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Biomedical subjects

T Yagi

Publications and source records attributed to T Yagi.

At least 181 records · Page 10Linked to original sources
PubMed1999

Removal of the bridging ligand atom at the Ni-Fe active site of [NiFe] hydrogenase upon reduction with H2, as revealed by X-ray structure analysis at 1.4 A resolution.

BACKGROUND: The active site of [NiFe] hydrogenase, a heterodimeric protein, is suggested to be a binuclear Ni-Fe complex having three diatomic ligands to the Fe atom and three bridging ligands between the Fe and Ni atoms in the oxidized form of the enzyme. Two of the bridging ligands are thiolate sidechains of cysteinyl residues of the large subunit, but the third bridging ligand was assigned as a non-protein monatomic sulfur species in Desulfovibrio vulgaris Miyazaki F hydrogenase. RESULTS: The X-ray crystal structure of the reduced form of D. vulgaris Miyazaki F [NiFe] hydrogenase has been solved at 1.4 A resolution and refined to a crystallographic R factor of 21.8%. The overall structure is very similar to that of the oxidized form, with the exception that the third monatomic bridge observed at the Ni-Fe site in the oxidized enzyme is absent, leaving this site unoccupied in the reduced form. CONCLUSIONS: The unusual ligand structure found in the oxidized form of D. vulgaris Miyazaki F [NiFe] hydrogenase was confirmed in the reduced form of the enzyme, with the exception that the electron density assigned to the monatomic sulfur bridge had almost disappeared. On the basis of this finding, as well as the observation that H2S is liberated from the oxidized enzyme under an atmosphere of H2 in the presence of its electron carrier, it was postulated that the monatomic sulfur bridge must be removed for the enzyme to be activated. A possible mechanism for the catalytic action of the hydrogenase is proposed.

Binding Sites↗
PubMed1999

Jejunal pouch to avoid stricture after esophagojejunostomy with circular stapler.

BACKGROUND: Anastomotic stricture is one of the most common problems in esophagojejunostomy using the end-to-end anastomosing (EEA) instrument (Auto Suture Co, Norwalk, CT) after total gastrectomy. To alleviate the stricture, several methods, such as incision to the scar, balloon dilatation, and steroid injection are available. To avoid stricture, the jejunal pouch may allow use of a larger EEA than Roux-en-Y (ReY) reconstruction does. STUDY DESIGN: A total of 45 patients underwent curative total gastrectomy and esophagojejunostomy with jejunal pouch construction (27 patients) or ReY (18 patients), using the EEA. The effects of jejunal pouch construction with a large EEA on avoidance of stricture and benefit to nutritional status were investigated by comparing it with the ReY in terms of postoperative morbidity, postprandial symptoms, and nutritional parameters (serum protein, serum albumin, body weight). RESULTS: EEA28 or larger could be used in 25 patients in the pouch group and 8 patients in the ReY group (p < 0.05). Stricture developed in one patient in the pouch group and in four patients in the ReY group (p < 0.05). Postprandial symptoms were experienced less frequently (p < 0.05) in the pouch group than in the ReY group. When stricture and symptoms were analyzed according to the size of EEA, they occurred more frequently (p < 0.05) in the patients with EEA25 than those with EEA28 or EEA31. No significant differences were evident in nutritional parameters. CONCLUSIONS: The choice of jejunal pouch technique allowed the use of a larger EEA than that of ReY reconstruction, resulting in avoidance of anastomotic stricture and postprandial symptoms, though little benefit in nutritional status was evident to the patients after total gastrectomy.

Anastomosis, Roux-en-Y↗
PubMed1999

Enrichment and efficient screening of ES cells containing a targeted mutation: the use of DT-A gene with the polyadenylation signal as a negative selection maker.

Gene targeting in embryonic stem (ES) cells via homologous recombination can occur at very low frequency. In order to enrich homologous recombinants before screening, a negative selection marker, such as thymidine kinase (TK) and diphtheria toxin A fragment (DT-A), has been commonly used. In this study, we developed a negative selection marker using DT-A gene with polyadenylation signal and it was designated DT-ApA. To determine the difference in targeting efficiency of the negative selections, we constructed three different targeting vectors for each negative selection (first, TK at the 3' end, second, TK at both the 5' and 3' ends < 2 X TK >, and third, DT-ApA at the 5' end of the homologous sequences). Gene targeting experiments using these constructs clearly showed that negative selection using DT-ApA was more efficient than that using TK for homologous recombination and that negative selection using DT-ApA was as efficient as that using 2 X TK. Considering the fact that the use of DT-ApA is more convenient for construction of targeting vectors than that of 2 X TK, DT-ApA is an efficient negative selection marker. In addition, we examined long and accurate PCR (LA-PCR) for screening gene targeted clones. The use of LA-PCR with genomic DNAs from ES cell clones facilitated simple detection of homologous recombinants, suggesting that the screening with LA-PCR is compatible with the use of longer homologous sequences of both arms in vector design. Our results indicate that the use of DT-ApA for negative selection together with the application of LA-PCR for screening ensures efficient and time-saving screening for homologous recombinants.

Animals↗
PubMed1999

Reduced hippocampal LTP in mice lacking a presynaptic protein: complexin II.

The SNAP receptor (SNARE) complex is a core complex specialized for synaptic vesicle exocytosis, and the binding of SNAPs to the complex is an essential step for neurotransmitter release. Complexin I and II have been identified as SNARE-complex-associated proteins. Importantly, complexins compete with alpha-SNAP for binding to the complex, suggesting that complexins may modulate neurotransmitter release process. To examine this possibility and to understand the physiological function of complexins, we generated complexin II knockout mice. The complexin-II-deficient mice (-/-) were viable and fertile, and appeared normal. Electrophysiological recordings in the mutant hippocampus showed that ordinary synaptic transmission and paired-pulse facilitation, a form of short-term synaptic plasticity, were normal. However, long-term potentiation (LTP) in both CA1 and CA3 regions was impaired, suggesting that complexin II may not be essential for synaptic vesicle exocytosis, but it does have a role in the establishment of hippocampal LTP.

Adaptor Proteins, Vesicular Transport↗
PubMed1999

Eotaxin synthesis by nasal polyp fibroblasts.

Nasal polyps is a chronic inflammatory disease of the upper airway characterized by structural abnormalities including stromal fibrosis. Fibroblasts are a rich source of cytokines and inflammatory mediators and are thought to play an important role in the development of fibrosis. In addition, there is considerable evidence for the participation of eosinophils in the pathophysiology of nasal polyps. Although increased numbers of eosinophils are present in nasal polyps, the mechanisms responsible for their selective accumulation are not completely clear. Eotaxin is a chemokine that promotes the selective recruitment of eosinophils. Thus, it may be an important molecule for the recruitment of eosinophils in nasal polyps. The purpose of this study was to investigate whether nasal polyp fibroblasts synthesize eotaxin after stimulation with lipopolysaccharide, IL-1beta or TNF-alpha. Using primary nasal polyp tissue-derived fibroblast lines, we demonstrated that LPS, IL-1beta and TNF-alpha induced the gene expression and protein production of eotaxin in nasal polyp fibroblasts. This responsiveness to LPS, IL-1beta and TNF-alpha was time- and dose-dependent. These findings support the hypothesis that fibroblasts could play an important role in the recruitment of eosinophils in nasal polyps through the production of eotaxin.

Cells, Cultured↗
PubMed1999

Experimental autoimmune labyrinthitis induced by cell-mediated immune reaction.

This study was designed to establish an experimental cell-mediated autoimmune labyrinthitis model in C57BL/6 mice, which could exhibit high reproducibility and be adopted for precise immunological analysis. Inner ear antigen (IEA) was prepared from bovine membranous labyrinth. Following pretreatment with cyclophosphamide (CP) and primary sensitization with IEA/FCA, many inflammatory cells infiltrated transiently into the perilymphatic and endolymphatic regions of the cochlea, vestibule and the endolymphatic sac, but not into the kidney, lung, brain or liver. This reaction occurred from day 7 and peaked on day 12 in all animals, and then rapidly reduced. This reaction occurred in all experimental mice during day 10 to day 12. The control mice showed no cellular reaction in the inner ear. These results suggest that the inner ear may possess cross-species organ-specific antigen and that a cell-mediated immune reaction may play an important role in the induction of autoimmune labyrinthitis.

Animals↗
PubMed1999

Three-dimensional analysis of pressure nystagmus in labyrinthine fistulae.

In this study we analysed pressure nystagmus three-dimensionally in subjects with labyrinthine fistulae. The fistulae were localized in the lateral canal in five patients, in the posterior canal in one patient and in both lateral and anterior canals in two patients. The eye velocities during the fistula test in patients with lateral canal fistula and in a patient with posterior canal fistula were closely aligned with the anatomical axes. On the other hand, pressure nystagmus in cases with fistulae of both the lateral and anterior canals were not aligned with the anatomical axes of the combination of both canals.

Ear Canal↗
PubMed1999

Involvement of interleukin-18 (IL-18) in mixed lymphocyte reactions (MLR).

The in vitro mixed lymphocyte reaction (MLR) is a useful model to study alloresponsiveness to histocompatibility antigens. Secretion of different cytokine proteins in the supernatant of allo-MLR cultures has been reported in a few studies. We studied the levels of the cytokines interferon gamma (IFN-gamma) and interleukin-6 (IL-6), IL-10, IL-12, and IL-18 in the supernatant in allo-MLR by ELISA assay. Supernatant levels of IFN-y, IL-6, IL-10, and IL-18 were detected at 12 h after MLR and markedly increased thereafter. In contrast, secretion of IL-12 was detected after 48-72 h. These results suggested that IFN-gamma production depended on IL-18 in the early phase of MLR and depended on both IL-18 and IL-12 in the late phase. An antibody (Ab) neutralizing test was also performed. The levels of IFN-gamma were significantly downregulated after the addition of anti-IL-18 Ab, anti-IL-12 Ab, or anti-IFN-y Ab, and the levels of IL-12 were significantly downregulated after the addition of anti-IL-12 Ab and anti-IL-18 Ab. Treatment with these Ab did not suppress IL-6 production at all. The two-way MLR showed the same tendency as the one-way MLR. These results suggest the importance of IL-18 and IL-12 in allogeneic cell interactions and also suggest the usefullness of these Ab as regulators of alloresponsiveness.

Culture Media↗
PubMed1999

Structure determination of rubredoxin from Desulfovibrio vulgaris Miyazaki F in two crystal forms.

The structures of two crystal forms (form I, P3221, a = b = 43.7, c = 50.7 A; form II, P21, a = 27.3, b = 44.9, c = 51.2 A and beta = 90. 6 degrees ) of the rubredoxin from Desulfovibrio vulgaris Miyazaki F have been solved by the molecular-replacement method. Form I has been refined at a resolution of 2.0 A to an R value of 20.8% and includes 32 water molecules. Form II includes 86 water molecules and has been refined at 1.9 A resolution to an R value of 17.5%. In form II, there are three molecules in the asymmetric unit with the molecules related by a non-crystallographic 32 symmetry axis. In both crystal forms, it was found that only a few residues effectively participate in the formation of intermolecular contacts along both the crystallographic (form I) and the non-crystallographic (form II) 32 axes. The crystal structure of the form II crystal is compared with those of other rubredoxin molecules from anaerobic bacteria. From this comparison, a similarity in the core region, which is composed of aromatic residues and includes the active centre, has been revealed.

Amino Acid Sequence↗
PubMed1999

Real time image processing with an analog vision chip system.

A linear analog network model is proposed to characterize the function of the outer retinal circuit in terms of the standard regularization theory. Inspired by the function and the architecture of the model, a vision chip has been designed using analog CMOS Very Large Scale Integrated circuit technology. In the chip, sample/hold amplifier circuits are incorporated to compensate for statistic transistor mismatches. Accordingly, extremely low noise outputs were obtained from the chip. Using the chip and a zero-crossing detector, edges of given images were effectively extracted in indoor illumination.

Adaptation, Ocular↗
PubMed1999

Distinct expression of RANTES and GM-CSF by lipopolysaccharide in human nasal fibroblasts but not in other airway fibroblasts.

Eosinophil infiltration of tissue is a hallmark of nasal polyposis and asthma in both atopic and nonatopic patients. Structural cells like airway fibroblasts are a rich source of cytokines and inflammatory mediators. In order to verify whether airway fibroblasts play a role in eosinophilic infiltration, we investigated the release of eosinophil chemotactic and activating factors from airway fibroblasts when stimulated with nonallergenic exogenous agents such as endotoxin (lipopolysaccharide; LPS). Using a number of primary human airway tissue-derived fibroblast lines, we demonstrated that LPS could induce the gene expression and production of RANTES (regulated and normal T cell expressed and presumably secreted) and granulocyte/macrophage colony-stimulating factor (GM-CSF) only in nasal but not in pharyngeal, tracheal, bronchial, and lung fibroblasts. This selective responsiveness of nasal fibroblasts to LPS was time and dose dependent. These findings suggest that nasal fibroblasts may play an important role in the recruitment and activation of eosinophils into nasal polyps through the release of RANTES and GM-CSF.

Cells, Cultured↗
PubMed1999

A requirement for neuropilin-1 in embryonic vessel formation.

Neuropilin-1 is a membrane protein that is expressed in developing neurons and functions as a receptor or a component of the receptor complex for the class 3 semaphorins, which are inhibitory axon guidance signals. Targeted inactivation of the neuropilin-1 gene in mice induced disorganization of the pathway and projection of nerve fibers, suggesting that neuropilin-1 mediates semaphorin-elicited signals and regulates nerve fiber guidance in embryogenesis. Neuropilin-1 is also expressed in endothelial cells and shown to bind vascular endothelial growth factor (VEGF), a potent regulator for vasculogenesis and angiogenesis. However, the roles of neuropilin-1 in vascular formation have been unclear. This paper reported that the neuropilin-1 mutant mouse embryos exhibited various types of vascular defects, including impairment in neural vascularization, agenesis and transposition of great vessels, insufficient aorticopulmonary truncus (persistent truncus arteriosus), and disorganized and insufficient development of vascular networks in the yolk sac. The vascular defects induced by neuropilin-1 deficiency in mouse embryos suggest that neuropilin-1 plays roles in embryonic vessel formation, as well as nerve fiber guidance.

Animals↗
PubMed1999

Direct measurement of inter-doublet elasticity in flagellar axonemes.

The outer doublet microtubules in ciliary and flagellar axonemes are presumed to be connected with each other by elastic links called the inter-doublet links or the nexin links, but it is not known whether there actually are such elastic links. In this study, to detect the elasticity of the putative inter-doublet links, shear force was applied to Chlamydomonas axonemes with a fine glass needle and the longitudinal elasticity was determined from the deflection of the needle. Wild-type axonemes underwent a high-frequency, nanometer-scale vibration in the presence of ATP. When longitudinal shear force was applied, the average position of the needle tip attached to the axoneme moved linearly with the force applied, yielding an estimate of spring constant of 2.0 (S.D.: 0.8) pN/nm for 1 microm of axoneme. This value did not change in the presence of vanadate, i.e., when dynein does not form strong cross bridges. In contrast, it was at least five times larger when ATP was absent, i.e., when dynein forms strong cross bridges. The measured elasticity did not significantly differ in various mutant axonemes lacking the central-pair microtubules, a subset of inner-arm dynein, outer-arm dynein, or the radial spokes, although it was somewhat smaller in the latter two mutants. It was also observed that the shear displacement in an axoneme in the presence of ATP often took place in a stepwise manner. This suggests that the inter-doublet links can reversibly detach from and reattach to the outer doublets in a cooperative manner. This study thus provides the first direct measure of the elasticity of inter-doublet links and also demonstrates its dynamic nature.

Animals↗
PubMed1999

Adhesive explant culture of allergic nasal mucosa: effect of emedastine difumarate, an anti-allergic drug.

Allergic reaction of the nose comprises of an immediate and a late reaction. To evaluate nasal allergic reactions, many experiments have been performed by investigators. In this study, we performed a new tissue culture technique (adhesive explant culture) to analyze the migration of cells into the culture medium from the cultured allergic nasal mucosa in response to an allergen. Basophilic cells (mast cells and basophils) and eosinophils, which were released into the culture medium after the allergen challenge, were evaluated by the analysis of histamine and eosinophil cationic protein (ECP) content in the culture medium. Histamine and basophilic cells in the culture medium were more abundant in the immediate phase (within 30 min) after challenge than in the late phase (from 30 min to 10 hr). On the other hand, ECP and eosinophils in the culture medium were more abundant in the late phase than in the immediate phase. The increase of histamine content in both phases were not inhibited by pre-treatment of emedastine difumarate (EME), an anti-allergic drug. However, the increase of ECP in the late phase was inhibited by pre-treatment with EME. Moreover, the number of EG2-positive cells was also decreased by pre-treatment with EME. These results suggest that EME might lower the activation of eosinophils in the late phase of the allergic reaction. The present study also indicates that this adhesive explant culture system is useful model for studying the cellular allergic responses to drugs ex vivo.

Anti-Allergic Agents↗
PubMed1999

Apoptosis of human tumor cells by chemotherapeutic anthracyclines is enhanced by Bax overexpression.

One of the major factors for efficacy of a chemotherapeutic drug is its activity to induce apoptosis of tumor cells. Doxorubicin and daunorubicin, radiomimetic anthracycline-group drugs, have been used for chemotherapy for about 30 years. Here we established the colorectal tumor and osteosarcoma cells in which Bax expression can be induced by the treatment of isopropyl-beta-D-thiogalactopyranoside, and examined the effect of the Bax overexpression on the cell death caused by these drugs. While the Bax overexpression neither affected growth nor morphology of the undamaged cells, it enhanced the cell death caused by these drugs. Increase in cellular nucleus fragmentation and DNA ladder formation indicates that the Bax-enhanced cell death is due to enhanced apoptosis of the drug-treated cells. The enhanced cell death was not observed when the cells were irradiated with X-ray or treated with other chemotherapeutic agents we examined. These results indicate that Bax may have a specific role to enhance the efficacy of chemotherapy with anthracycline-group agents.

Anthracyclines↗
PubMed1999

Relationship between coronary blood flow velocity waveform and transmural distribution of myocardial blood flow in coronary artery.

It is important to know the transmural distribution of myocardial blood flow in assessing the severity of ischemia in coronary heart disease. We analyzed the relation between phasic waveform of epicardial coronary flow velocity with a Doppler flow probe in the left anterior descending artery in dogs and regional myocardial blood flow using a colored microsphere technique. Time-velocity integral in an average of 5 cardiac cycles was measured as an index of coronary blood flow during diastole (TVId) and systole (TVIs). The diastolic fraction of coronary blood flow (%DF) was defined as TVId/(TVId + TVIs). Myocardial specimens were divided into inner (subendocardial), middle, and outer (subepicardial) layers, and the inner layer to outer layer myocardial blood flow ratio (endo/epi ratio) was used as an index of transmural distribution of myocardial perfusion. The mean endo/epi ratio and the mean %DF decreased as the pressure gradient increased. There was a moderate but significant correlation (r = 0.57) between the endo/epi ratio and the %DF. In conclusion, analysis of the phasic pattern of coronary blood flow velocity provides some information about the transmural distribution of blood flow in the myocardium. The %DF may be a useful index for evaluating subendocardial ischemia.

Animals↗
PubMed1999

Modulatory effect of a serine protease inhibitor on surgical stress: its clinical implications.

The relationship between endogenous cytokine antagonists and surgical stress is poorly understood. Surgical stress induces immunosuppression, and the reversed therapy of postoperative immunosuppression has been expected. The aim of the present study was to assess the effect of a serine protease inhibitor on postoperative immune reactivity. Twenty patients with colorectal cancer were randomly separated into experimental and control groups of 10 patients each. The experimental group received perioperative administration of a serine protease inhibitor while the control group did not. Plasma levels of cytokine antagonists, which suppress cell-mediated immunity, such as cortisol, interleukin-1 receptor antagonist, soluble interleukin-2 receptor (sIL-2R) and soluble tumor necrosis factors p55, p75 (sTNF-R55, -R75) were simultaneously measured. Significant reductions of plasma concentration of sIL-2R and sTNF-R55 were observed. Perioperative administration of a serine protease inhibitor may contribute to ameliorating immunosuppression after major surgery.

Adult↗