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H Sakamoto

Publications and source records attributed to H Sakamoto.

At least 19 recordsLinked to original sources
PubMed1999

Structural characterization of chemically derivatized oligosaccharides by nanoflow electrospray ionization mass spectrometry.

Oligosaccharides released from several glycoproteins were derivatized with either 4-aminobenzoic acid 2-(diethylamino)ethyl ester (ABDEAE) (Yoshino, K.; et al. Anal. Chem. 1995, 67, 4028-4031) or 2-aminopyridine. The resulting derivatives were analyzed on a nanoflow electrospray ionization (ESI) quadrupole-inlet time-of-flight mass spectrometer using the low-energy collision-induced dissociation technique. In the MS/MS spectra, the oxonium (b or internal series) and y series ions, which are derived from the multiply charged precursor ions, were predominant and were used for the structural readout. Some oxonium ions that were observed in the low-mass region, but that were not found in the PSD analyses (Mo, W.; et al. Anal. Chem. 1998, 70, 4520-4526), rendered a more detailed structural insight. The oxonium ions at m/z 512.2, which are derived from the fucosylated oligosaccharides of immunoglobulin Y and thyroglobulin, were observed, suggesting that fucosylation had occurred proximal to the outer nonreducing terminus. In addition, the data herein show that structural elucidation can be routinely achieved at a low sample concentration. For the case of ABDEAE derivatives, this can be achieved at the 50 fmol/microL level and with the actual sample consumption at the attomole level using nanoflow ESI MS/MS.

Aminopyridines↗
PubMed1999

Lateromedial and dorsoplantar borders among supplying areas of the nerves innervating the intrinsic muscles of the foot.

The branching patterns of nerves supplying the intrinsic muscles of the foot were analyzed as a basis to confirm the muscle layer structure. Thirty-eight feet of 20 Japanese cadavers were examined in detail in this study. The first dorsal interosseus was innervated by a branch from the deep peroneal nerve as well as a branch of the lateral plantar nerve in 92.1%, the second dorsal interosseus in 10. 5% and the third dorsal interosseus in 2.6%. In three specimens, branches from the deep peroneal nerve innervated the oblique head of the adductor hallucis or the lateral head the flexor hallucis brevis. In addition, branches from the medial and lateral plantar nerves and the deep peroneal nerve formed communication loops in three specimens. The first dorsal interosseus, the oblique head of the adductor hallucis and the lateral head of the flexor hallucis and their innervating nerve branches are closely related within the first intermetatarsal space. Since the tibial part of the first interosseus muscle primordium is occupied in the space during development, the variations of innervation patterns and formation of the communicating nerve loops may be explained by various combinations of the part and the other muscle primordia.

Adult↗
PubMed1999

Neuronal expression of a Caenorhabditis elegans elav-like gene and the effect of its ectopic expression.

We examined the expression of a Caenorhabditis elegans (C. elegans) elav-like gene, which we designated elr-1. The elr-1 gene encodes a predicted 456-amino-acid protein containing three putative RNA-binding domains and belongs to the ELAV family, which is functionally involved in neuronal differentiation. Northern blot analysis suggested that the levels of elr-1 mRNA are regulated developmentally. A elr-1::gfp reporter gene under the control of the elr-1 promoter was expressed specifically in the ring ganglia near the nerve ring, the ventral nerve cord (VNC), and the pre-anal and lumbar ganglia. In the VNC, GFP-positive cells were shown to be acetylcholine-producing motor neurons which increased in number as development proceeded, suggesting that elr-1 is expressed in mature neurons. Ectopic expression of ELR-1 protein at the L4 larval and adult stages, but not earlier stages, caused irreversible death, accompanied by uncoordinated movement (Unc), clear (Clr), and egg-laying defective (Egl) phenotypes, which are often observed in mutants with neuronal defects. These results suggest that ELR-1 may have important functions in specific mature neurons in C. elegans.

Amino Acid Sequence↗
PubMed1999

Ferric alpha-hydroxyheme bound to heme oxygenase can be converted to verdoheme by dioxygen in the absence of added reducing equivalents.

Whether or not reducing equivalents are indispensable for the conversion of ferric alpha-hydroxyheme bound to heme oxygenase-1 to verdoheme remains controversial (Matera, K. M., Takahashi, S., Fujii, H., Zhou, H., Ishikawa, K., Yoshimura, T., Rousseau, D. L., Yoshida, T., and Ikeda-Saito, M. (1996) J. Biol. Chem. 271, 6618-6624; Liu, Y., Moënne-Loccoz, P., Loehr, T. M., and Ortiz de Montellano, P. R. (1997) J. Biol. Chem. 272, 6906-6917). To resolve this controversy, we have prepared a ferric alpha-hydroxyheme-heme oxygenase-1 complex and titrated the complex with O2 under strictly anaerobic conditions. The formation of verdoheme was monitored by optical and electron spin resonance spectroscopies. Electron spin resonance spectra of the complex showed that alpha-hydroxyheme exists as a mixture of resonance structures composed of the iron(III) porphyrin and the iron(II) porphyrin pi neutral radical. Upon addition of CO the latter species becomes dominant. The results obtained from these titration experiments indicate that alpha-hydroxyheme can be converted to verdoheme by an approximately equimolar amount of O2 without any requirement for exogenous electrons. The verdoheme formed from alpha-hydroxyheme was shown to be in the ferrous oxidation state by the addition of CO or potassium ferricyanide to the resultant verdoheme-heme oxygenase-1 complex.

Electron Spin Resonance Spectroscopy↗
PubMed1999

Induction of apoptosis and suppression of clonogenicity of ovarian carcinoma cells with estrogen mustard.

BACKGROUND: This study was conducted to evaluate whether estramustine (estrogen mustard [EM]) is a promising alternative in the treatment of patients with epithelial ovarian carcinoma (OVCA). EM is a microtubule-associated protein (MAP) specific antimicrotubule agent with low toxicity. METHODS: The authors investigated the ability of EM to induce apoptosis and suppress colony formation of OVCA cells. Paclitaxel was used as a positive control. DNA electrophoresis and terminal deoxynucleotidyl dUTP-X nick end labeling (TUNEL) assays were used to detect internucleosomal DNA fragmentation. Flow cytometry was used to identify apoptotic cells and disturbance of the cell cycle of EM-treated OVCA cells further. Quantitation of detached cultured cells also provided a relative estimate of the apoptotic response of OVCA cells to treatment with EM. The colony formation assay was used to evaluate the effects of EM on clonogenicity. RESULTS: The effects of EM on four OVCA cell lines in culture were highly similar to those of paclitaxel in causing apoptosis and inhibiting clonogenicity. DNA electrophoresis and TUNEL assays showed that EM induced internucleosomal DNA fragmentation in OVCA cells. Flow cytometry showed changes typical of apoptotic changes and cell cycle block and synchronization at the G2/M-phase. Counting of detached cells showed a log-dose response to EM treatment. The colony formation assay also showed a log-dose response suppression of OVCA cell clonogenicity after treatment with EM. CONCLUSIONS: EM may be a promising candidate in the clinical treatment of patients with OVCA. The lower toxicity and MAP specific action of EM is a novel chemotherapy for OVCA.

Antineoplastic Agents, Alkylating↗
PubMed1999

Naturally occurring somatic motoneuron death in a teleost angelfish, Pterophyllum scalare.

Naturally occurring somatic motoneuron death in a teleost angelfish, Pterophyllum scalare, was investigated histochemically and electron microscopically. The number of motor axons in the ventral root, which corresponds to the motoneuron number in spinal hemisegment, was rapidly increased beyond the adult value within 3 days after hatching, and then decreased to reach the adult value within a few weeks. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) histochemistry, which detects fragmented nuclear DNA characteristic to apoptotic cells, showed that the apoptotic cells are located in the motor column of the cord in the larvae at specific developmental stages. Electron microscopic observations of the spinal cells further confirmed the motoneuron apoptosis. The present data suggest that the massive death of somatic motoneurons at certain ontogenic stages which has been known to occur in higher vertebrates also takes place in fish.

Animals↗
PubMed1999

Identification of a novel promoter and exons of the c-ERBB-2 gene.

The c-ERBB-2 gene is frequently amplified in various cancers. During the course of studies on the structural characterization of the amplification units, we isolated four cDNA clones, A39, GRB7, C51 and CAB1 with corresponding genes localized on the c-ERBB-2 locus. A tentative gene, C51, was located at about 12 kb upstream of the c-ERBB-2 gene. By molecular cloning of a full-length cDNA clone of C51 and the reverse transcription PCR (RT-PCR) method, we identified a novel transcript of c-ERBB-2 containing new 5' sequences including the C51 sequence, demonstrating that the c-ERBB-2 gene has a novel promoter and new exons. The structural organization of the novel promoter and exons of c-ERBB-2 was revealed by complete sequence analysis of a total size of about 20 kb of genomic DNA clones containing the 5'-flanking region of the previously described c-ERBB-2 gene. Transient expression of the newly identified promoter-reporter gene constructs in breast cancer cell line MCF-7 showed that the elements responsible for promoter activity were contained in a 697 bp region upstream of the transcriptional start site. The new transcript may encode a protein different in the portion of the extracellular domain. Although the presence of the predicted protein product was not examined, this report is important in that it provides a new aspect of the c-ERBB-2 protooncogene products.

Adult↗
PubMed1999

Structural basis for recognition of the tra mRNA precursor by the Sex-lethal protein.

The Sex-lethal (Sxl) protein of Drosophila melanogaster regulates alternative splicing of the transformer (tra) messenger RNA precursor by binding to the tra polypyrimidine tract during the sex-determination process. The crystal structure has now been determined at 2.6 A resolution of the complex formed between two tandemly arranged RNA-binding domains of the Sxl protein and a 12-nucleotide, single-stranded RNA derived from the tra polypyrimidine tract. The two RNA-binding domains have their beta-sheet platforms facing each other to form a V-shaped cleft. The RNA is characteristically extended and bound in this cleft, where the UGUUUUUUU sequence is specifically recognized by the protein. This structure offers the first insight, to our knowledge, into how a protein binds specifically to a cognate RNA without any intramolecular base-pairing.

Amino Acid Sequence↗
PubMed1999

The JAK-binding protein JAB inhibits Janus tyrosine kinase activity through binding in the activation loop.

The Janus family of protein tyrosine kinases (JAKs) regulate cellular processes involved in cell growth, differentiation and transformation through their association with cytokine receptors. However, compared with other kinases, little is known about cellular regulators of the JAKs. We have recently identified a JAK-binding protein (JAB) that inhibits JAK signaling in cells. In the studies presented here we demonstrate that JAB specifically binds to the tyrosine residue (Y1007) in the activation loop of JAK2, whose phosphorylation is required for activation of kinase activity. Binding to the phosphorylated activation loop requires the JAB SH2 domain and an additional N-terminal 12 amino acids (extended SH2 subdomain) containing two residues (Ile68 and Leu75) that are conserved in JAB-related proteins. An additional N-terminal 12-amino-acid region (kinase inhibitory region) of JAB also contributes to high-affinity binding to the JAK2 tyrosine kinase domain and is required for inhibition of JAK2 signaling and kinase activity. Our studies define a novel type of regulation of tyrosine kinases and might provide a basis for the design of specific tyrosine kinase inhibitors.

Amino Acid Sequence↗
PubMed1999

APS, an adaptor protein containing PH and SH2 domains, is associated with the PDGF receptor and c-Cbl and inhibits PDGF-induced mitogenesis.

Previously we cloned a novel adaptor protein, APS (adaptor molecules containing PH and SH2 domains) which was tyrosine phosphorylated in response to c-kit or B cell receptor stimulation. Here we report that APS was expressed in some human osteosarcoma cell lines, markedly so in SaOS-2 cells, and was tyrosine-phosphorylated in response to several growth factors, including platelet derived growth factor (PDGF), insulin-like growth factor (IGF), and granulocyte-macrophage colony stimulating factor (GM-CSF). Ectopic expression of the wild type APS, but not C-terminal truncated APS, in NIH3T3 fibroblasts suppressed PDGF-induced MAP kinase (Erk2) activation, c-fos and c-myc induction as well as cell proliferation. In vitro binding experiments suggest that APS bound to the beta type PDGF receptor, mainly via phosphotyrosine 1021 (pY1021). Indeed, tyrosine phosphorylation of PLC-gamma, which has been demonstrated to bind to pY1021, but not that of PI3 kinase and associated proteins, was reduced in APS transformants. PDGF induced phosphorylation of the tyrosine residue of APS close to the C-terminal end. In vitro and in vivo binding experiments indicate that the tyrosine phosphorylated C-terminal region of APS bound to c-Cbl, which has been shown to be a negative regulator of tyrosine kinases. Since coexpression of c-Cbl with wild type APS, but not C-terminal truncated APS, synergistically inhibited PDGF-induced c-fos promoter activation, c-Cbl could be a mechanism of inhibitory action of APS on PDGF receptor signaling.

Adaptor Proteins, Signal Transducing↗
PubMed1999

Enhancement of glucuronosyl etoposide transport by glutathione in multidrug resistance-associated protein-overexpressing cells.

Multidrug resistance-associated protein (MRP) has been shown to transport glutathione (GSH) S-conjugates such as leukotriene C4 (LTC4) and S-(2,4-dinitrophenyl)-glutathione (DNP-SG). On the other hand, it has while it has been reported that MRP-overexpressing cells exhibit decreased sensitivity to drugs which do not form GSH S-conjugates. In this study, we found that GSH affects the transport of glucuronosyl etoposide as a major metabolite of etoposide in MRP-overexpressing KB/VP-4 cells. The relative resistance level of KB/VP-4 cells to etoposide was 70-fold that of wild-type KB cells. Membrane vesicles prepared from KB/VP-4 cells exhibited markedly enhanced ATP-dependent transport of glucuronosyl etoposide as well as LTC4. Transport of glucuronosyl etoposide was augmented in the presence of GSH. Treatment of KB/VP-4 cells with buthionine sulfoximine (BSO), an inhibitor of GSH synthesis, resulted in about 75% depletion of cellular GSH levels, a four-fold increase of the sensitivity to etoposide and depression of glucuronosyl etoposide efflux. These results suggest that GSH plays a role in the enhancement of MRP-mediated glucuronosyl etoposide transport.

ATP-Binding Cassette Transporters↗
PubMed1999

Fish calcitonin genes: primitive bony fish genes have been conserved in some lower vertebrates.

Using the polymerase chain reaction method, we amplified calcitonin genes from the cDNA of the ultimobranchial glands or from the genomic DNA of the blood cells and liver of various fishes. The fishes examined were primitive bony fishes (lungfish, polypterus, sturgeon, and gar), 16 species of teleosts, and cartilaginous fishes (stingray). Sequenced calcitonin genes were compared among fishes and with those of reptiles and chickens. The similarity of the calcitonin genes was the highest between the reptile and chicken groups and the primitive bony fishes (sequence similarity of the nucleotides 81-90%). The values between teleosts and the primitive bony fishes were 70-81%, with the exception of eels. Eel calcitonin genes were very similar to those of primitive bony fishes (83-88%). Stingray calcitonin genes were relatively more similar to those of the primitive bony fishes (74-78%) than to teleosts (63-73%). In goldfish and sardine, two types of calcitonin genes were found. We concluded that the genes of primitive bony fish are placed at a fundamental position in this hormone, at least among these vertebrates.

Animals↗
PubMed1999

Thymidine phosphorylase expression is predominantly observed in stroma of well-differentiated adenocarcinoma of endometrium and correlates with a frequency of vascular involvement.

Thymidine phosphorylase (dThdPase) expression was studied in 20 cases of well-differentiated endometrial carcinomas to examine the clinicopathological significance. Immunohistochemical study showed predominant dThdPase expression in tumor stroma. No expression was detected in coexisting normal endometrial stroma or stroma of endometriosis. Seven cases (35%) showed strong stain and 13 (65%) showed weak stain. Based on the strength of immunoreactivity, patients were grouped into two groups: group S (strong) and group W (weak). In group S, 5 (71%) showed vascular involvement whereas in group W, only 3 (23%) cases were positive, giving a significant difference in the frequency of vascular involvement between the two groups (P < 0.03, chi2 analysis). No correlation was found between dThdPase expression and myometrial invasion. Patients with vascular involvement resulted in poorer outcome (P < 0.003) whereas between group S and W, there showed no difference in survival (group S vs group W; P < 0.15). A multivariate analysis including stage, vascular involvement, and dThdPase stain as variables showed none as independent risk factors. However, univariate analysis showed that the presence of a vascular involvement was a risk factor for a shorter survival (relative risk = 6.277, 95% range = 1.2-32.6, P < 0. 03). It is concluded that expression of dThdPase in endometrial carcinoma is a marker of a desmoreaction and a risk factor for vascular invasion. Since vascular invasion is a risk factor for poor outcome, the significance of dThdPase expression in endometrial carcinoma requires further clinical evaluations.

Adenocarcinoma↗
PubMed1999

Intraoperative irradiation after surgery for locally recurrent rectal cancer.

PURPOSE: This study retrospectively evaluated the effects of intraoperative electron beam irradiation on patients with locally recurrent (pelvic) rectal cancer. METHODS: From November 1, 1975, to December 31, 1997, 51 patients underwent surgery for locally recurrent rectal or rectosigmoid cancer, and 27 patients received intraoperative electron beam irradiation. The intraoperative electron beam irradiation dose was 15 to 30 Gy. Kaplan-Meier survival estimates at three and five years were analyzed for the 47 patients who recovered postoperatively. RESULTS: Statistically significant factors related to survival included intraoperative electron beam irradiation vs. no intraoperative electron beam irradiation (P=0.0007), amount of residual tumor (slight vs. gross; P=0.0022), and symptom status (P=0.0024). Factors not associated with survival included distant metastases at reoperation, type of surgery for the recurrent tumor, external beam irradiation, pathologic grade, age, and gender. Surgical resection without intraoperative electron beam irradiation resulted in three-year and five-year survival rates of 5 and 0 percent, respectively. For patients who received intraoperative electron beam irradiation, the three-year survival rate was 43 percent and five-year survival rate was 21 percent. Intraoperative electron beam irradiation was a statistically significant factor related to survival in patients with and without distant metastasis (P=0.04 and P=0.0035, respectively), with slight residual tumor (P=0.0003), or with palliative surgery (P=0.0276). CONCLUSION: The trends seen in resection with intraoperative electron beam irradiation are encouraging with regard to improvements in survival as compared with studies not using intraoperative electron beam irradiation treatment.

Adult↗
PubMed1999

Two cases of chronic tonsillitis studied by FDG-PET.

We report two cases of chronic tonsillitis studied by FDG-PET. Symmetrical high FDG uptake by the tonsils was observed in both cases. On histopathologic examination of the resected tonsils, follicular hyperplasia was observed with proliferation of lymphocytes in the germinal centers. Increased glucose metabolism in active inflammation involving lymphocyte proliferation was thought to be a cause of high FDG uptake by tonsils in chronic tonsillitis.

Adolescent↗
PubMed1999

Motor innervation by enteric nerve fibers containing both nitric oxide synthase and galanin immunoreactivities in the striated muscle of the rat esophagus.

The relationship between nitric oxide synthase (NOS)- and galanin-immunoreactive nerve terminals and the origin of NOS-immunoreactive nerve terminals on the motor endplates in the striated muscles of the rat esophagus was investigated. Double immunohistochemical staining revealed a dual innervation of motor endplates by calcitonin gene-related peptide (CGRP)-immunoreactive axons and by axons that were immunoreactive for both NOS and galanin. On average, 91% of NOS terminals were galanin immunoreactive. NOS-immunoreactive fibers were revealed at 67% of endplates, identified by the presence of CGRP terminals. The left vagus and superior laryngeal nerve were cut and 15 days allowed for terminals to degenerate. This caused a significant loss of CGRP fibers, but did not affect the density of innervation of the striated muscle by NOS-immunoreactive fibers. Thus the NOS/galanin fibers are deduced to originate from ganglia in the esophageal wall. This is supported by our observation of numerous NOS-immunoreactive nerve cell bodies in the myenteric ganglia of the esophagus, 74% of which were galanin immunoreactive. There were no CGRP-immunoreactive nerve cell bodies in the wall of the esophagus.

Animals↗
PubMed1999

"White coat effect" induced by therapist's presence during speech therapy for stroke rehabilitation: a single case study.

The excessive pressor response triggered in patients by an alerting reaction to a doctor's presence has been termed the "white coat effect." A 68-year-old man with verbal apraxia after multiple lacunar infarctions was referred to the hospital for speech rehabilitation. He experienced difficulty in talking with the speech therapist during therapy sessions but not when talking with his friends or family. Because the therapist's presence was stressful to the patient, it was considered that his anxiety might produce an excessive increase in blood pressure. Blood pressure monitoring was performed during 2 separate days of speech therapies consisting of two sessions each. In one session, therapy was directed by the therapist; in the other, therapy was self-directed. The therapist-directed approach substantially increased both systolic and diastolic blood pressures, whereas the self-directed therapy slightly increased only systolic pressure. It was concluded that the excessive pressor response seen in this patient during therapist-directed speech therapy resulted from the white coat effect induced by the therapist's presence.

Aged↗