Showing papers by "Hokkaido University published in 1995"

Identification of a Member of the MAPKKK Family as a Potential Mediator of TGF-β Signal Transduction

Abstract: The mitogen-activated protein kinase (MAPK) pathway is a conserved eukaryotic signaling module that converts receptor signals into various outputs. MAPK is activated through phosphorylation by MAPK kinase (MAPKK), which is first activated by MAPKK kinase (MAPKKK). A genetic selection based on a MAPK pathway in yeast was used to identify a mouse protein kinase (TAK1) distinct from other members of the MAPKKK family. TAK1 was shown to participate in regulation of transcription by transforming growth factor-β (TGF-β). Furthermore, kinase activity of TAK1 was stimulated in response to TGF-β and bone morphogenetic protein. These results suggest that TAK1 functions as a mediator in the signaling pathway of TGF-β superfamily members.

Reduced hippocampal LTP and spatial learning in mice lacking NMDA receptor ε1 subunit

Abstract: THE NMDA (TV-methyl-D-aspartate) receptor channel is important for synaptic plasticity, which is thought to underlie learning, memory and development1, 2. The NMDA receptor channel is formed by at least two members of the glutamate receptor (GluR) channel subunit families, the GluRe (NR2) and GiuRζ (NR1) sub-unit families3–8. The four e subunits are distinct in distribution, properties and regulation5–14. On the basis of the Mg2+ sensitivity and expression patterns, we have proposed that the ei (NR2A) and e2 (NR2B) subunits play a role in synaptic plasticity6, 14. Here we show that targeted disruption of the mouse el subunit gene resulted in significant reduction of the NMDA receptor channel current and long-term potentiation at the hippocampal CA1 synapses. The mutant mice also showed a moderate deficiency in spatial learning. These results support the notion that the NMDA receptor channel-dependent synaptic plasticity is the cellular basis of certain forms of learning.

Bmpr encodes a type I bone morphogenetic protein receptor that is essential for gastrulation during mouse embryogenesis.

Abstract: Bone morphogenetic proteins (BMPs) are secreted proteins that interact with cell-surface receptors and are believed to play a variety of important roles during vertebrate embryogenesis. Bmpr, also known as ALK-3 and Brk-1, encodes a type I transforming growth factor-beta (TGF-beta) family receptor for BMP-2 and BMP-4. Bmpr is expressed ubiquitously during early mouse embryogenesis and in most adult mouse tissues. To study the function of Bmpr during mammalian development, we generated Bmpr-mutant mice. After embryonic day 9.5 (E9.5), no homozygous mutants were recovered from heterozygote matings. Homozygous mutants with morphological defects were first detected at E7.0 and were smaller than normal. Morphological and molecular examination demonstrated that no mesoderm had formed in the mutant embryos. The growth characteristics of homozygous mutant blastocysts cultured in vitro were indistinguishable from those of controls; however, embryonic ectoderm (epiblast) cell proliferation was reduced in all homozygous mutants at E6.5 before morphological abnormalities had become prominent. Teratomas arising from E7.0 mutant embryos contained derivatives from all three germ layers but were smaller and gave rise to fewer mesodermal cell types, such as muscle and cartilage, than controls. These results suggest that signaling through this type I BMP-2/4 receptor is not necessary for preimplantation or for initial postimplantation development but may be essential for the inductive events that lead to the formation of mesoderm during gastrulation and later for the differentiation of a subset of mesodermal cell types.

Loss of Apc heterozygosity and abnormal tissue building in nascent intestinal polyps in mice carrying a truncated Apc gene.

Abstract: Mutations in the APC (adenomatous polyposis coli) gene appear to be responsible for not only familial adenomatous polyposis but also many sporadic cases of gastrointestinal cancers. Using homologous recombination in mouse embryonic stem cells, we constructed mice that contained a mutant gene encoding a product truncated at a 716 (Apc delta 716). Mendelian transmission of the gene caused most homozygous mice to die in utero before day 8 of gestation. The heterozygotes developed multiple polyps throughout the intestinal tract, mostly in the small intestine. The earliest polyps arose multifocally during the third week after birth, and new polyps continued to appear thereafter. Surprisingly, every nascent polyp consisted of a microadenoma covered with a layer of the normal villous epithelium. These microadenomas originated from single crypts by forming abnormal outpockets into the inner (lacteal) side of the neighboring villi. We carefully dissected such microadenomas from nascent polyps by peeling off the normal epithelium and determined their genotype by PCR: all microadenomas had already lost the wild-type Apc allele, whereas the mutant allele remained unchanged. These results indicate that loss of heterozygosity followed by formation of intravillous microadenomas is responsible for polyposis in Apc delta 716 intestinal mucosa. It is therefore unlikely that the truncated product interacts directly with the wild-type protein and causes the microadenomas by a dominant negative mechanism.

Inhibitory Effect of Tumor Metastasis in Mice by Saponins, Ginsenoside-RB2, 20(R)- and 20(S)-Ginsenoside-Rg3, of Red Ginseng

Abstract: We examined the inhibitory effect of two saponin preparations from Red ginseng, 20(R)- and 20(S)-ginsenoside-Rg3, in comparison with that of ginsenoside-Rb2, on lung metastasis produced by two highly metastatic tumor cells, B16-BL6 melanoma and colon 26-M3.1 carcinoma, in syngeneic mice. In an in vitro analysis, both saponin preparations showed a significant inhibition of adhesion to fibronectin (FN) and laminin (LM) by B16-BL6 melanoma. Similarly, they significantly inhibited the invasion of B16-BL6 cells into the reconstituted basement membrane (Matrigel)/FN in a dose-dependent manner. In an experimental metastasis model using B16-BL6 melanoma, consecutive intravenous (i.v.) administrations of 100 micrograms/mouse of 20(R)- or 20(S)-ginsenoside-Rg3 1, 2, 3 and 4 d after tumor inoculation led to a significant decrease in lung metastasis. The inhibitory effect of i.v. administration of both ginseng saponins on the tumor metastasis of B16-BL6 melanoma was also recognized in a low dose of 10 micrograms/mouse. The oral administration (p.o.) of both saponins (100-1000 micrograms/mouse) induced a significant decrease in lung metastasis of B16-BL6 melanoma. Moreover, both ginseng saponins were effective in inhibiting of lung metastasis produced by colon 26-M3.1 carcinoma. When 20(R)- or 20(S)-ginsenoside-Rg3 was orally administered consecutively after tumor inoculation in a spontaneous metastasis model using B16-BL6 melanoma, both of them significantly inhibited lung metastasis. In the experiment involving neovasculization by tumor cells in vivo, both mice groups given each saponin preparation after tumor inoculation exhibited a significant decrease in the number of blood vessels oriented toward the tumor mass, with no repression of tumor size. These findings suggest that both ginseng saponins, 20(R)- and 20(S)-ginsenoside-Rg3, possess an ability to inhibit the lung metastasis of tumor cells, and the mechanism of their antimetastatic effect is related to inhibition of the adhesion and invasion of tumor cells, and also to anti-angiogenesis activity.

Graft Site Morbidity with Autogenous Semitendinosus and Gracilis Tendons

Abstract: To distinguish between morbidity caused by harvesting semitendinosus and gracilis tendons and morbidity associated with anterior cruciate ligament reconstruction surgery, we performed a prospective randomized study using 65 patients who underwent anterior cruciate ligament reconstruction using these tendons. The patients underwent either contralateral (N = 34) or ipsilateral (N = 31) graft harvest. For the nonoperated knees in the ipsilateral harvest group, isometric and isokinetic strength of the quadriceps and hamstring muscles increased to approximately 120% of the preoperative value at 12 months after surgery. Compared with these knees, the tendon harvest did not affect quadriceps muscle strength at all. However, harvest did decrease hamstring muscles strength for 9 months after surgery. The graft harvest in the knees with anterior cruciate ligament reconstruction also did not significantly affect quadriceps muscle strength, but it did significantly decrease hamstring muscles strength only at 1 month. Activity-related soreness at the donor site was rarely restricting and resolved by 3 months. This study demonstrated that the semitendinosus and gracilis tendon graft is a reasonable choice to minimize the donor site morbidity in ligament reconstruction using autografts.

Steam reforming of methanol over Pd/ZnO: Effect of the formation of PdZn alloys upon the reaction

Abstract: The catalytic performance of Pd/ZnO for steam reforming of methanol (CH 3 OH + H 2 O → CO 2 + 3H 2 ) was greatly improved by previously reducing the catalysts at higher temperatures. The original catalytic functions of metallic palladium were greatly modified as a result of the formation of PdZn alloys. Over the catalysts containing alloys, formaldehyde species formed in the reaction were suggested to be effectively attacked by water, being transformed into carbon dioxide and hydrogen. By contrast, the formaldehyde species decomposed selectively to carbon monoxide and hydrogen over catalysts containing metallic palladium.

A novel protein phosphatase-1 inhibitory protein potentiated by protein kinase C. Isolation from porcine aorta media and characterization.

Abstract: A novel phosphorylation-dependent inhibitory protein (IP) of porcine aorta myosin light chain phosphatase (PA-MLCP) was purified to homogeneity from porcine aorta media. The molecular mass of IP was 20 kDa. IP phosphorylated by endogenous potentiating kinase (IP-K) inhibited not only PA-MLCP activity, but also that of the catalytic subunit of protein phosphatase-1. The amino acid sequence of a peptide derived from IP phosphorylated with IP-K, RHARVT*VK, shared one of the consensus sequences phosphorylatable by protein kinase C (PKC), where T* was phosphorylated. IP was phosphorylated by PKC and the phosphorylated product inhibited PA-MLCP as strongly as IP phosphorylated with IP-K.

Role of the integument in insect defense: pro-phenol oxidase cascade in the cuticular matrix

Abstract: The cuticle of the silkworm Bombyx mori was demonstrated to contain pro-phenol oxidase [zymogen of phenol oxidase (monophenol, L-dopa:oxygen oxidoreductase, EC 1.14.18.1)] and its activating cascade. The activating cascade contained at least one serine proteinase zymogen (latent form of pro-phenol oxidase activating enzyme). When the extracted cascade components were incubated with Ca2+, the latent form of pro-phenol oxidase activating enzyme was itself activated and, in turn, converted through a limited proteolysis of pro-phenol oxidase to phenol oxidase. Immuno-gold localization of prophenol oxidase in the cuticle using a cross-reactive hemolymph anti-pro-phenol oxidase antibody revealed a random distribution of this enzyme in the nonlamellate endocuticle and a specific orderly arrayed pattern along the basal border of the laminae in the lamellate endocuticle of the body wall. Furthermore, prophenol oxidase was randomly distributed in the taenidial cushion of the tracheal cuticle. At the time of pro-phenol oxidase accumulation in the body wall cuticle, no pro-phenol oxidase mRNA could be detected in the epidermal tissue, whereas free-circulating hemocytes contained numerous transcripts of pro-phenol oxidase. Our results suggest that the pro-phenol oxidase is synthesized in the hemocytes and actively transported into the cuticle via the epidermis.

Perpetuation of influenza A viruses in Alaskan waterfowl reservoirs.

Abstract: To provide information on the mechanism of perpetuation of influenza viruses among waterfowl reservoirs in nature, virological surveillance was carried out in Alaska during their breeding season in summer from 1991 to 1994. Influenza viruses were isolated mainly from fecal samples of dabbling ducks in their nesting places in central Alaska. The numbers of subtypes of 108 influenza virus isolates were 1 H2N3, 37 H3N8, 55 H4N6, 1 H7N3, 1 H8N2, 1 H10N2, 11 H10N7, and H10N9. Influenza viruses were also isolated from water samples of the lakes where they nest. Even in September of 1994 when the most ducks had left for migration to south, viruses were still isolated from the lake water. Phylogenetic analysis of the NP genes of the representative isolates showed that they belong to the North American lineage of avian influenza viruses, suggesting that the majority of the waterfowls breeding in central Alaska migrate to North America and not to Asia. The present results support the notion that influenza viruses have been maintained in waterfowl population by water-borne transmission and revealed the mechanism of year-by-year perpetuation of the viruses in the lakes where they breed.

Collective Josephson plasma resonance in the vortex state of Bi2Sr2CaCu2O8+ delta.

Abstract: We have measured the microwave surface resistance (30-60 GHz) for different crystallographic orientations in the vortex state of ${\mathrm{Bi}}_{2}{\mathrm{Sr}}_{2}\mathrm{Ca}{\mathrm{Cu}}_{2}{\mathrm{O}}_{8+\ensuremath{\delta}}$. A sharp magnetoabsorption resonance is observed below ${T}_{c}$ when ac electric fields and magnetic fields are applied parallel to the $c$ axis (${E}_{\mathrm{ac}}\ensuremath{\parallel}B\ensuremath{\parallel}c$). We argue that the observed resonance arises from collective Josephson plasma oscillations generated by interlayer Josephson currents. From the frequency and temperature dependence of the resonance, we discuss the interlayer phase coherence in the vortex liquid and solid states quantitatively.

Weakened cellular scavenging activity against oxidative stress in diabetes mellitus: regulation of glutathione synthesis and efflux

Abstract: Glutathione functions to scavenge oxidants or xenobiotics by covalently binding them and transporting the resulting metabolites through an adenosine 5′-triphosphate-dependent transport system. It has been reported that the intracellular concentration of glutathione decreases in diabetes mellitus. In order to elucidate the physiological significance and the regulation of anti-oxidants in diabetic patients, changes in the activity of the glutathione-synthesizing enzyme, γ-glutamylcysteine synthetase, and transport of thiol [S-(2,4-dinitrophenyl)glutathione] were studied in erythrocytes from patients with non-insulin-dependent diabetes and K562 cells cultured with 27 mmol/l glucose for 7 days. The activity of γ-glutamylcysteine synthetase, the concentration of glutathione, and the thiol transport were 77%, 77% and 69%, respectively in erythrocytes from diabetic patients compared to normal control subjects. Treatment of patients with an antidiabetic agent for 6 months resulted in the restoration of γ-glutamylcysteine synthetase activity, the concentration of glutathione, and the thiol transport. A similar impairment of glutathione metabolism was observed in K562 cells with high glucose levels. The cytotoxicity by a xenobiotic (1-chloro-2,4-dinitrobenzene) was higher in K562 cells with high glucose than in control subjects (50% of inhibitory concentration. 300±24 Μmol/l vs 840±29 Μmol/l, p<0.01). Expression of γ-glutamylcysteine synthetase protein was augmented in K562 cells with high glucose, while enzymatic activity and expression of mRNA were lower than those in the control subjects. These results suggest that inactivation of glutathione synthesis and thiol transport in diabetic patients increases the sensitivity of the cells to oxidative stresses, and these changes may lead to the development of some complications in diabetes mellitus.

Two distinct oscillators in the rat suprachiasmatic nucleus in vitro

Abstract: In the rat suprachiasmatic nucleus slice culture, circadian rhythms in the release of arginine vasopressin and vasoactive intestinal polypeptide were measured simultaneously and longitudinally. The phase relationship between the two peptide rhythms was relatively constant in the culture without a treatment of antimitotic drugs but became diverse by an introduction of antimitotics, which is generally used to reduce the number of glial cells. By monitoring the two rhythms continuously for 6 days, different periods were detected in culture with the antimitotic treatment. Furthermore, N-methyl-D-aspartate shifted the phase of the two peptide rhythms in the same culture differently. These results indicate that the arginine vasopressin and vasoactive intestinal polypeptide release are under control of different circadian oscillators.

Optical Microfabrication of Chalcogenide Glasses

Abstract: It was found that chalcogenide glasses can be shaped by stressing the glass under light illumination because light illumination enhances the fluidity of the glass. The mechanism of photoinduced fluidity was found to be photoelectronic, that is, athermal. The process can be applied to microfabrication of optical fibers and glassy films with a typical dimension of 10 to 100 micrometers.

Gelsolin: a candidate for suppressor of human bladder cancer

Abstract: Human transitional cell carcinomas of the bladder frequently reveal chromosomal abnormalities that span a range between chromosome 9p12 and 9qter, even at early stages of bladder carcinogenesis. Because the gene that encodes an actin-regulatory protein, gelsolin, is localized in chromosome 9q33, we examined the expression of gelsolin in a number of human bladder cancer cell lines and tissues. In all 6 cell lines and in 14 of the 18 tumor tissues (77.8%), gelsolin expression was undetectable or extremely low in comparison with its expression in normal bladder epithelial cells. Furthermore, upon the introduction of the exogenous human or mouse authentic gelsolin cDNA into a human bladder cancer cell line, UMUC-2, gelsolin transfectants of UMUC-2 greatly reduced the colony-forming ability and the tumorigenicity in vivo. These results suggest that gelsolin plays a key role as a tumor suppressor in human urinary bladder carcinogenesis.

Flexion-relaxation phenomenon in the back muscles. A comparative study between healthy subjects and patients with chronic low back pain

Abstract: At a certain position of trunk flexion, there is a sudden onset of electrical silence in back muscles. This is called "flexion-relaxation (F-R) phenomenon." The goals of this study were (1) to evaluate the relationship between flexion angle and activity of back muscles during flexion movement and (2) to determine what the difference is between healthy subjects and patients with chronic low back pain (CLBP). Twenty-five healthy subjects (13 males and 12 females; average age, 28.3 yr) and 20 patients with CLBP (12 males and 8 females; average age, 34.1 yr) volunteered for this study. The subjects were asked to flex forward maximally from the erect position and to maintain full flexion, followed by returning to the initial upright position. Flexion angle of trunk and hip was measured during the examination. Electromyographic activity of erector spinae was also monitored simultaneously. F-R phenomenon was observed in all healthy subjects before reaching the maximum flexion. Electrical silence continued even after extending the trunk began. In contrast, no patients with CLBP demonstrated F-R phenomenon. A significant difference in muscular activities of erector spinae between the groups was obtained when returning to the erect position from the maximum flexion. Moreover, time lag between trunk and hip movement was much greater in patients than in healthy subjects. This study demonstrated that neuromuscular coordination between trunk and hip could be abnormal in patients with CLBP.

Template-switching during DNA synthesis by Thermus aquaticus DNA polymerase I

Abstract: Recombinant DNA molecules are often generated during the polymerase chain reaction (PCR) when partially homologous templates are available [e.g., see Paabo et al. (1990) J. Biol. Chem. 265, 4718-4721]. It has been suggested that these recombinant molecules are a consequence of truncated extension products annealing to partially homologous templates on subsequent PCR cycles. However, we demonstrate here that recombinants can be generated during a single round of primer extension in the absence of subsequent heat denaturation, indicating that template-switching produces some of these recombinant molecules. Two types of template-switches were observed: (i) switches to pre-existing templates and (ii) switches to the complementary nascent strand. Recombination is reduced several fold when the complementary template strands are physically separated by attachment to streptavidin magnetic beads. This result supports the hypothesis that either the polymerase or at least one of the two extending strands switches templates during DNA synthesis and that interaction between the complementary template strands is necessary for efficient template-switching.

Symmetry, degeneracy, and uncoupled modes in two-dimensional photonic lattices

Abstract: The photonic bands of two-dimensional triangular and square lattices composed of circular rods were classified by means of the group theory based on the symmetry of the lattice structure. According to this classification, it was shown that the uncoupled mode, or the mode that cannot be excited by an external plane wave, which we previously found for the triangular lattice by the numerical calculation of the transmittance, is an antisymmetric mode under the relevant mirror reflection, and this fact is consistent with the observation by Robertson et al. It was also shown that triangular and square lattices with ${\mathit{C}}_{6\mathit{v}}$ or ${\mathit{C}}_{4\mathit{v}}$ symmetry have many other uncoupled modes with relatively low eigenfrequencies and some of them can be easily identified as the spectral ranges of total reflection in spite of their nonzero state density.

Exchange Monte Carlo Method and Application to Spin Glass Simulations

Abstract: We propose an efficient Monte Carlo algorithm for simulating a ``hardly-relaxing" system, in which many replicas with different temperatures are simultaneously simulated and a virtual process exchanging configurations of these replica is introduced. This exchange process is expected to let the system at low temperatures escape from a local minimum. By using this algorithm the three-dimensional $\pm J$ Ising spin glass model is studied. The ergodicity time in this method is found much smaller than that of the multi-canonical method. In particular the time correlation function almost follows an exponential decay whose relaxation time is comparable to the ergodicity time at low temperatures. It suggests that the system relaxes very rapidly through the exchange process even in the low temperature phase.

Effects of conditioned fear stress on 5-HT release in the rat prefrontal cortex.

Abstract: The effects of conditioned fear stress (CFS) on 5-HT release in the medial prefrontal cortex were studied by in vivo microdialysis. CFS (exposure to an environment in which foot-shock had been delivered previously) induced a marked suppression of motility-that is, freezing behavior. The extracellular concentration of 5-HT in the medial prefrontal cortex increased during this freezing behavior, but no significant changes were observed in the concentration of its metabolite, 5-HIAA. The increased 5-HT concentration returned to pretreatment levels when the animals were returned to their home cages. Diazepam (0.5 mg/kg, intraperitoneally) reduced the CFS-induced freezing behavior and prevented the increases in extracellular 5-HT levels. A 5-HT3 receptor antagonist, tropisetron (10 and 100 micrograms/kg), also inhibited both the CFS-induced increase in 5-HT release and the freezing behavior. These findings suggest that there is a relationship between anxiety and 5-HT release in the prefrontal cortex and that the 5-HT3 receptor antagonist tropisetron might have anxiolytic properties.

Tsunami deposits from the 1993 Southwest Hokkaido earthquake and the 1640 Hokkaido Komagatake eruption, northern Japan

Abstract: The southwest Hokkaido tsunami of July 12th, 1993, left continuous onshore sand deposits along the west coast of Oshima Peninsula, Hokkaido, northern Japan. We investigated spatial distribution and lithofacies of the new tsunami deposits for its identification of ancient tsunami deposits. An eyewitness account and bent plants helped our interpretation of the onshore tsunami behavior. We regard the following properties as typical of the coastal tsunami sand deposits: (1) The deposits cover the surface almost continuously on gentle topography. (2).Deposit thicknesses and mean grain sizes decrease with distance from the sea. (3) Deposit thicknesses and lithofacies vary greatly across local surface undulation. (4) Graded bedding reflecting tsunami runup and backwash is present in thick deposits. (5) The deposits are widely distributed along the coast and extend inland several tens of meters to 100 m. We examined a candidate for the paleo-tsunami deposits associated with the 1640 Komagatake eruption, and confirmed that the similar patterns are typical of ancient tsunami deposits.