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Biomedical subjects

S Uchiyama

Publications and source records attributed to S Uchiyama.

At least 19 recordsLinked to original sources
PubMed2002

Localization of group IB phospholipase A(2) isoform in the gills of the red sea bream, Pagrus (Chrysophrys) major.

We previously reported that PLA(2) activity in the gills is higher than that in other tissues in red sea bream and purified PLA(2) from the gills belongs to the group IB PLA(2) as well as other red sea bream PLA(2)s. In this study, we reconfirmed that the level of PLA(2) activity is extremely high in the gills compared with other tissues, and gill PLA(2) was detected only in the gills by immunoblotting and inhibition test using anti-gill PLA(2) monoclonal antibody. The level of PLA(2) activity and protein expression in the gills are well correlated. Fish can be roughly divided into high and low groups based on the level of PLA(2) activity. Gill PLA(2) was detected in the gills of the high group, but not the low group by immunoblotting. In the gills of the high group, gill PLA(2) was detected in the mucous cells and pavement cells located on the surface of gill epithelia by immunohistochemistry. On the other hand, positive signals were observed only in the mucous cells by in situ hybridization. We also isolated inactive proPLA(2), having AR propeptide, preceding the mature enzyme from the gill extract. These results suggest that gill PLA(2) is synthesized as an inactive proPLA(2) in the mucous cells and is secreted to the surface of gill epithelia.

Animals↗
PubMed2002

Postoperative nerve conduction changes after open and endoscopic carpal tunnel release.

OBJECTIVE: To assess the improvement of motor distal latency (MDL), sensory nerve conduction velocity (SCV) of the median nerve and the amplitudes of compound muscle action potential (CMAP) and sensory nerve action potential (SNAP) in patients with idiopathic carpal tunnel syndrome subjected to surgical treatment according to the open carpal tunnel release method and the endoscopic carpal tunnel release. METHODS: Sixty-six hands of sixty-six patients were divided into two groups: the ECTR group and the OCTR group. The patients were evaluated preoperatively, and at 1, 3, 6, and 12 months postoperatively. RESULTS: Although no statistically significant difference of the recovery of MDL and the amplitude of CMAP and SNAP was detected between the two groups at any time point during follow-up, one patient in the ECTR group in whom the operation had been converted to OCTR, showed delay of MDL and decrease in the amplitude of CMAP. CONCLUSIONS: There is a risk of nerve damage in patients undergoing ECTR. Although statistical analysis suggests that nerve conduction improves by about the same degree 12 months after ECTR or OCTR, slightly faster improvement after OCTR cannot be excluded.

Action Potentials↗
PubMed2001

Measurement of platelet fibrinogen binding and p-selectin expression by flow cytometry in patients with cerebral infarction.

We determined the percentages of fibrinogen-bound and P-selectin-expressed platelets in whole blood using flow cytometry in 254 patients with various cerebral infarction subtypes, as well as in 30 age-matched controls. Patients with atherothrombotic stroke showed significant increases in both fibrinogen-bound and P-selectin-expressed platelets. Patients with lacunar stroke also showed significant increases in both of them, but the percentage of P-selectin expression was significantly lower than that in atherothrombotic stroke. Patients with cardioembolic stroke showed a significant increase in P-selectin-expressed platelets without any increase in fibrinogen-bound platelets. Platelet fibrinogen binding and P-selectin expression were significantly lower in patients treated with ticlopidine but not with aspirin than in those not treated with any antiplatelet agent, and were lowest in those treated with both ticlopidine and aspirin. Our findings suggest that expression of adhesion molecules on platelet membrane surface differs among the patients with subtypes of ischemic stroke and differs among the types of antiplatelet regimens.

Adult↗
PubMed2001

A fluorogenic reagent, 4-mercapto-7-methylthio-2,1,3-benzoxadiazole for carboxylic acids, designed by prediction of the fluorescence intensity.

During the course of our studies of the development of fluorogenic reagents having a 4,7-disubstituted benzofurazan structure, we previously proposed 7-acetylamino-4-mercapto-2,1,3-benzoxadiazole (AABD-SH) as a fluorogenic reagent for carboxylic acids. Since then, progress has made it possible to estimate the fluorescence quantum yields of the 4,7-disubstituted benzofurazan compounds on the basis of the PM3 calculation of their S1-T2 energies. Subsequently, a new fluorogenic reagent, 4-mercapto-7-methylthio-2,1,3-benzoxadiazole (MTBDSH) was designed and synthesized. In the presence of condensation reagents, triphenylphosphine (TPP) and 2,2'-dipyridyl disulfide (DPDS), MTBD-SH readily reacted with n-caprylic acid within 1 min at room temperature. The derivatives of five carboxylic acids (n-caprylic acid, n-capric acid, lauric acid, myristic acid, and palmitic acid) were well-separated on a reversed-phase column and were fluorimetrically detected at 519 nm with excitation at 391 nm. The detection limits (S/N = 3) were 2.4-5.0 fmol. Thus, MTBD-SH had properties that were considered to be superior. For carboxylic acids, itwas superior not only to AABD-SH, but also to many other conventional reagents. The superiority was examined in terms of its reactivity and sensitivity and the avoidance of interfering peaks that were derived from the reagent itself or degradation products in the chromatogram.

Carboxylic Acids↗
PubMed2001

Solution structure of the ribosome recycling factor from Aquifex aeolicus.

The solution structure of ribosome recycling factor (RRF) from hyperthermophilic bacterium, Aquifex aeolicus, was determined by heteronuclear multidimensional NMR spectroscopy. Fifteen structures were calculated using restraints derived from NOE, J-coupling, and T1/T2 anisotropies. The resulting structure has an overall L-shaped conformation with two domains and is similar to that of a tRNA molecule. The domain I (corresponding to the anticodon stem of tRNA) is a rigid three alpha-helix bundle. Being slightly different from usual coiled-coil arrangements, each helix of domain I is not twisted but straight and parallel to the main axis. The domain II (corresponding to the portion with the CCA end of tRNA) is an alpha/beta domain with an alpha-helix and two beta-sheets, that has some flexible regions. The backbone atomic root-mean-square deviation (rmsd) values of both domains were 0.7 A when calculated separately, which is smaller than that of the molecule as a whole (1.4 A). Measurement of 15N-[1H] NOE values show that the residues in the corner of the L-shaped molecule are undergoing fast internal motion. These results indicate that the joint region between two domains contributes to the fluctuation in the orientation of two domains. Thus, it was shown that RRF remains the tRNA mimicry in solution where it functions.

Amino Acid Sequence↗
PubMed2001

A compactly integrated flow cell with a chemiluminescent FIA system for determining lactate concentration in serum.

We have fabricated an integrated flow cell as a total microanalysis system (microTAS). This flow cell (size, 15 x 20 mm; total inner volume, 12.2 microL) was designed for a rational analyzing system of lactate determination for serum. This cell was made by micromachining techniques and consisted of two hollows of a lactate oxidase (LOD) reactor and a mixing cell, a spiral groove, and three penetrated holes. To form the reactor and capillary, these patterns, etched on a silicon wafer, were attached to a glass plate by the anodic bonding method. A photodiode was put under part of the spiral capillary. The compactly accumulated devices were integrated into a flow injection analysis (FIA) system. In the flow cell, lactate was catalyzed to pyruvate and hydrogen peroxide at the LOD reactor; subsequently, hydrogen peroxide reacted with the luminol-ferricyanic reagent at the mixing cell. The resulting chemiluminescent light was detected by the photodiode. Using the miniaturized flow cell, the sample volume for one measurement was greatly reduced to 0.2 microL. The response to lactate was obtained within 30 s and was linear between 0.5 and 5.0 mM (4.5 and 45 mg/dL) lactate with excellent correlative variances of 3.2% (average of three measurements at 5.0 mM). For practical application, the lactate concentration in control human serum was determined using this system. The results showed a good correlation coefficient (r = 0.979) with the results obtained by the spectrophotometric reference method. No difference in sera (normal or pathological) was found. Consequently, this integrated flow cell shows potential as a clinical device for lactate determination in serum. In this article, the effect of the design on the chemiluminescent FIA system is also described.

Enzymes, Immobilized↗
PubMed2001

Fluorogenic and fluorescent labeling reagents with a benzofurazan skeleton.

Fluorogenic and fluorescent labeling reagents having a benzofurazan (2,1,3-benzoxadiazole) skeleton such as 4-fluoro-7-nitro-2,1,3-benzoxadiazole (NBD-F), 4-N,N-dimethylaminosulfonyl-7-fluoro-2,1,3-benzoxadiazole (DBD-F), 4-aminosulfonyl-7-fluoro-2,1,3-benzoxadiazole (ABD-F), ammonium 7-fluoro-2,1,3-benzoxadiazole-4-sulfonate (SBD-F), 4-hydrazino-7-nitro-2,1,3-benzoxadiazole (NBD-H), 4-N,N-dimethylaminosulfonyl-7-hydrazino-2,1,3-benzoxadiazole (DBD-H), 4-nitro-7-N-piperazino-2,1,3-benzoxadiazole (NBD-PZ), 4-N,N-dimethylaminosulfonyl-7-N-piperazino-2,1,3-benzoxadiazole (DBD-PZ), 4-(N-chloroformylmethyl-N-methyl)amino-7-N,N-dimethylaminosulfonyl-2,1,3-benzoxadiazole (DBD-COCl) and 7-N,N-dimethylaminosulfonyl-4-(2,1,3-benzoxadiazolyl) isothiocyanate (DBD-NCS) are reviewed in terms of synthetic method, reactivity, fluorescence characteristics, sensitivity and application to analytes.

Benzoxazoles↗
PubMed2001

Three-dimensional evaluation of lumbar disc hernia and prediction of absorption by enhanced MRI.

Both the spontaneous shrinkage and the disappearance of disc hernia have been confirmed through the use of computed tomography (CT) and magnetic resonance imaging (MRI). There is, however, no practical method to predict the likely absorption of the herniated mass. The objective of this study was to predict the spontaneous absorption of disc hernia by MRI, and to select the optimum treatment. The study involved 65 patients with lumbar disc hernias. Conservative treatment was carried out in 21 patients, while 44 patients underwent herniotomy. In the nonoperated patients, an MRI was taken both during the painful period, and shortly after pain remission. Hernial shrinkage was evaluated according to the decrease in the calculated volume, in addition to the decrease in hernial area, calculated by MRI. In the operated group, preoperative MRI enhancement, type of hernia, and invasion of granulation tissue in the histological specimens were studied. In the 21 nonoperated patients, the volume (mean +/- SD) was 0.488 +/- 208 cm(3) (range, 0.197-0.931 cm(3)) in the painful period and 0.214 +/- 0.181 cm(3) (range, 0.0-0.744 cm(3)) in the remission period. This decrease in volume was statistically significant. There was also a greater decrease in hernias exhibiting positive enhancement by MRI. In the operated patients, hernias that penetrated the posterior longitudinal ligament (PLL) had high rates of preoperative enhancement, and these hernias showed invasion of granulation tissue with marked neovascularization. Positive enhancement by MRI confirms an ongoing absorption process. Enhanced MRI can be a good method for the prediction of spontaneous absorption of lumbar disc hernias.

Absorption↗
PubMed2001

Cloning and expression of group IB phospholipase A2 isoforms in the red sea bream, Pagrus major.

Two cDNA encoding red sea bream DE-1 and DE-2 phospholipases A2 (PLA2) were cloned from the hepatopancreas of red sea bream, Pagrus (Chrysophrys) major. The cDNA of DE-1 PLA2 encoded a mature protein of 125 amino acid residues with an apparent signal peptide of 20 residues and propeptide of 5 residues, and that of DE-2 PLA2, a mature protein of 126 amino acid residues with an apparent signal peptide of 17 residues and propeptide of 6 residues. Comparison of the predicted amino acid sequences for mature DE-1 and DE-2 PLA2 showed that both proteins contain 14 cysteines including Cys 11 and 77 and a pancreatic loop, which are commonly conserved in group IB PLA2; however, the identity in amino acid sequence between DE-1 and DE-2 PLA2 was low (47%). A previous report concerning the cDNA cloning of red sea bream gill G-3 PLA2 and the present results represent the first cloning and sequencing of three distinct isoforms of group IB PLA2 in a single fish species, red sea bream. Reverse transcription-polymerase chain reaction analysis showed that DE-1 PLA2 mRNA was expressed in the hepatopancreas, pyloric ceca, intestine, spleen, gonad, stomach, and kidney, whereas gill G-3 PLA2 mRNA was expressed only in the gills and gonad. The expression of DE-2 PLA2 mRNA was detected in all of the tissues analyzed. These results indicate that three distinct isoforms of group IB PLA2, DE-1 and DE-2 PLA2 in hepatopanceas and gill G-3 PLA2, are expressed in a tissue-specific manner in red sea bream.

Amino Acid Sequence↗
PubMed2001

Anabolic steroids increase exercise tolerance.

The influence of an anabolic androgenic steroid (AAS) on thymidine and amino acid uptake in rat hindlimb skeletal muscles during 14 days after a single exhaustive bout of weight lifting was determined. Adult male rats were divided randomly into Control or Steroid groups. Nandrolone decanoate was administered to the Steroid group 1 wk before the exercise bout. [3H]thymidine and [14C]leucine labeling were used to determine the serial changes in cellular mitotic activity, amino acid uptake, and myosin synthesis. Serum creatine kinase (CK) activity, used as a measure of muscle damage, increased 30 and 60 min after exercise in both groups. The total amount of weight lifted was higher, whereas CK levels were lower in Steroid than in Control rats. [3H]thymidine uptake peaked 2 days after exercise in both groups and was 90% higher in Control than in Steroid rats, reflecting a higher level of muscle damage. [14C]leucine uptake was approximately 80% higher at rest and recovered 33% faster postexercise in Steroid than in Control rats. In a separate group of rats, the in situ isometric mechanical properties of the plantaris muscle were determined. The only significant difference was a higher fatigue resistance in the Steroid compared with the Control group. Combined, these results indicate that AAS treatment 1) ameliorates CK efflux and the uptake of [3H]thymidine and enhances the rate of protein synthesis during recovery after a bout of weight lifting, all being consistent with there being less muscle damage, and 2) enhances in vivo work capacity and the in situ fatigue resistance of a primary plantarflexor muscle.

Anabolic Agents↗
PubMed2001

Structural and thermodynamic behavior of eukaryotic initiation factor 4E in supramolecular formation with 4E-binding protein 1 and mRNA cap analogue, studied by spectroscopic methods.

The structural and thermodynamic behavior of the complex formation of eIF4E with either or both mRNA cap analogue (m7GTP, m7GpppA, or m7GpppG) and 4EBP1 has been investigated by spectroscopic measurements. Although the circular dichroism (CD) spectrum of eIF4E was little affected by the association with any cap analogue, the association constant of eIF4E with m7GpppA/G, estimated from the fluorescence quenching, was about 10 times larger than that with m7GTP. The van't Hoff analyses showed that the m7GpppA/G binding is enthalpy-driven with a large negative deltaH(o), and this is in contrast with the entropy-driven binding of m7GTP, where the positive deltaS(o) is large enough to overcome an increase of deltaH(o). This different behavior obviously originates in the interaction of the second nucleotide in m7GpppA with eIF4E, suggesting the importance of the nucleotide sequence linked to the m7Gppp terminal moiety, in addition to the specific interaction with the m7G base, for the recognition of mRNA cap structure by eIF4E. On the other hand, the CD spectra indicated that the binding of 4EBP1, an endogenous eIF4E-regulatory protein without having any defined secondary structure, shifted the m7GTP- or m7GpppA/G-bound eIF4E to an irregular structure, although such a structural change was not observed for eIF4E alone. The association constant of 4EBP1 with m7GTP- or m7GpppA/G-bound eIF4E was by two orders of magnitude larger than that with eIF4E alone. These results suggest the close interrelation in the supramolecular formation of 4EBP-eIF4E-mRNA cap structure.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Difference in the occurrence of cardiovascular events according to class of antihypertensive agent, based on a follow-up study of Japanese hypertension patients.

We conducted a nested case-control study to evaluate the relationship between antihypertensive agent class and the incidence of initial cardiovascular events. A total of 7.443 patients being treated with an antihypertensive agent in 1985-1986 were enrolled for follow-up of up to 5 years. A total of 362 patients (186 males and 176 females) developed cardiovascular events. Age (5-year interval) and sex-matched controls were then randomly selected. A Multiple logistic regression analysis was done to control for the effects of confounding factors. The results showed that the use of diuretics and beta-blockers was associated with a reduced risk of cardiovascular events (odds ratio [OR] =0.65, 95% confidence interval [CI]: 0.49-0.86, and OR=0.75, 95% CI: 0.56-1.02, respectively), against a significantly raised risk associated with the use of calcium antagonists (OR=1.34, 95% CI: 1.03-1.80). However, as far as stroke was concerned, there was no significant association of risk with the use of any agent. The control group was found to be similar to the case group with respect to the changes in the treatment program during the follow-up period. The results suggest that the calcium antagonists used in Japan during the period of 1985-1990 constituted a potential risk for the occurrence of cardiovascular events.

Adrenergic beta-Antagonists↗
PubMed2000

Selected mutations in a mesophilic cytochrome c confer the stability of a thermophilic counterpart.

Mesophilic cytochrome c(551) of Pseudomonas aeruginosa (PA c(551)) became as stable as its thermophilic counterpart, Hydrogenobacter thermophilus cytochrome c(552) (HT c(552)), through only five amino acid substitutions. The five residues, distributed in three spatially separated regions, were selected and mutated with reference to the corresponding residues in HT c(552) through careful structure comparison. Thermodynamic analysis indicated that the stability of the quintuple mutant of PA c(551) could be partly attained through an enthalpic factor. The solution structure of the mutant showed that, as in HT c(552), there were tighter side chain packings in the mutated regions. Furthermore, the mutant had an increased total accessible surface area, resulting in great negative hydration free energy. Our results provide a novel example of protein stabilization in that limited amino acid substitutions can confer the overall stability of a natural highly thermophilic protein upon a mesophilic molecule.

Bacteria↗
PubMed2000

Structural basis of the interaction between IgG and Fcgamma receptors.

The binding of multivalent antigen-antibody complexes to receptors for the Fc portion of IgG (FcgammaR) induces the clustering of the FcgammaR and triggers cell activation leading to defence reactions against pathogens. The Fc portion of IgG consists of two identical polypeptide chains which are related to each other by a 2-fold axis and are folded in two structural domains, the C(H)2 domain, near the flexible hinge region of the IgG molecule, and the C(H)3 domain. We studied the interaction in solution between the Fc fragment of mouse IgG2b and the extracellular region of mouse FcgammaRII. We find that one Fc molecule binds one FcgammaRII molecule only. Using NMR spectroscopy, we show that FcgammaRII binds to a negatively charged area of the C(H)2 domain, corresponding to the lower hinge region, and that the binding of FcgammaRII onto one of the two symmetrically related sites on the Fc induces a conformational change in the other site. We therefore propose a model that explains why IgG molecules are unable to trigger FcgammaR-mediated cellular responses spontaneously in the absence of crosslinking by multivalent antigens.

Animals↗