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Biomedical subjects

T Yamazaki

Publications and source records attributed to T Yamazaki.

At least 199 records · Page 11Linked to original sources
PubMed2000

[A case of non-small-cell lung cancer successfully treated using combination chemotherapy with CDDP and vinorelbine].

A 67-year-old woman presented to our hospital with a chief complaint of bloody sputum. A plain chest X-ray a CT scan revealed a tumor shadow 3 cm in size in the middle lobe of the right lung, multiple nodular shadows in the bilateral lung fields and enlarged hilar and mediastinal lymph nodes. A tumor biopsy done under bronchoscopy revealed poorly differentiated adenocarcinoma of the lungs (cT2N3M1). She was given two courses of combination therapy consisting of cisplatin (80 mg/m2) and vinorelbine (20 mg/m2). The primary tumor in the middle lobe of the right lung and the lung metastases were markedly reduced in size, and a complete response was obtained. The only adverse events were grade 4 neutropenia and grade 2 nausea and vomiting.

Aged↗
PubMed2000

[Surgical treatment of middle cranial fossa arachnoid cyst in children with atypical symptoms--report of three cases].

We report three cases of symptomatic arachnoid cyst in the middle cranial fossa who presented with atypical symptoms including speech delay, vomiting and personality changes. They were improved by a cyst-peritoneal shunt operation. In many patients with a small or middle sized arachnoid cyst in the middle cranial fossa, their symptom tends to be so trivial that surgery is not the choice of treatment. The present cases suggest that peculiar symptoms caused by a limbic system lesion may be improved by treating the arachnoid cyst in the middle cranial fossa.

Arachnoid Cysts↗
PubMed1999

Kinetic studies on the successive reaction of neuronal nitric oxide synthase from L-arginine to nitric oxide and L-citrulline.

Rat neuronal nitric oxide synthase (nNOS) was heterologously expressed in Escherichia coliand purified. The conversion of L-arginine to N(omega)-hydroxy-L-arginine and further to L-citrulline in one cycle of the reaction of the purified nNOS was measured with the reaction rapid quenching method using (3)H-L-arginine as the substrate. It was found that most of the produced (3)H-N(omega)-hydroxy-L-arginine was successively hydroxylated to (3)H-L-citrulline without leaving the enzyme. From the analysis of time courses, the rate constants for each reaction step, and also for the dissociation of the intermediate, were estimated at various temperature in which the rates for the first and the second reactions were not much different each other but the rate for the dissociation of (3)H-N(omega)-hydroxy-L-arginine from the enzyme was significantly slow. Under the steady-state reaction condition, almost all of the nNOS was estimated to be active from the amount of burst formation of L-citrulline in the pre-steady state. The rate constant for the dissociation of the product L-citrulline from nNOS was calculated from the combination of results of the rapid quenching experiments and the metabolism of L-arginine in the presence of an excess amount of substrate, which was the smallest among all the rate constants in one cycle of the nNOS reaction. The activation energies for all the reaction steps were determined from the temperature dependence of the rate constants, which revealed that the rate-determining step of the nNOS reaction in the steady state was the dissociation of the product L-citrulline from the enzyme.

Animals↗
PubMed1999

NMR observation of selected segments in a larger protein: central-segment isotope labeling through intein-mediated ligation.

Peptide segments in a protein, which can include an active site of interest or be a series of parts constituting the entire structure, are now selectively observed by nuclear magnetic resonance (NMR) spectroscopy using samples prepared by the intein-mediated ligation method. Two separate inteins were used to ligate NMR-transparent segments to both the ends of an NMR-visible segment, producing a partly visible intact protein molecule. The (15)N-(1)H correlation spectrum of a 370-residue maltose binding protein labeled with (15)N at a continuous segment comprising residues Gly(101)-Ser(238) showed the essential elimination of signal overlapping, the signals being at the same positions as for the uniformly labeled sample. This method will allow structural analysis by NMR of over 50-kDa proteins in combination with contemporary NMR techniques suppressing the signal decays of larger proteins.

ATP-Binding Cassette Transporters↗
PubMed1999

MAPK superfamily plays an important role in daunomycin-induced apoptosis of cardiac myocytes.

BACKGROUND: Although anthracyclines, such as daunomycin (DM) and adriamycin, are potent chemotherapeutic agents, they have serious adverse effects, including cardiac toxicity. In the present study, we investigated the molecular mechanisms of DM-induced cardiomyocyte impairment. METHODS AND RESULTS: When cultured cardiac myocytes of neonatal rats were exposed to 1 micromol/L DM for 24 hours, many cells became positive for TUNEL staining, with morphological changes characteristic of apoptosis. Fragmentation of DNA into oligonucleosome-size fragments was recognized by agarose gel electrophoresis in DM-treated myocytes. DM activated 3 members of the mitogen-activated protein kinase (MAPK) family dose-dependently, such as extracellular signal-regulated protein kinases (ERKs), c-Jun NH(2)-terminal kinases, and p38 MAPK in cardiac myocytes. Oxyradical scavengers or Ca(2+) chelators inhibited DM-induced activation of ERKs and p38 MAPK. DM-induced activation of ERKs was also inhibited by overexpression of dominant negative mutants of Ras (D.N.Ras), and the p38 MAPK activation was attenuated by D.N.Rho. The number of DM-induced apoptotic cells was markedly increased when the ERK signaling pathway was selectively blocked by a specific MAPK/ERK kinase inhibitor, PD98059, whereas pretreatment with a specific inhibitor of p38 MAPK, SB203580, significantly reduced the amount of apoptosis. CONCLUSIONS: These results suggest that DM activates MAPKs through reactive oxygen species and Ca(2+) and that the MAPK family plays important roles in DM-induced apoptosis in cardiac myocytes. ERKs protect cardiomyocytes from apoptosis, whereas p38 MAPK is involved in the induction of cardiomyocyte apoptosis.

Animals↗
PubMed1999

Localization of opioid-binding cell adhesion molecule (OBCAM) in adult rat brain.

We investigated the tissue distribution and brain localization of opioid-binding cell adhesion molecule (OBCAM) in the adult rats by immunoblotting and immunohistochemistry using a monoclonal anti-OBCAM peptide antibody that is specific for OBCAM. OBCAM was preferentially expressed in the central nervous system (CNS) and at a very low level in the spleen. Within the brain, OBCAM was distributed in almost all the gray matter, but little or no immunoreactive OBCAM was found in the white matter. Morphologically, the distribution pattern of OBCAM immunoreactivity was very similar to that of synaptophysin, suggesting a role in the synaptic machinery.

Animals↗
PubMed1999

Structure-activity relationships of calcicludine and dendrotoxin-I, homologous peptides acting on different targets, calcium and potassium channels.

Calcicludine (CaC) and dendrotoxin-I (DTX-I) possess high homology of their primary structures despite their different biological activities acting on calcium and potassium channels, respectively. In order to elucidate the channel specificity displayed by these toxins, their three-dimensional structures were compared by NMR. These analyses revealed that their overall conformations are similar except for the structure at the N-terminus. To demonstrate the significance of this N-terminal, chimeric peptides, CaC(1-30)/DTX-I(31-60) and DTX-I(1-30)/CaC(31-60), were synthesized. The CD spectra and receptor-binding measurements of chimeric peptides indicated that the contribution to the overall conformation and to the affinity of the N-terminal part of molecule seem to be more important than that of the C-terminal one. These results suggest that the N-terminal part may participate in distinguishing between calcium and potassium channels.

Amino Acid Sequence↗
PubMed1999

A heterozygous mutation of beta-actin associated with neutrophil dysfunction and recurrent infection.

A human disorder caused by mutation in nonmuscle actin has not been reported. We report here a variant of nonmuscle actin in a female patient with recurrent infections, photosensitivity, and mental retardation. She also had abnormalities in neutrophil chemotaxis, superoxide production, and membrane potential response. Two-dimensional PAGE analysis of proteins from neutrophils and other cell types from this patient demonstrated a unique protein spot migrating at 42 kDa with pI shifted slightly to neutral relative to normal beta- and gamma-actin. Digestion peptide mapping and Western blotting showed this spot to be an abnormal actin. A full-length cDNA library was constructed by using mRNA from patient's cells and cDNA encoding the mutant beta-actin molecule was identified by an in vitro translation method. Sequencing of the clones demonstrated a G-1174 to A substitution, predicting a glutamic acid-364 to lysine substitution in beta-actin and eliminating a HinfI DNase restriction site found in normal beta-actin sequence. By HinfI digestion and by sequencing, the mutation in one allele of patient's genomic DNA was confirmed. Though no defect in cell-free polymerization of actin was detected, this defect lies in a domain important for binding to profilin and other actin-regulatory molecules. In fact, the mutant actin bound to profilin less efficiently than normal actin did. Heterozygous expression of mutant beta-actin in neutrophils and other cells of this patient may act in a dominant-negative fashion to adversely affect cellular activities dependent on the function of nonmuscle actin.

Actins↗
PubMed1999

Identification of the DNA binding surface of H-NS protein from Escherichia coli by heteronuclear NMR spectroscopy.

The DNA binding domain of H-NS protein was studied with various N-terminal deletion mutant proteins and identified by gel retardation assay and heteronuclear 2D- and 3D-NMR spectroscopies. It was shown from gel retardation assay that DNA binding affinity of the mutant proteins relative to that of native H-NS falls in the range from 1/6 to 1/25 for H-NS(60-137), H-NS(70-137) and H-NS(80-137), whereas it was much weaker for H-NS(91-137). Thus, the DNA binding domain was defined to be the region from residue A80 to the C-terminus. Sequential nuclear Overhauser effect (NOE) connectivities and those of medium ranges revealed that the region of residues Q60-R93 in mutant protein H-NS(60-137) forms a long stretch of disordered, flexible chain, and also showed that the structure of the C-terminal region (residues A95-Q137) in mutant H-NS(60-137) was nearly identical to that of H-NS(91-137). 1H and 15N chemical shift perturbations induced by complex formation of H-NS(60-137) with an oligonucleotide duplex 14-mer demonstrated that two loop regions, i.e. residues A80-K96 and T110-A117, play an essential role in DNA binding.

Amino Acid Sequence↗
PubMed1999

The molecular mechanism of cardiac hypertrophy and failure.

Mechanical stretch induced by high blood pressure is an initial factor leading to cardiac hypertrophy. In an in vivo study, an angiotensin II (AngII) type 1 receptor antagonist TCV116 reduced left ventricular (LV) weight, LV wall thickness, transverse myocyte diameter, relative amount of V3 myosin heavy chain, and interstitial fibrosis, while treatment with hydralazine did not. In an in vitro study using cultured cardiomyocytes, mechanical stretch activated second messengers such as mitogen-activated protein (MAP) kinase, followed by increased protein synthesis. Additionally, in the stretch-conditioned medium AngII and endothelin-1 concentrations were increased. Furthermore, the Na+/H+ exchanger activated by mechanical stretch modulated the hypertrophic responses of cardiomyocytes. The pathways leading to MAP kinase activation differed between cell types. In cardiac fibroblasts AngII activated MAP kinase via G beta gamma subunit of Gi, Src, Shc, Grb2, and Ras, whereas Gq and protein kinase C were critical in cardiomyocytes.

Angiotensin II↗
PubMed1999

Role of ecto-kinase in phorbol ester-enhanced growth hormone-binding protein release from human IM-9 cells.

Previously we reported that a phorbol ester, phorbol 12, 13-dibutyrate (PDBu), increased the release of human growth hormone-binding protein (hGH-BP) in IM-9 cells, and that this phorbol ester-enhanced release was mediated by protein kinase Ca (PKCalpha). In the present study, the mechanisms of the phorbol ester-enhanced hGH-BP release were further investigated. Treatment of IM-9 cells with PDBu did not increase hGH-BPs (55-60 kDa) in the intracellular soluble fraction. When the cells were treated with trypsin to remove human growth hormone receptors (hGHRs) on the cell surface after stimulation, no hGH-BPs were detected in the culture supernatants, nor did treatment with bafilomycin A1 or chloroquine affect the PDBu-enhanced hGH-BP release. These results suggest that hGH-BPs released by PDBu stimulation are derived from cell surface hGHRs and not generated within the cells. Protein kinase inhibitors with broad specificities, K-252a and K-252b, inhibited the PDBu-enhanced release with almost the same dose-dependency, although only a trace amount of K-252b was incorporated into IM-9 cells than K-252a, suggesting that K-252b probably inhibits an ecto-kinase extracellularly. PDBu actually enhanced the phosphorylation of several extracellular proteins, and this enhanced phosphorylation was completely inhibited by K-252b treatment. Moreover, the PKCalpha-specific inhibitor bisindolylmaleimide III which inhibits PDBu enhanced hGH-BP release inhibited the PDBu-enhanced phosphorylation of extracellular proteins. On the other hand, the impermeable PKC inhibitor PKC inhibitor peptide 19-31 did not inhibit PDBu-enhanced release, suggesting that the target PKCalpha for PDBu is not present on the extracellular surface. Taken together, these results suggest that, in addition to intracellular PKCalpha, activation of an undefined ecto-kinase may also be involved in the PDBu-enhanced hGH-BP release.

Carbazoles↗
PubMed1999

CAST, a novel CD3epsilon-binding protein transducing activation signal for interleukin-2 production in T cells.

Antigen recognition through T cell receptor (TCR)-CD3 complex transduces signals into T cells, which regulate activation, function, and differentiation of T cells. The TCR-CD3 complex is composed of two signaling modules represented by CD3zeta and CD3epsilon. Signaling through CD3zeta has been extensively analyzed, but that via CD3epsilon, which is also crucial in immature thymocyte development, is still not clearly understood. We isolated cDNA encoding a novel CD3epsilon-binding protein CAST. CAST specifically interacts in vivo and in vitro with CD3epsilon but not with CD3zeta or FcRgamma via a unique membrane-proximal region of CD3epsilon. CAST is composed of 512 amino acids including a single tyrosine and undergoes tyrosine phosphorylation upon TCR stimulation. Overexpression of two dominant-negative types of CAST, a minimum CD3epsilon-binding domain and a tyrosine-mutant, strongly suppressed NFAT activation and interleukin-2 production. These results demonstrate that CAST serves as a component of preformed TCR complex and transduces activation signals upon TCR stimulation and represents a new signaling pathway via the CD3epsilon-containing TCR signaling module.

Amino Acid Sequence↗
PubMed1999

Role of the renin-angiotensin system in cardiac hypertrophy.

Hypertrophy is an adaptive response of the heart to hemodynamic overload such as hypertension. However, it is generally accepted that cardiac hypertrophy is one of the most critical risk factors of heart disease. Therefore, for the treatment of hypertension it is important to understand the mechanism of cardiac hypertrophy and to establish effective pharmaceutical interventions. Mechanical stretch induced by hypertension is an initial factor leading to cardiac hypertrophy. In an in vivo study using spontaneously hypertensive rats, an angiotensin II type 1 receptor antagonist, TCV116, decreased left ventricular weight, left ventricular wall thickness, transverse myocyte diameter, relative amount of V3 myosin heavy chain, and interstitial fibrosis, whereas treatment with hydrolazine did not. In an in vitro study using cultured cardiomyocytes of neonatal rats, mechanical stretch activated second messengers, such as extracellular signal-regulated protein kinase (ERK), followed by increased protein synthesis. Additionally, in the stretch-conditioned medium, the levels of angiotensin II and endothelin-1 concentrations were increased. Moreover, the Na+/H+ exchanger activated by mechanical stretch modulated the hypertrophic responses of cardiomyocytes. To further elucidate whether angiotensin II is indispensable for mechanical stress-induced cardiac hypertrophy, mechanical stretch-induced ERK activation was examined in angiotensin II type 1a receptor knock-out mice. Although the addition of angiotensin II had no effects on the ERK activity in cardiomyocytes of angiotensin II type 1a receptor knockout mice, mechanical stretch induced a larger increase in the ERK activity in cardiac myocytes from these mice compared with cardiac myocytes of wild-type mice. These results suggest that mechanical stretch could induce hypertrophic responses in cardiac myocytes even in the absence of angiotensin II. The pathways leading to ERK activation differed between cell types. In cardiac fibroblasts, angiotensin II activated ERK via the G(beta)gamma subunit of Gi, Src, Shc, Grb2, and Ras, whereas Gq and protein kinase C were critical in cardiomyocytes.

Angiotensin II↗
PubMed1999

Bacillus subtilis histone-like protein, HBsu, is an integral component of a SRP-like particle that can bind the Alu domain of small cytoplasmic RNA.

Small cytoplasmic RNA (scRNA) is metabolically stable and abundant in Bacillus subtilis cells. Consisting of 271 nucleotides, it is structurally homologous to mammalian signal recognition particle RNA. In contrast to 4.5 S RNA of Escherichia coli, B. subtilis scRNA contains an Alu domain in addition to the evolutionarily conserved S domain. In this study, we show that a 10-kDa protein in B. subtilis cell extracts has scRNA binding activity at the Alu domain. The in vitro binding selectivity of the 10-kDa protein shows that it recognizes the higher structure of the Alu domain of scRNA caused by five consecutive complementary sequences in the two loops. Purification and subsequent analyses demonstrated that the 10-kDa protein is HBsu, which was originally identified as a member of the histone-like protein family. By constructing a HBsu-deficient B. subtilis mutant, we showed that HBsu is essential for normal growth. Immunoprecipitating cell lysates using anti-HBsu antibody yielded scRNA. Moreover, the co-precipitation of HBsu with (His)6-tagged Ffh depended on the presence of scRNA, suggesting that HBsu, Ffh, and scRNA make a ternary complex and that scRNA serves as a functional unit for binding. These results demonstrated that HBsu is the third component of a signal recognition particle-like particle in B. subtilis that can bind the Alu domain of scRNA.

Amino Acid Sequence↗
PubMed1999

Depletion of Bacillus subtilis histone-like protein, HBsu, causes defective protein translocation and induces upregulation of small cytoplasmic RNA.

Small cytoplasmic RNA (scRNA) is a metabolically stable homologue of mammalian SRP RNA that contains an Alu-like domain. The Bacillus subtilis histone-like protein HBsu can bind this domain. We demonstrate here that repressing the level of HBsu results in slow growth and the accumulation of precursor of beta-lactamase fusion proteins having the signal sequence of alkaline protease, penicillin binding protein 5* (PBP5*) or CGTase. The degree of the translocation defect varied among the various signal sequences tested. A pulse-chase experiment showed that processing the alpha-amylase signal sequence is significantly inhibited in HBsu-depleted cells. Northern blot analysis indicated that repressing the HBsu gene induces scRNA upregulation, indicating that the defective translocation of presecretory proteins is not due to a reduced scRNA level. The data presented here suggest that HBsu plays a pivotal role in SRP function rather than simply stabilizing the other SRP components such as scRNA.

Bacillus subtilis↗