substitution expressed in Saccharomyces cerevisiae.

A single amino acid-substituted mutant protein, CYP2D6 (G42R) was expressed in Saccharomyces cerevisiae and its enzymatic properties were compared with those of other single (P34S, R296C and S486T) and double amino acid-substituted mutant proteins (P34S/S486T and R296C/S486T) expressed in yeast cells, all of which were known to occur in the CYP2D6 gene as single nucleotide polymorphisms. The protein levels of G42R, P34S and P34S/S486T in microsomal fractions and their oxidation capacities towards debrisoquine as a prototypic substrate and bunitrolol as a chiral substrate were different from those of wild-type CYP2D6, while the R296C, S486T and R296C/S486T behaved similarly to the wild-type in these indices. The CYP contents both in yeast microsomal and in whole cell fractions indicated that some part of G42R protein was localized in the endoplasmic reticulum membrane fraction, whereas most of G42R protein was in some subcellular fractions other than endoplasmic reticulum. In kinetic analysis, the G42R substitution increased apparent Km and decreased Vmax for debrisoquine 4-hydroxylation, while it increased both Km and Vmax for bunitrolol 4-hydroxylation. The P34S substitution did not drastically change Km but decreased Vmax for debrisoquine 4-hydroxylation, whereas Km was increased and Vmax unchanged or decreased for bunitrolol 4-hydroxylation by P34S substitution. These results suggest that the G42R substitution causes a change in the CYP2D6 conformation, which may be different from the change produced by the P34S substitution.

Amino Acid Substitution↗
PubMed2001

Detection of virulence associated genes in clinical strains of vibrio mimicus.

A total of 42 clinical strains of Vibrio mimicus were examined for the presence of virulence associated genes toxR, toxS, toxT, tcpP, ctx and tcpA by PCR assay. Almost all strains were shown to have the toxR gene, while the toxS gene was found in 27 strains. On the other hand, five strains possessed both toxT and tcpP genes, but others had neither. Only two strains were positive for amplification of the ctx gene, whereas no PCR product with tcpA primers was detected. The results indicate the incomplete copies of virulence cascade in V mimicus strains. The pathogenesis and epidemic potential of this species is also discussed.

Bacterial Outer Membrane Proteins↗
PubMed2001

Analysis of seawaters for the recovery of culturable Vibrio parahaemolyticus and some other vibrios.

We investigated the recovery of dormant and injured cells along with the normally culturable cells of Vibrio species with special emphasis on V. parahaemolyticus using both selective and non-selective media at moderate (20 C) and standard (37 C) culture temperatures from a bay water environment. Culture temperatures (20 or 37 C) did not affect the recovery of V. parahaemolyticus but did for other vibrios. We observed similar seasonality of V parahaemolyticus as in most other environmental studies. V. parahaemolyticus and other Vibrio species were recovered in higher numbers by a replica plating method compared to most probable number (MPN) and direct TCBS (thiosulfate citrate bile-salt sucrose) agar counts. Even with the replica plating method, however, vibrios number goes down to a minimum level and V. parahaemolyticus was undetectable during the cool temperature period of the year, although total bacterial cells and CFU on nutrient agar (with 2% NaCl) did not vary so much during the study period.

Colony Count, Microbial↗
PubMed2001

[QOL questionnaire version 2001 for pediatric patients with bronchial asthma and their parents or caregivers--preparation and evaluation].

We already reported that the QOL questionnaire for pediatric patients with bronchial asthma and their parents or caregivers reflects reliability (including test retest), factorial validity, and changes in paroxysmal attacks of asthma. In this study, we revised the questionnaire for use in routine medical care. We investigated the importance of questions for affected children and their parents or caregivers and selected and discarded questions on the basis of average ranks of the importance and medical standpoint so that the questionnaire might have physical and emotional domains. The QOL questionnaire version 2001 for pediatric patients with bronchial asthma and their parents or caregivers (hereafter to be abbreviated as the version 2001) includes 15 questions for patients under the age of 4 years and 20 questions for patients over the age of 4 years and 4 year-old. The "present" and "absent" groups, which were categorized with respect to the events (e.g., attack) that developed in 2 weeks prior to the QOL investigation, showed a significant difference (Mann-Whitney U-test) in terms of the total score of replies which constitute each domain, i.e., physical domain (p = 0.0052) in patients under the age of 4 years and physical domain (p < 0.0001) and emotional domain (p < 0.0001) in patients over the age of 4 years and 4 year-old. Regarding the average value of each reply which constitutes the domain as well, the physical domain showed a decrease in symptom instability and the "> or = 4 years" group showed a decrease in exertional load and mental load. Therefore, a QOL investigation like ours which investigates disorders in which attacks develop suddenly, e.g., asthma, should examine the presence or absence of all events prior to the investigation. The version 2001 was considered useful for affected children and their parents or caregivers to comprehend the content and severity of impairments which were provoked by asthma.

Asthma↗
PubMed2000

Supramolecular complex of cytochrome c with lariat ether: solubilization, redox behavior and catalytic activity of cytochrome c in methanol.

A variety of lariat ethers were employed to solubilize water-soluble cytochrome c in methanol, in which alcohol, ether, ester, amine, and amide functionalities were attached as cation-ligating side arms to 18-crown-6, 15-crown-5, and 12-crown-4 rings. Among these lariat ethers, the alcohol-armed 18-crown-6 derivative offered the highest solubilization efficiency for cytochrome c via supramolecular complexation. The resulting cytochrome c-lariat ether complexes were electrochemically and spectroscopically characterized and confirmed to have redox-active heme structures of 6-coordinate low-spin population in methanol. Some of them catalyzed the oxidation of pinacyanol chloride with hydrogen peroxide in methanol and exhibited higher activities than unmodified cytochrome c and its poly(ethylene glycolated) derivative. Since the supramolecular complexation between lariat ether and cytochrome c includes extremely simple procedures, it provides a facile preparation method of effective biocatalysts working in organic solvents from metalloproteins.

Carbocyanines↗
PubMed2000

Autonomic regulation after exercise evidenced by spectral analysis of heart rate variability in asthmatic children.

BACKGROUND: Bronchial asthma is associated with abnormal autonomic nervous function in childhood. Exercise is one of the most common precipitating factors of acute asthmatic crises although the exact mechanism of autonomic regulation in asthmatic children after exercise is unclear. OBJECTIVE: The aim of this study was to investigate the features of autonomic regulation after exercise in asthmatic and control children. METHODS: Pulmonary function tests and heart rate variability spectral analysis were performed in 15 asthmatic children and 7 control children (age 6 to 15 years) during and after an exercise challenge. RESULTS: The maximum % fall of forced expiratory volume in 1 second (FEV1) was significantly greater (P < .01) in asthmatic subjects (9.1 +/- 5.1%) than in normal control subjects (1.0 +/- 2.5%). The high frequency band (HF) amplitude, an index of cardiac vagal tone, 5 minutes after exercise was significantly higher (P < .05) in the asthmatic subjects (14.4 +/- 7.9 msec) than in control subjects (5.9 +/- 2.6 msec). Furthermore, the difference in the HF amplitude between the control group and the exercise-induced asthma group was significant both 5 minutes (P < .01) and 10 minutes (P < .05) after challenge. There was a significant correlation (P = .565, P = .0165) between HF amplitude 5 minutes after exercise and the magnitude of the decrease in FEV1. On the other hand, no significant difference was observed in the low frequency band amplitude between the controls and the asthmatic subjects. The ratio of low frequency to high frequency power, which is suggested to correlate with cardiac sympathetic activity, did not differ between the two groups. CONCLUSION: These findings suggest that autonomic nervous activities, particularly vagal response after exercise, in asthmatic children is different from that in control children.

Adolescent↗
PubMed2000

Microbial metalloproteases and pathogenesis.

Zinc metalloproteases produced by human pathogenic microorganisms show a wide variety of pathological actions. In local infections, the proteases cause necrotic or hemorrhagic tissue damage through digestion of structural components of the ground substance, and also form edematous lesions through generation of inflammatory mediators, while in systemic infections, the proteases act as a synergistic virulence factor through disordered proteolysis of many plasma proteins. Clostridial neurotoxins, Bacteroides fragilis enterotoxin and Bacillus anthracis lethal factor are also zinc metalloproteases.

Animals↗
PubMed2000

Analysis of Vibrio mimicus clinical strains by arbitrarily primed polymerase chain reaction.

A total of 51 Vibrio mimicus clinical strains from different geographic locations were examined by arbitrarily primed polymerase chain reaction (AP-PCR). The primer VMH-3 divided them into 28 groups, although 18 groups consisted of a single strain at present. All groups had a common 1.0-kb amplification fragment. Most of the groups consisted of strains from same region, although two exceptional groups showed a few amplification fragments including strains from different regions. AP-PCR groups were not consistently associated with serogroups. AP-PCR is thought to be a valuable and easy method for the epidemiological study of V. mimicus.

DNA Primers↗
PubMed2000

Characterization of Vibrio parahaemolyticus manganese-resistant mutants in reference to the function of the ferric uptake regulatory protein.

In many bacteria, the ferric uptake regulatory protein (Fur) has a central role in the negative regulation of genes affected by iron limitation. In this study, Vibrio parahaemolyticus strains carrying mutations in the fur gene encoding Fur were isolated by the manganese selection method to assess the function of Fur in connection with alternations in the coordinate expression of the siderophore vibrioferrin (VF) and iron-repressible outer membrane proteins (IROMPs). Ten out of 25 manganese-resistant mutants constitutively produced VF and expressed at least two IROMPs irrespective of the iron concentration in the medium. PCR-direct DNA sequencing of the fur genes in these mutants identified four different point mutations causing amino acid changes. Moreover, a fur overexpressing plasmid was constructed to prepare antiserum against V. parahaemolyticus Fur. Western blotting with this antiserum revealed that the intracellular abundance of the wild-type Fur was not significantly affected by the iron concentrations in the growth medium, and that the Fur proteins of the mutant strains occurred at substantially smaller amounts and/or migrated more rapidly in sodium dodecyl sulfate-polyacrylamide gel electrophoresis than the wild-type Fur. These data afford an additional insight into the structure-function relationship of Fur and imply its involvement in the iron acquisition systems of V. parahaemolyticus, although it is yet unknown whether its action on the target genes is direct or indirect.

Bacterial Proteins↗
PubMed2000

Cloning and characterization of the ddc homolog encoding L-2,4-diaminobutyrate decarboxylase in Enterobacter aerogenes.

L-2,4-diaminobutyrate decarboxylase (DABA DC) catalyzes the formation of 1,3-diaminopropane (DAP) from DABA. In the present study, the ddc gene encoding DABA DC from Enterobacter aerogenes ATCC 13048 was cloned and characterized. Determination of the nucleotide sequence revealed an open reading frame of 1470 bp encoding a 53659-Da protein of 490 amino acids, whose deduced NH2-terminal sequence was identical to that of purified DABA DC from E. aerogenes. The deduced amino acid sequence was highly similar to those of Acinetobacter baumannii and Haemophilus influenzae DABA DCs encoded by the ddc genes. The lysine-307 of the E. aerogenes DABA DC was identified as the pyridoxal 5'-phosphate binding residue by site-directed mutagenesis. Furthermore, PCR analysis revealed the distribution of E. aerogenes ddc homologs in some other species of Enterobacteriaceae. Such a relatively wide occurrence of the ddc homologs implies biological significance of DABA DC and its product DAP.

Amino Acid Sequence↗