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Biomedical subjects

T Ueno

Publications and source records attributed to T Ueno.

At least 91 records · Page 5Linked to original sources
PubMed2001

Prefabricated bone graft induced from grafted periosteum for the repair of jaw defects: an experimental study in rabbits.

PURPOSE: The purpose of the study was to determine whether a prefabricated graft of new bone induced from periosteum grafted into muscle was an effective material for the repair of jaw defects. MATERIAL AND METHODS: Artificial mandibular jaw defects in young Japanese rabbits were covered either with free grafted periosteum (n = 5) or a prefabricated graft of newly formed bone induced from periosteum, which was first grafted into the floor of the mouth, and placed as a revascularized muscle-pedicled bone flap (n = 5). Bone formation in jaw defects was examined radiographically and histologically 28 days after grafting into defects. RESULTS: Bone formation was confirmed radiographically and histologically in both groups. However, the free grafted periosteum formed thin bone and fibrous tissue existed between the new and the original bone. In contrast, more active bone formation was observed with the prefabricated graft. This grafted new bone developed further and fused to the mandible. Blood vessels surrounding the new bone were observed histologically. CONCLUSION: These experimental findings suggested that prefabricated bone grafts induced from periosteum grafts are potentially useful for correction of jaw defects.

Animals↗
PubMed2001

Nuclear factor-kappa B decoy attenuates neuronal damage after global brain ischemia: a future strategy for brain protection during circulatory arrest.

OBJECTIVES: Recent studies have reported that cis element decoy oligodeoxynucleotides against nuclear factor-kappa B block the activation of genes that mediate ischemic injury. To improve brain protection during circulatory arrest in cardiac surgery, we evaluated the efficacy of nuclear factor-kappa B decoy oligodeoxynucleotides in preventing neuronal damage after global brain ischemia. METHODS: Hemagglutinating virus of Japan-liposome complex with fluorescein isothiocyanate-labeled nuclear factor-kappa B decoy oligodeoxynucleotides was injected through the carotid artery during 20 minutes of global brain ischemia in rats to evaluate the efficacy of transfecting the decoy oligodeoxynucleotides. The messenger RNA levels of several factors related to ischemia-reperfusion injury in the hippocampus were estimated by a real-time polymerase chain reaction method 1 hour after reperfusion. Neuronal damage was evaluated by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling staining and by using immunohistochemical study of microtubule-associated protein 2 in the hippocampus CA-1 region 7 days after ischemia. RESULTS: Introduction of the nuclear factor-kappa B decoy oligodeoxynucleotides into rat brain neurons through the carotid artery during global brain ischemia was markedly successful. The polymerase chain reaction study showed that the transfected nuclear factor-kappa B decoy oligodeoxynucleotides effectively inhibited the expression of tumor necrosis factor alpha interleukin 1 beta and intracellular adhesion molecule 1 messenger RNA 1 hour after global brain ischemia. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling staining and microtubule-associated protein 2 immunohistochemistry showed that the transfected nuclear factor-kappa B decoy oligodeoxynucleotides significantly attenuated the neuronal damage 7 days after global brain ischemia. CONCLUSIONS: Therapeutic transfection of nuclear factor-kappa B decoy oligodeoxynucleotides during brain ischemia may be useful for attenuating neuronal damage, suggesting a strategy for cerebral protection against global ischemia.

Animals↗
PubMed2001

Effect of glucagon, glicentin, glucagon-like peptide-1 and -2 on interdigestive gastroduodenal motility in dogs with a vagally denervated gastric pouch.

BACKGROUND: We previously reported that inhibition of gastric motility and hypertrophy of the small intestinal mucosa were observed after ileo-jejunal transposition which induced hypersecretion of enteroglucagon. Our aim was to study the effect of four enteroglucagon-related peptides (glucagon, glucagon-like peptide (GLP)-1, -2 and glicentin) on gastroduodenal motility and their mechanisms of action. METHODS: The effect of these four peptides on motilin-induced interdigestive contractions was studied in dogs with vagally denervated gastric pouches equipped with four strain gauge force transducers on the pouch, gastric body, antrum and duodenum. Whether or not nitric oxide synthase inhibitor or phentolamine and propranolol reverses the inhibitory effect of those peptides was also studied. RESULTS: Glucagon inhibited contractions in the pouch and stomach but had no effect on duodenal contractility. GLP-1 inhibited contractions at all sites. GLP-2 inhibited contractions in the pouch but did not affect motility in the neurally intact gastroduodenum. Glicentin had no effect on contractions at any site. Pretreatment with either a nitric oxide synthase inhibitor or phentolamine and propranolol reversed the inhibitory effect of glucagon, GLP-1 and GLP-2 on contractions in the pouch, but did not alter the inhibitory effect of glucagon and GLP-1 on motility in the neurally intact stomach and duodenum. CONCLUSIONS: These results suggest that the effects of four peptides on gastroduodenal motility differ, and changes occur in the enteric neural modulation of motor activity after chronic surgical extrinsic denervation.

Animals↗
PubMed2001

Leupeptin-induced appearance of partial fragment of betaine homocysteine methyltransferase during autophagic maturation in rat hepatocytes.

A cytosolic enzyme, betaine homocysteine methyltransferase (BHMT), and its partial fragments were discovered as autolysosomal membrane proteins from rat liver in the presence of leupeptin [Ueno et al. (1999) J. Biol. Chem. 274, 15222-15229]. The present study was undertaken to further characterize the transport and processing of BHMT from cytosol to autolysosome and to test if the fragment can be used as an in vitro probe for the maturation step of macroautophagy. Upon subcellular fractionation, BHMT (p44) was found in all fractions, while its 32-kDa fragment (p32) was found only in the mitochondrial-lysosomal (ML) fraction. Incubation of isolated hepatocytes with leupeptin induced time-dependent accumulation of p32 in the ML fraction from 30 to 90 min after the start of incubation. However, chloroquine completely inhibited the appearance of p32, indicating that the processing from p44 to p32 is lysosomal. Incubation with Bafilomycin A(1), a vacuolar H(+)-ATPase inhibitor, together with leupeptin, led to linear accumulation of p44, but not of p32. The p44 accumulation rate was calculated to be 4.9%/h, which was comparable to autophagic sequestration rate. The distribution of p44 within the ML fraction turned out to be dual, i.e., the membrane-surface attached and luminal/sedimentable forms. Amino acids and 3-methyladenine, both of which specifically suppress macroautophagy, inhibited the accumulation of p32 as well as of p44. Finally, energy-dependent appearance of p32 was demonstrated during incubation of postnucler supernatant fractions, making it possible to establish an in vitro assay system. All the results strongly support the idea that BHMT is taken up and degraded to p32 through the macroautophagic pathway, and that p32 could be a novel probe for the maturation of macroautophagy.

Animals↗
PubMed2001

Organ-specific autoimmunity in mice whose T cell repertoire is shaped by a single antigenic peptide.

Organ-specific autoimmune diseases have been postulated to be the result of T cell response against organ-specific self-peptides bound to MHC molecules. Contrary to this paradigm, we report here that transgenic mice lacking MHC class I expression and expressing an MHC class II I-A(b) molecule that presents only a single peptide (E alpha 52-68) spontaneously develops peripheral nervous system-specific autoimmune disease with many of the histopathological features found in experimental allergic neuritis. Reciprocal bone marrow chimeras produced using susceptible and resistant lines revealed that bone marrow-derived cells determined disease susceptibility. While the expression of the I-A(b)-E alpha 52-68 complex in the periphery was readily detectable in both lines, its expression on thymic dendritic cells responsible for tolerance induction was markedly lower in the susceptible line than in the resistant line. Consistent with this, CD4(+) T cells that can be activated by the I-A(b)-E alpha 52-68 complex were found in the susceptible line, but not in the resistant line. Such CD4(+) T cells conferred the disease to the resistant line by adoptive transfer, and administration of Ab specific for the I-A(b)-E alpha 52-68 complex inhibited disease manifestation in the susceptible line. These results indicate that disease development involves systemic T cell reactivity to I-A(b)-E alpha 52-68 complex, probably caused by incomplete negative thymocyte selection.

Animals↗
PubMed2001

Insulin and somatostatin releasing islet cell tumor caused hypoglycemia.

We report a hypoglycemic case with normal insulin levels, which was caused by an islet cell tumor that was releasing insulin and somatostatin. A fasting test suggested the over secretion of insulin. Moreover, this hypoglycemia was enhanced by the inhibitory effect of somatostatin on the secretion of insulin counter-regulatory hormones, such as glucagon, in addition to the autonomous secretion of insulin from the tumor. In cases of hypoglycemia with apparently normal insulin levels, the measurement of somatostatin and various provocative tests are recommended. Arterial stimulation venous sampling (ASVS) was useful to detect the location of this functioning islet cell tumor.

Adenoma, Islet Cell↗
PubMed2001

Relation of type II transforming growth factor-beta receptor to hepatic fibrosis and hepatocellular carcinoma.

Hepatocarcinogenesis is closely related to hepatic fibrosis. In this study, we investigated the relationship of type II transforming growth factor-beta receptor (T beta RII) to hepatic fibrosis and hepatocellular carcinoma (HCC). In vivo: liver tissues were obtained from 30 patients (10 chronic hepatitis, 7 cirrhosis, 13 HCC). Protein expression and immunolocalization of T beta RII were examined by Western blot analysis and immunohistochemistry. In vitro: T beta RII protein expression in hepatoma cell lines (HepG2, Hep3B, HLE, HLF and Huh7) was examined by Western blot analysis. Next, we transfected T beta RII cDNA to Huh7, and compared the change of cell number and observed the induction of apoptosis after TGF-beta1 treatment using a FACScan flow cytometer. In vivo: T beta RII immunolocalization in liver tissues was significantly decreased in patients with HCC compared with that of patients with chronic hepatitis or liver cirrhosis. In Western blot analysis, T beta RII expression in tissues attenuated in comparison with that in non-tumor tissues in some patients with HCC. In vitro: T beta RII protein expression in HLE, HLF and Huh7 cells was weaker than that in HepG2 and Hep3B cells. In Huh7 cells transfected T beta RII cDNA, cell arrest and apoptosis were obviously induced. These results indicated that human HCC has a reduced expression of T beta RII for TGF-beta1. This may provide a selective growth advantage to HCC to escape the inhibitory growth signals of TGF-beta1, and may be linked with critical steps in the growth of hepatoma cells.

Aged↗
PubMed2001

[An approach for the quality control in clinical microbiology laboratory by antibiotic susceptibility].

It is important to develop a system for the prevention of various errors and the maintenance of the high quality level in the department of laboratory medicine. We developed a computer program to infer the species of microorganisms by assessing the minimal inhibitory concentration of respective microorganisms to various antibiotics, antibiotic susceptibility patterns(ASP). Retrospective analysis of 3,405 strains from 39 species using this program resulted in a sensitivity of 91.4% and specificity of 98.9%. Furthermore, in 1,660 strains isolated from September 1999 to August 2000, the efficacy of this program was evaluated using both this program and conventional identifying method such as VITEK, api and ATB. The discrepancy between this inferring method and conventional method was noted in 179 strains, revealing errors in 12 strains. We conclude that this ASP inferring program is useful for preventing errors and the maintenance of high quality examination by combining with conventional methods in the laboratory.

Anti-Bacterial Agents↗
PubMed2001

[Acute myocardial infarction in the postoperative period in an elderly man with low coronary risk factor before the operation].

An 86-year-old man complicated by severe asthma developed myocardial infarction after the operation of abdominal aortic aneurysm. Emergency coronary angiography (CAG) revealed 99% stenosis of main trunks of left coronary artery (LMT). Because the patient had been without major coronary risk factors, it was difficult to predict coronary lesions preoperatively. Elderly patients have frequently latent complications not detected by usual consultation. Therefore, in elderly patients, we have to evaluate each individual risk factor more rigidly, and perform more thorough preoperative tests, including CAG.

Aged↗
PubMed2001

[Anterior chest wall reconstruction with myocutaneous flap after open drainage for infectious bulla and empyema following T4 lung cancer resection via anterior approach].

A 48-year-old man with T4 lung cancer underwent resection of the right upper lobe, chest wall, and superior vena cava, and reconstruction of the last 2 tissues via anterior approach after induction chemo-radiotherapy. However, a bulla in the apical segment of the lower lobe divided by stapling was infected after the operation, and open drainage through the anterior chest wall was mandatory due to massive air leak and spread of infection to the pleural space. As the incision were fortunately restricted in the anterior chest, he could undergo successful reconstruction of a full-thickness defect of the anterior chest wall using a serratus anterior muscle flap and a myocutaneous flap of lattissimus dorsi with minimum rib resection after 2 months. He has in regular work with a small limitation of raising the right upper extremitus these 7 years.

Carcinoma, Large Cell↗
PubMed2000

Interaction of the Unc-51-like kinase and microtubule-associated protein light chain 3 related proteins in the brain: possible role of vesicular transport in axonal elongation.

We identified two mammalian ULK1 (Unc-51-like kinase involved in neurite extension) binding proteins by yeast two-hybrid screening. Both proteins showed high structural similarity to microtubule-associated protein (MAP) light chain 3 (LC3). One is identical to the Golgi-associated ATPase Enhancer of 16 kDa (GATE-16), an essential factor for intra-Golgi transport [39]. The other is identical to the gamma 2-subunit of GABA-A receptor associated protein (GABARAP) which has a possible role in receptor transport [46]. Using the yeast two-hybrid system and the in vitro GST pull-down assay, we found that the N-terminal proline/serine rich (PS) domain of ULK1 (amino acid 287-416) is required for ULK1-GATE-16 and ULK1-GABARAP protein interactions. However, the kinase activity of ULK1 affected neither ULK1-GATE-16 nor ULK1-GABARAP interaction. Immunohistochemical analysis using ULK1 and GABARAP antibodies showed that the ULK1 and the GABARAP proteins co-localized to many kind of neurons such as pyramidal cells of the hippocampus, mitral cells of the olfactory bulb, and Purkinje cells of the cerebellum. In HeLa cells, endogenous ULK1 and tagged GABARAP showed punctate structures in the cytosol, and were colocalized. These results suggest that the interaction of ULK1 and GABARAP is important to vesicle transport and axonal elongation in mammalian neurons.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Autophagosome-associated variant isoforms of cytosolic enzymes.

In a search for autophagosome-associated proteins, two-dimensional gel separations of proteins from purified autophagosomes, postnuclear supernatant, cytosol, lysosomes, mitochondria, endosomes and a cytomembrane fraction (mostly endoplasmic reticulum) were compared. Three proteins, with monomeric molecular masses of 43, 35 and 31 kDa, were enriched in total or sedimentable fractions of autophagosomes relative to the corresponding fractions of postnuclear supernatant, suggesting an association with the autophagosomal delimiting membrane. These proteins were also present on lysosomal membranes, but they were absent from mitochondria, and detected only in small amounts in the cytomembrane fraction and in endosomes, indicating that they were not associated with organelles sequestered by autophagy. However, all three proteins were present in the cytosol, suggesting that they were cytosolic proteins binding peripherally to the delimiting membrane of autophagosomes, probably to its innermost surface as indicated by their resistance to treatment of intact autophagosomes with proteinase or protein-stripping agents. Amino acid sequencing identified these proteins as an isoform of argininosuccinate synthase, an N-truncated variant of glyceraldehyde-3-phosphate dehydrogenase, and a sequence variant of short-chain 2-enoyl-CoA hydratase.

Amino Acid Sequence↗
PubMed2000

CDKN2A germ-line mutations in individuals with multiple cutaneous melanomas.

Germ-line CDKN2A mutations are present in some kindreds with hereditary cutaneous melanoma, and in Sweden a founder mutation with an extra arginine in codon 113 (113insR) has been identified. We screened 80 individuals with at least two primary cutaneous melanomas, who were identified mainly by a search of a regional cancer registry, for germ-line CDKN2A mutations. In nine patients, CDKN2A alterations that may contribute to melanoma predisposition were detected. In six individuals with a family history of melanoma, the 113insR founder mutation was present. One patient, who also had a family history of melanoma, had a 24-bp deletion that included codons 62-69. An in vitro binding assay established that the resulting mutant p16 protein was unable to bind cyclin-dependent kinase 4 and cyclin-dependent kinase 6. Two patients without a family history of melanoma had CDKN2A alterations: (a) one had a mutation in the 5' noncoding sequence (-14C/T); and (b) the other had an insertion of an extra T in codon 28, which results in a stop signal in codon 43. The median age at diagnosis of the first melanoma was significantly lower, the number of primary melanomas was significantly higher, and the presence of a family history of melanoma was significantly more common in patients with CDKN2A mutations than in those without germ-line mutations. The proportion of CDKN2A mutation carriers was significantly higher among patients treated for three or more primary melanomas compared with those with two tumors only. We conclude that mutation screening of individuals with multiple primary melanomas is a useful strategy to identify new melanoma kindreds with CDKN2A germ-line mutations.

Adolescent↗
PubMed2000

The human homolog of Saccharomyces cerevisiae Apg7p is a Protein-activating enzyme for multiple substrates including human Apg12p, GATE-16, GABARAP, and MAP-LC3.

Autophagy is a process that involves the bulk degradation of cytoplasmic components by the lysosomal/vacuolar system. In the yeast, Saccharomyces cerevisiae, an autophagosome is formed in the cytosol. The outer membrane of the autophagosome is fused with the vacuole, releasing the inner membrane structure, an autophagic body, into the vacuole. The autophagic body is subsequently degraded by vacuolar hydrolases. Taking advantage of yeast genetics, apg (autophagy-defective) mutants were isolated that are defective in terms of formation of autophagic bodies under nutrient starvation conditions. One of the APG gene products, Apg12p, is covalently attached to Apg5p via the C-terminal Gly of Apg12p as in the case of ubiquitylation, and this conjugation is essential for autophagy. Apg7p is a novel E1 enzyme essential for the Apg12p-conjugation system. In mammalian cells, the human Apg12p homolog (hApg12p) also conjugates with the human Apg5p homolog. In this study, the unique characteristics of hApg7p are shown. A two-hybrid experiment indicated that hApg12p interacts with hApg7p. Site-directed mutagenesis revealed that Cys(572) of hApg7p is an authentic active site cysteine residue essential for the formation of the hApg7p.hApg12p intermediate. Overexpression of hApg7p enhances the formation of the hApg5p.hApg12p conjugate, indicating that hApg7p is an E1-like enzyme essential for the hApg12p conjugation system. Cross-linking experiments and glycerol-gradient centrifugation analysis showed that the mammalian Apg7p homolog forms a homodimer as in yeast Apg7p. Each of three human Apg8p counterparts, i.e. the Golgi-associated ATPase enhancer of 16 kDa, GABA(A) receptor-associated protein, and microtubule-associated protein light chain 3, coimmunoprecipitates with hApg7p and conjugates with mutant hApg7p(C572S) to form a stable intermediate via an ester bond. These results indicate that hApg7p is an authentic protein-activating enzyme for hApg12p and the three Apg8p homologs.

Adaptor Proteins, Signal Transducing↗
PubMed2000

LC3, a mammalian homologue of yeast Apg8p, is localized in autophagosome membranes after processing.

Little is known about the protein constituents of autophagosome membranes in mammalian cells. Here we demonstrate that the rat microtubule-associated protein 1 light chain 3 (LC3), a homologue of Apg8p essential for autophagy in yeast, is associated to the autophagosome membranes after processing. Two forms of LC3, called LC3-I and -II, were produced post-translationally in various cells. LC3-I is cytosolic, whereas LC3-II is membrane bound. The autophagic vacuole fraction prepared from starved rat liver was enriched with LC3-II. Immunoelectron microscopy on LC3 revealed specific labelling of autophagosome membranes in addition to the cytoplasmic labelling. LC3-II was present both inside and outside of autophagosomes. Mutational analyses suggest that LC3-I is formed by the removal of the C-terminal 22 amino acids from newly synthesized LC3, followed by the conversion of a fraction of LC3-I into LC3-II. The amount of LC3-II is correlated with the extent of autophagosome formation. LC3-II is the first mammalian protein identified that specifically associates with autophagosome membranes.

Amino Acid Sequence↗
PubMed2000

The biochemical mode of inhibition of DNA polymerase beta by alpha-rubromycin.

Quinone antibiotics, alpha- and beta-rubromycin, were originally found as inhibitors of retroviral reverse transcriptase. We investigated the effects of these agents on DNA metabolic enzymes including DNA and RNA polymerases as retroviral reverse transcriptase is a kind of the polymerase. As expected, we found that alpha- and beta-rubromycin strongly inhibited not only the retroviral reverse transcriptase activity, but the activities of the mammalian DNA polymerases, telomerase and terminal deoxynucleotidyl transferase in vitro. These agents should therefore be classified as DNA polymerase inhibitors. The Ki values of alpha-rubromycin against nucleotide substrate were 0.66 and 0.17 microM for DNA polymerase alpha and beta (pol. alpha and beta), respectively, and those of beta-rubromycin was 2.40 and 10.5 microM, respectively. Alpha-rubromycin strongly inhibited the pol. beta activity, and showed the strongest pol. beta inhibitory effect reported to date. At least on pol. beta, alpha-rubromycin was suggested to bind to the active region competing with the nucleotide substrate, and subsequently inhibit the catalytic activity. alpha-Rubromycin directly competed with the nucleotide substrate, and indirectly but simultaneously and non-competitively disturbed the template-DNA interaction with pol. beta.

Animals↗
PubMed2000

A transgenic mouse expressing human CYP1A2 in the pancreas.

A transgenic mouse line expressing the human cytochrome P450 CYP1A2 in the pancreas under the control of the mouse elastase promoter was established. The expression of CYP1A2 was specific to the transgenic pancreas and was not found in the control wild-type mouse pancreas. The level of CYP1A2 expressed in pancreatic microsomes from transgenic mice was comparable to that of the endogenously expressed CYP1A2 protein in the liver, as judged by western blotting analyses. Estrone metabolism was used to determine the activity of CYP1A2 expressed in the pancreas of the transgenic mouse. The transgenic pancreas exhibited almost one-third to one-half of the activity of wild-type or CYP1A2 transgenic mouse liver, whereas the wild-type pancreas demonstrated no activity. The addition of NADPH-cytochrome P450 oxidoreductase to the reaction mixture containing pancreatic microsomes from the transgenic mice did not increase the estrone metabolism activity significantly. This transgenic mouse line provides another useful tool to study human CYP1A2 and its relation to chemical toxicity and carcinogenesis.

Animals↗