Biomedical subjects

A Yamazaki

Publications and source records attributed to A Yamazaki.

At least 73 records · Page 4Linked to original sources

PubMed1996

Experimental autoimmune uveoretinitis induced by the gamma-subunit of cyclic guanosine monophosphate phosphodiesterase in rats.

PURPOSE: To investigate the capacity of the recombinant gamma-subunit (P gamma) of cyclic guanosine monophosphate phosphodiesterase to induce experimental autoimmune uveoretinitis in Lewis rats. METHODS: Bovine P gamma was expressed in Escherichia coli cells and purified by fast protein liquid chromatography. Lewis rats were immunized by a single footpad injection of P gamma emulsified in complete Freund's adjuvant. Clinical and histopathologic changes in the eye and pineal gland were examined. Lymphocytes were prepared from the lymph nodes of rats with uveitis and transferred by intraperitoneal injection to naive recipient rats. RESULTS: Immunization of rats with P gamma induced panuveitis and pinealitis with clinical and histopathologic changes similar to those induced by S-antigen. Lymphocytes from the lymph nodes of diseased rats transferred uveitis to naive recipients. CONCLUSIONS: P gamma, a retina-specific protein of molecular weight less than 10,000 kDa, is capable of inducing uveoretinitis in Lewis rats. The disease can be transferred adoptively to naive rats by injection of lymphocytes from donor rats with experimental autoimmune uveoretinitis. Inflammation of the pineal gland of immunized rats suggests that P gamma is not only localized to the retina but also to the pineal gland.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed1996

Cytotoxicity of synthetic barium hydroxyapatite.

Barium hydroxyapatite (Ba10(PO4)6(OH)2, Ba-HAp) was synthesized by a wet method using Ba(OH)2.8H2O and (NH4)2HPO4 as starting materials. The Ba-HAp obtained had a Ba/P atomic ratio of 1.76 and contained CO3 groups. The Ba-HAp was sintered at 1073 K for 12 hours. The sintered Ba-HAp had a three point bending strength of 29 MPa and Young's modulus of 27 GPa. Cytotoxicity of the sintered bodies and particles was tested using L-cells. The sintered Ba-HAp showed no cytotoxicity, and the cells were closely in contact with the surfaces of sintered Ba-HAp. Morphological observation of the cell around the Ba-HAp particles also showed no cytotoxicity. However, cell growth was inhibited by Ca adsorption on the Ba-HAp particles. These results suggested that the Ba-HAp had no cytotoxicity and can be applied as a bioactive X-ray opaque material.

Adsorption↗

PubMed1995

Protective immune responses induced by secretion of a chimeric soluble protein from a recombinant Mycobacterium bovis bacillus Calmette-Guérin vector candidate vaccine for human immunodeficiency virus type 1 in small animals.

A recombinant Mycobacterium bovis bacillus Calmette-Guérin (BCG) vector-based vaccine that secretes the V3 principal neutralizing epitope of human immunodeficiency virus (HIV) could induce immune response to the epitope and prevent the viral infection. By using the Japanese consensus sequence of HIV-1, we successfully constructed chimeric protein secretion vectors by selecting an appropriate insertion site of a carrier protein and established the principal neutralizing determinant (PND)-peptide secretion system in BCG. The recombinant BCG (rBCG)-inoculated guinea pigs were initially screened by delayed-type hypersensitivity (DTH) skin reactions to the PND peptide, followed by passive transfer of the DTH by the systemic route. Further, immunization of mice with the rBCG resulted in induction of cytotoxic T lymphocytes. The guinea pig immune antisera showed elevated titers to the PND peptide and neutralized HIVMN, and administration of serum IgG from the vaccinated guinea pigs was effective in completely blocking the HIV infection in thymus/liver transplanted severe combined immunodeficiency (SCID)/hu or SCID/PBL mice. In addition, the immune serum IgG was shown to neutralize primary field isolates of HIV that match the neutralizing sequence motif by a peripheral blood mononuclear cell-based virus neutralization assay. The data support the idea that the antigen-secreting rBCG system can be used as a tool for development of HIV vaccines.

AIDS Vaccines↗

PubMed1995

Posterior migration of cervical spinal cord between split laminae as a complication of laminoplasty.

STUDY DESIGN: This is a case report. OBJECTIVE: To focus attention on a possible neurologic complication of laminoplasty. SUMMARY OF BACKGROUND DATA: An iatrogenic spinal cord lesion developed after laminoplasty as an operative complication of unknown cause. METHODS: Two patients had iatrogenic damage to the spinal cord from an unknown cause after laminoplasty of the cervical spine, but recovered soon after an additional laminectomy. They were evaluated with regard to the mechanism of damage and prevention. RESULTS: The spinal cord damage occurred when the spinal cord migrated posteriorly and deformed between the split laminae at the apex of the lordosis, where the maximum posterior displacement of the spinal cord was achieved. CONCLUSION: This damage was caused by a relatively narrower canal resulting from the split laminae of the apex of the adjacent laminae.

Aged↗

PubMed1995

Delayed-type hypersensitivity to a recombinant mycobacterial antigen, MPB64, in guinea pigs sensitized to Mycobacterium tuberculosis or Mycobacterium bovis BCG.

Recombinant MPB64 (rMPB64), a mycobacterial antigen, was obtained from an Escherichia coli clone transformed with a recombinant expression vector, pMAL64c. The rMPB64 was examined for the activity to elicit delayed-type hypersensitivity (DTH) in guinea pigs injected with liver Mycobacterium tuberculosis H37Rv or live M. bovis BCG Tokyo. It was found that rMPB64 has the same reactivity as native MPB64 (nMPB64) or MPT64 (nMPT64) and the potency to elicit DTH was 13.4 times higher than that of PPD. Because MPB64 is secreted only by living M. tuberculosis and some strains of BCG, it is possible to use this antigen for the diagnosis of tuberculosis.

Amino Acid Sequence↗

PubMed1995

Potentiation of retinoic acid-induced U-937 differentiation into respiratory burst-competent cells by nitric oxide donors.

All-trans retinoic acid (tretinoin) is a known inducer of differentiation of the human monoblastic cell line, U-937. We now report that the ability of retinoic acid (RA) to induce differentiation of U-937 cells into cells possessing respiratory burst activity is enhanced by the known nitric oxide-donating drugs glyceryl trinitrate, molsidomine and CAS 936, and by tetranitromethane in combination with cysteine. RA alone was a strong inducer of U-937 differentiation as indicated by the following responses to 12-O-tetradecanoylphorbol-13-acetate (TPA) stimulation: (1) increase in the percentage of cells staining with nitroblue tetrazolium (NBT); (2) increase in the total amount of formazan (the product of NBT reduction by O2-.) as determined spectrophotometrically; (3) increase in hexose monophosphate shunt (HMPS) activity as assessed by [14C]CO2 released from D-[1-14C]glucose. RA was also able to increase mRNA levels for two respiratory burst-related genes and for glucose-6-phosphate dehydrogenase (G6PD), an HMPS enzyme. Other indications of differentiation were reduced cell proliferation, increased adherence and altered nuclear morphology. The observed increase in formazan production and HMPS activity and the reduction of cell proliferation due to RA were augmented by co-treatment with either glyceryl trinitrate, molsidomine, CAS 936 or tetranitromethane plus cysteine. Glyceryl trinitrate alone increased HMPS activity and G6PD mRNA levels and also reduced cell proliferation. Glyceryl trinitrate, molsidomine and CAS 936 are presumed to release nitric oxide and increase intracellular cGMP levels by stimulation of soluble guanylate cyclase. The mechanism of action of tetranitromethane is less certain, although it may also generate reactive nitrogen intermediates. These data suggest that a NO./cGMP pathway may augment a retinoic acid-mediated pathway to enhance maturation of U-937 cells with respect to the respiratory burst. Glyceryl trinitrate may act additionally by another pathway.

Cell Differentiation↗

PubMed1995

Secretion of MPB64 antigen by a recombinant clone of Mycobacterium smegmatis: characterization and application for the diagnosis of tuberculosis.

MPB64, a specific antigen to Mycobacterium tuberculosis complex (TB complex), was produced and secreted by a clone of M. smegmatis-MPB64 where the structural gene of MPB64 was inserted using a new mycobacteria, E. coli shuttle plasmid pNIS vector. Antibodies against the recombinant MPB64 (rMPB64) were used for the reverse particle latex agglutination (RPLA) test to detect the MPB64 antigen rapidly. RPLA tests were applied to the shock cultures and the clinical isolates of mycobacteria to identify TB complex. RPLA with anti-MPB64 antibody-coated latex beads completely distinguished TB complex from other mycobacteria. Thus, it is suggested that RPLA with anti-MPB64 antibody would be a new, easy and inexpensive method for the rapid diagnosis of tuberculosis.

Antigens, Bacterial↗

PubMed1994

Phosphorylation of an inhibitory subunit of cGMP phosphodiesterase in Rana catesbeiana rod photoreceptors. II. A possible mechanism for the turnoff of cGMP phosphodiesterase without GTP hydrolysis.

In amphibian rod photoreceptor membranes, P gamma, an inhibitory subunit of cGMP phosphodiesterase, is phosphorylated by a protein kinase when P gamma is complexed with the guanosine 5'-O-(3-thiotriphosphate)-bound alpha subunit of transducin (GTP gamma S.T alpha). Five different experiments support the conclusion that the phosphorylated P gamma loses its interaction with GTP gamma S.T alpha. These observations include 1) detection of the inhibitory effect of the GTP gamma S.T alpha.P gamma complex on cGMP phosphodiesterase activity after P gamma in the complex is phosphorylated in a system reconstituted from isolated components, 2) no stimulating effect of GTP gamma S.T alpha on the phosphorylated P gamma-inhibited cGMP phosphodiesterase in the reconstituted system, 3) physical release of phosphorylated P gamma from GTP gamma S.T alpha in the reconstituted system, 4) no inhibitory effect of phosphorylated P gamma on both GTP hydrolysis by T alpha and GTP gamma S/GDP exchange on T alpha in the reconstituted system, and 5) no enhanced activity of cGMP phosphodiesterase by GTP gamma S.T alpha in rod outer segment membranes after incubation of the membranes with the kinase preparation in the presence of ATP. Together with our data that P gamma released with GTP.T alpha is not phosphorylated, and that phosphorylated P gamma inhibits more effectively cGMP phosphodiesterase activity than nonphosphorylated P gamma (Tsuboi, S., Matsumoto, H., Jackson, K. W., Tsujimoto, K., Williams, T., and Yamazaki, A. (1994) J. Biol. Chem. 269, 15016-15023), these observations suggest that, after P gamma is released with GTP.T alpha from catalytic subunits of cGMP phosphodiesterase, P gamma complexed with GTP.T alpha is phosphorylated by a kinase. Then, the phosphorylated P gamma is released from GTP.T alpha and binds to active cGMP phosphodiesterase to inhibit the cGMP hydrolysis. It is suggested that in some G-protein-dependent signal transduction systems G-protein-activated effector may be phosphorylated with a specific kinase and that phosphorylation of the effector results in the turnoff of signal transduction without GTP hydrolysis.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed1994

Phosphorylation of an inhibitory subunit of cGMP phosphodiesterase in Rana catesbeiana rod photoreceptors. I. Characterization of the phosphorylation.

Interaction between the inhibitory subunit (P gamma) and catalytic subunits of cGMP phosphodiesterase is essential for the regulation of cGMP phosphodiesterase in vertebrate rod photoreceptors. P gamma phosphorylation in vitro has been studied using a kinase which is extracted from amphibian rod outer segments. Various chromatographies of the kinase preparation using ionic exchange, gel filtration, and heparin-Sepharose columns indicate that a kinase with M(r) 70,000 is responsible for the P gamma phosphorylation. The kinase does not require any of the known activators for protein kinases but is inhibited by cGMP in a concentration-dependent manner. Together with analysis by laser-desorption mass spectrometry, measurement of 32P radioactivity in phosphorylated P gamma indicates that P gamma extracted with GTP-bound transducin alpha subunit is not phosphorylated and that a phosphate is incorporated into more than 80% of the P gamma by the kinase. Phosphoamino acid analysis, sequencing of phosphorylated peptides derived from phosphorylated P gamma, and phosphorylation of synthetic peptides indicate threonine 22 in P gamma is phosphorylated by the kinase. Phosphorylated P gamma has a higher inhibitory activity for active cGMP phosphodiesterase than non-phosphorylated P gamma. These data suggest that threonine 22 in P gamma is phosphorylated by a specific kinase and that the P gamma phosphorylation governs the interaction between P gamma and catalytic subunits of cGMP phosphodiesterase in vertebrate rod photoreceptors.

3',5'-Cyclic-GMP Phosphodiesterases↗

PubMed1994

Immunoselection of GRP94/endoplasmin from a KNRK cell-specific lambda gt11 library using antibodies directed against a putative heparanase amino-terminal peptide.

Induction of an invasive phenotype by metastatic tumour cells results in part from inappropriate expression of extracellular matrix-degrading enzymes normally involved in embryonic morphogenesis, tissue remodelling, angiogenesis and wound healing. Such enzymes include endoglycosidases that degrade heparan sulfate (HS) in endothelial basement membrane, as well as better characterized proteases. Heparanase, an endo-beta-D-glucuronidase initially detected in B16 melanoma cells, has been described as a M(r) 96,000 glycoprotein with pI of 5.2, and has been immunolocalized to the cell surface and cytoplasm. We have utilized a polyacrylamide-gel-based HS degradation assay to demonstrate that KNRK, a rat kidney fibroblast cell line transformed by v-K-ras, exhibits HS-degrading activity similar to that of B16F10 mouse melanoma cells. To immunoselect heparanase-expressing clones from a KNRK-cell-specific lambda gt11 cDNA library, we have also prepared a rabbit anti-serum directed against a putative amino-terminal peptide of B16F10 cellular heparanase. Lysogens from one clone expressed a beta-galactosidase fusion protein whose staining with peptide anti-serum was inhibited by competition with excess peptide. Dideoxy-mediated sequencing of the insert termini of this recombinant revealed that it represents a rat homologue of M(r) 94,000 glucose-regulated protein (GRP94/endoplasmin), a molecular chaperone that contains the exact amino-terminal sequence previously attributed to heparanase. Our results call into question the specificity of this peptide sequence, as well as previous immunolocalization studies of heparanase carried out using such anti-sera.

Amino Acid Sequence↗

PubMed1994

Ultrastructural localization of retinal guanylate cyclase in human and monkey retinas.

Immuno-imaging with confocal and electron microscopy revealed the localization of retinal guanylate cyclase (RetGC) in human and monkey retinas. Using an antibody against a peptide derived from human RetGC, RetGC was found predominantly in the photoreceptor layer in these retinas, although a small amount of RetGC was detected in various other retinal cells. In particular, the cone outer segments were more densely labeled with the antibody than the rod outer segments. The RetGC in outer segments was localized exclusively in the membrane-rich domains, and appeared to be associated with the marginal region of the disk membrane and/or the plasma membrane. The connecting cilium and its cytoplasmic extension never showed immunoreactivity with the antibody. The localization of RetGC in photoreceptor cells is discussed from the viewpoint of mechanisms for the recovery of photoreceptors to the dark level.

Animals↗

PubMed1994

An in situ hybridization study on interleukin-1 beta mRNA induced by transient forebrain ischemia in the rat brain.

Expression of interleukin-1 beta (IL-1 beta) mRNA in the rat brain after transient forebrain ischemia was investigated by in situ hybridization histochemistry. Thirty min after the start of recirculation, IL-1 beta mRNA was induced in the several brain regions, including the olfactory bulb, cerebral cortex, hippocampus, striatum and thalamus where neuronal degeneration was reported to be observed after transient forebrain ischemia. The hybridization signals were observed both on the glial cells and around the vascular walls.

Animals↗

PubMed1994

Cytotoxic T lymphocyte response in mice induced by a recombinant BCG vaccination which produces an extracellular alpha antigen that fused with the human immunodeficiency virus type 1 envelope immunodominant domain in the V3 loop.

The host immune response of cell-mediated immunity, particularly that of cytotoxic T lymphocytes (CTLs), is a major immune defence mechanism which may provide resistance to a human immunodeficiency virus type 1 (HIV-1) spread leading to acquired immune deficiency syndrome (AIDS). To prevent the accompanying activity of HIV-1 proteins responsible for the loss of helper T-lymphocyte function, it is crucial to develop a live attenuated recombinant vaccine expressing only T- or both T- and B-cell epitopes. Here, we examined the expression of the HIV-1 Env protein V3 region (15 amino acids from Arg315 to Lys329) in Mycobacterium bovis BCG as a fused form with an extracellular alpha antigen of Mycobacterium kansasii. Balb/c mice inoculated with this recombinant BCG (rBCG), rapidly induced V3 peptide-specific CTLs. Target cell lysis was restricted to the murine class I major histocompatibility complex, H-2d. A similar CTL response was also elicited after Balb/c mice were immunized with the same rBCG even when pre-inoculated with non-recombinant BCG. Thus, the rapid induction of HIV-1-specific CTLs indicates that this vaccine may be a therapeutic approach to preventing progression to AIDS.

AIDS Vaccines↗

PubMed1994

Postprandial increase in the duodenal pH and the effect of intravenous secretin injection on ionic compositions of duodenal fluid and plasma in sheep.

Testing five castrated male sheep with a pH-electrode inserted into the duodenal digesta through a T-shaped cannula, we found that the pH of the duodenal digesta was significantly increased from 3.11 +/- 0.11 to 3.47 +/- 0.09 after a meal. Secretin (2.5 CHR U/kg) was intravenously injected to mimic the duodenal pH increase. The administration significantly increased the duodenal digesta pH from 3.32 +/- 0.18 to 4.85 +/- 0.61, which was accompanied by an increase in sodium concentration, but by a decrease in potassium and chloride concentrations. It also significantly decreased the arterial blood pH from 7.543 +/- 0.002 to 7.515 +/- 0.008, which was accompanied by a reduced plasma HCO3- concentration. From these results, we conclude that in sheep, feeding increases the duodenal digesta pH, and the postprandial pH increase in the duodenal fluid would be due to the raised HCO3- secretion from the gastrointestinal tract.

Acid-Base Equilibrium↗

PubMed1994

An extra phosphorylation of Na+,K(+)-ATPase by paranitrophenylphosphate (pNPP): evidence for the oligomeric nature of the enzyme.

Paranitrophenylphosphate (pNPP) induced fluorescence changes in fluorescence isothiocyanate (FITC)-labeled Na+,K(+)-ATPase preparations. The extents of changes were similar to those induced by acetylphosphate (AcP) accompanying accumulation of a K(+)-sensitive phosphoenzyme (E2P) and an ouabain bound phosphoenzyme in the presence of Mg2+ and 16 mM Na+. Phosphoenzymes formed from [32P]pNPP were shown to turn over. The ratio of the maximum amount of the phosphoenzyme formed from pNPP to that of the phosphoenzyme formed from ATP and that of the ouabain-enzyme complex under steady-state conditions was shown to be close to 1:0.5:1. Such extra phosphorylation has hitherto only been observed in a transient state with the additions of high concentrations of ATP [Peluffo, R.D., Garrahan, P.J., and Rega, A. (1992) J. Biol. Chem. 267, 6596-6601]. Our data are compatible with the simultaneous presence of high and low affinity ATP-binding sites in Na+,K(+)-ATPase [Hamer, E. and Schoner, W. (1993) Eur. J. Biochem. 213, 743-748]. The maximum amount of paranitrophenol-sensitive fraction to synthesize [32P]pNPP in fully accumulated ADP-sensitive phosphoenzyme (E1P) from [32P]ATP was around 1/4 of the amount of ouabain-enzyme complex. These data and others indicate that a much higher degree of oligomerization, rather than (alpha beta)2, may be the functional unit of the enzyme in the membranes.

Adenosine Triphosphate↗

PubMed1994

FR901451, a novel inhibitor of human leukocyte elastase from Flexibacter sp. II. Pharmacological effect of FR901451.

Intratracheal (i.t.) or intravenous (i.v.) administration of FR901451, a potent inhibitor of human leukocyte elastase (HLE) prevented HLE-induced lung hemorrhage in hamsters with ED50 values of 10.5 micrograms/site and 8.1 mg/kg, respectively. alpha 1-Antitrypsin (alpha 1-AT) also showed inhibitory effect in this model. However, the ED50 value by i.t. injection of FR901451 was 20-fold lower than that of alpha 1-AT. Moreover, FR901451 i.t. significantly modulated porcine pancreas elastase (PPE)-induced changes of the respiratory mechanics in hamsters. The ED50 values were 529 micrograms/site and 244 micrograms/site, which were expressed by static lung compliance (Cst) and vital capacity (VC) of the lungs, respectively. These results suggest that FR901451 could be clinically useful agent for the treatment of the destructive lung disease such as pulmonary emphysema.

Animals↗

PubMed1994

Application of hydroxyapatite-sol as drug carrier.

The application of hydroxyapatite-sol as a drug carrier is being developed. Hydroxyapatite-sol which is a suspension consisting of hydroxyapatite nano-crystals, was synthesized using an ultrasonic homogenizer. The size of the crystals was 40 x 15 x 10 mm3 on average and their specific surface area was 100 m2/g. An amount of a glycoside antibiotics adsorbed onto hydroxyapatite nano-crystals was measured. The drug adsorbed 0.2 mg per 1 mg of hydroxyapatite. The affect of the drug adsorbed onto the hydroxyapatite was investigated using cancer cells. The drug, adsorbed onto the hydroxyapatite nano-crystals, inhibited cancer cell growth.

Adsorption↗

PubMed1994

Breeding of the gad-mdx mouse: influence of genetically induced denervation on dystrophic muscle fibers.

A new double mutant mouse strain, gad-mdx, was established. The transmission of mdx and gad genes was monitored by determining their chemical markers, creative kinase activity and phosphoglucomutase-1 isoenzyme, respectively, in blood samples. This new strain was characterized by high creatine kinase activity in the plasma, lack of dystrophin in the muscle, and the presence of axonal swellings in the neural tissue. Although the body weight and the limb muscle size of the mutant mice were significantly lower than those of either gad or mdx mice, the clinical signs were not evident until the animals were 80 days old. After that time, the disease followed the course seen in the gad strain, and muscle weakness was exhibited in the advanced stages. Histologic examination revealed that the prevalence of muscle fiber necrosis, a deleterious consequence of the mdx gene, was significantly lower in the double mutant strain than in the mdx strain. These results supported the idea that small-caliber muscle fibers, which are induced by gad gene expression, are resistant to dystrophic necrosis. We believe that this double mutant strain will be valuable for the analysis of neural influence on diseased muscle fibers, and that it will also provide an opportunity for the testing of new therapeutic strategies for human Duchenne muscular dystrophy.

Animals↗