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Biomedical subjects

T Murata

Publications and source records attributed to T Murata.

At least 145 records · Page 8Linked to original sources
PubMed2000

OTX2 directly interacts with LIM1 and HNF-3beta.

Otx2 is a paired-class homeobox gene, and its functions in anterior visceral endoderm and/or anterior mesendoderm have been suggested to be vital for head development in mammals. Several transcription factors are expressed in these tissues, and mutant mice analyses have suggested the interactions of the Otx2 gene cascade with the Lim1 or HNF-3beta cascade. Here we show that OTX2 directly associates with LIM1 and HNF-3beta; OTX2 binds to the LIM1 homeodomain (HD) with its C-terminal region, whereas both HD and C-terminal regions of OTX2 bind to the HNF-3beta fork head domain or OTX2 HD. The luciferase assay with the P3C sequence, a specific DNA binding sequence for paired-class homeobox genes, has demonstrated that LIM1 enhances, but HNF-3beta represses, OTX2-directed gene expression. Direct interactions of OTX2 with LIM1 or HNF-3beta may play important roles in anterior visceral endoderm and/or anterior mesendoderm to constitute transcriptional regulatory networks for head development.

Animals↗
PubMed2000

Posthypoxic reoxygenation-induced neurotoxicity prevented by free radical scavenger and NMDA/non-NMDA antagonist in tandem as revealed by dynamic changes in glucose metabolism with positron autoradiography.

Using a positron autoradiography technique, dynamic changes in the cerebral glucose metabolic rate (CMRglc) induced by hypoxia/reoxygenation were investigated in living brain slices. After incubating fresh rat brain slices (300 microm thick) with [(18)F]2-fluoro-2-deoxy-D-glucose ([(18)F]FDG) in oxygenated Krebs-Ringer solution at 36 degrees C, serial two-dimensional time-resolved images of [(18)F]FDG uptake in the slices were obtained on imaging plates. As compared to the unloaded control values, with hypoxia-loading [(18)F]FDG uptake increased markedly, suggesting enhanced glycolysis. The net influx constant (K) of [(18)F]FDG at pre-hypoxia-loading and after reoxygenation with loading hypoxia for various periods of time was quantitatively evaluated by applying the Patlak graphical method to the image data. Regardless of the brain region, with hypoxia of </=10-min duration, the K value returned to the preloading level, whereas with hypoxia of >/=20 min duration only partial or no recovery was seen, indicating irreversible neuronal damage. The 30-min administration of either N-methyl-D-aspartate (NMDA)/non-NMDA antagonist or a free radical scavenger at the same time as reoxygenation after 20 min hypoxia showed a neuroprotective effect inhibiting the decrease in the post-hypoxia-loading K value. In contrast, no such neuroprotective effect was evident with administration of either of these agents only during hypoxia loading, possibly indicating that immediately after reoxygenation neuronal damage was induced mediated by excitatory amino acids and free radicals in tandem. These results demonstrate that serial quantitative evaluation of CMRglc using this technique may be of use in investigating the brain tissue injury associated with hypoxia/reoxygenation as well as clarifying the underlying mechanisms and protective effect of various drugs against such injury.

6-Cyano-7-nitroquinoxaline-2,3-dione↗
PubMed2000

Cementless cup supporter for protrusio acetabuli in patients with rheumatoid arthritis.

The MC cup supporter is an acetabular reinforcement ring for the Harris-Galante (HG) cementless porous-coated socket. This supporting ring has been used with autogenous bone graft since 1991 in the management of protrusio acetabuli in patients with rheumatoid arthritis (RA). We have reviewed 15 hips in 13 patients at an average of 53.6 months (range 24-84 months) after operation. The clinical results of total hip arthroplasty (THA) in protrusio acetabuli using this ring were satisfactory. All grafts appeared to have united radiologically with no radiolucent lines between graft and acetabulum.

Acetabulum↗
PubMed2000

Glycine substitution mutations by different amino acids in the same codon of COL7A1 lead to heterogeneous clinical phenotypes of dominant dystrophic epidermolysis bullosa.

Dystrophic epidermolysis bullosa (DEB), caused by mutations in the gene encoding type VII collagen (COL7A1), is known to show heterogeneous clinical phenotypes. Certain correlations between the nature or position of COL7A1 mutations and the resultant DEB phenotypes have been suggested, although such relationships may be more complex than initially thought. The purpose of the present study was to clarify the molecular basis of two different subtypes of dominant DEB (DDEB), EB pruriginosa and classical type. Interestingly, we found that both cases were caused by a missense glycine substitution mutation by different amino acids in the same codon of COL7A1 (G2028R and G2028A). These results further support the notion that different glycine substitution mutations in the same codon can lead to heterogeneous clinical phenotypes of DDEB, EB pruriginosa and classical type.

Adult↗
PubMed2000

CD18 and ICAM-1-dependent corneal neovascularization and inflammation after limbal injury.

Extensive limbal injury is a leading cause of irreversible blindness. The destruction of corneal limbal stem cells often results in corneal neovascularization and an optically inferior epithelium. Previous work has shown that the neovascularization after limbal injury is vascular endothelial growth factor (VEGF)-dependent, with much of the VEGF emanating from the inflammatory cells that invade the cornea. Using a relevant mouse model of limbal injury, we examined the role of CD18 and intercellular adhesion molecule-1 (ICAM-1) in limbal injury-induced neovascularization. The results show that CD18- and ICAM-1-deficient mice developed 35% (n = 5, P = 0.003) and 36% (n = 5, P = 0.002) less neovascularization than strain-specific normal controls, respectively. The corneal neutrophil counts were similarly reduced by 51% (n = 5, P < 0.003) and 46% (n = 5, P < 0.006), respectively. When VEGF mRNA levels were analyzed, they were reduced by 66% (n = 3, P = 0.004) and 48% (n = 3, P = 0.024), respectively. Taken together, these data identify CD-18 and ICAM-1 as mediators of the inflammatory and VEGF-dependent corneal neovascularization that follows limbal injury. The targeting of CD18 and ICAM-1 may prove useful in the treatment of inflammation-associated neovascularization in the cornea and elsewhere.

Animals↗
PubMed2000

Expression of phosphodiesterase 3 in rat submandibular gland.

The expression of phosphodiesterase (PDE) 3 isoforms was investigated in extracts of rat submandibular gland by reverse transcription-polymerase chain reaction (RT-PCR) and the PCR fragments were then sequenced. PDE3 activity was detected in gland homogenates; about 90% of the activity was in the supernatant fraction and about 10% in the particulate fraction. PDE3A and 3B mRNA was detected by RT-PCR in RNA from the gland. The nucleotide sequences of the fragments were identical to those of rat PDE3A and 3B. The results indicate that two PDE3 isoforms are present in rat submandibular gland and may regulate an important cAMP pool.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed2000

The possibility of gene therapy for the treatment of choroidal neovascularization.

PURPOSE: Choroidal neovascularization (CNV) is responsible for most cases of severe visual loss in age-related macular degeneration. Recently, the possibility of gene therapy has been proposed for the treatment of CNV. The purpose of this study was to examine the feasibility of ex vivo and in situ gene therapy approaches for CNV. DESIGN: Experimental study. METHODS: Human retinal pigment epithelial (RPE) cells were transduced with a retroviral vector coding for beta-galactosidase. Transduced cells were grown on type II collagen sheets and transplanted under the retina of 20 rabbits. Animals were observed for 3 to 56 days, and transplanted cells were examined histologically and with X-gal staining. Bovine choroidal endothelial cells (CEC) were transduced with retroviral vectors coding for tissue inhibitor of metalloproteinase-2 (TIMP-2) or control vector. Production of TIMP-2 by transduced cells was determined by immunohistochemical analysis and enzyme-linked immunosorbent assay. Effect of transduction on in vitro proliferation, migration, and tube formation was examined in response to vascular endothelial growth factor (VEGF). Four CNV lesions were induced in one cynomolgus monkey by laser photocoagulation. Two days later, retroviral vector coding for TIMP-2 or control vector was injected into the subretinal space overlying the CNV lesions. The monkey was observed for 12 weeks using fluorescein angiography. RESULTS: Transplantation of transduced RPE cells was technically achieved in 10 of 20 animals. In these animals, RPE cells at the site of transplantation formed a monolayer and expressed beta-galactosidase for 14 days. beta-Galactosidase-positive cells were not identified at 56 days. Choroidal endothelial cells transduced with TIMP-2 secrete TIMP-2 into the media and show decreased migration and tube formation in vitro. In the in vivo monkey model, the control CNV lesions (n = 2) showed prominent leakage, whereas the experimental lesions (n = 2) showed minimal hyperfluorescence. CONCLUSIONS: Retrovirally transduced RPE cells survive in the subretinal space for at least 14 days and continue to express the gene product coded for by the vector. Choroidal endothelial cells retrovirally transduced for TIMP-2 produce TIMP-2 in vitro and show decreased angiogenic responses in vitro in response to VEGF. A preliminary study attempting in situ delivery of TIMP-2 vector to CNV lesions in a monkey eye supports the feasibility of this approach and encourages further study.

Animals↗
PubMed2000

Comparative analysis of the whole set of rRNA operons between an enterohemorrhagic Escherichia coli O157:H7 Sakai strain and an Escherichia coli K-12 strain MG1655.

Two primer sets for direct sequence determination of all seven rRNA operons (rrn) of Escherichia coli have been developed; one is for specific-amplification of each rrn operon and the other is for direct sequencing of the amplified operons. Using these primer sets, we determined the nucleotide sequences of seven rrn operons, including promoter and terminator regions, of an enterohemorrhagic E. coli (EHEC) O157:H7 Sakai strain. To elucidate the intercistronic or intraspecific variation of rrn operons, their sequences were compared with those for the K-12 rrn operons. The rrn genes and the internal transcribed spacer regions showed a higher similarity to each other in each strain than between the corresponding operons of the two strains. However, the degree of intercistronic homogeneity was much higher in the EHEC strain than in K-12. In contrast, promoter and terminator regions in each operons were conserved between the corresponding operons of the two strains, which exceeded intercistronic similarity.

Base Sequence↗
PubMed2000

A new murine zinc finger gene, Opr.

Here we report a novel murine zinc-finger gene, Opr, belonging to the opa/Zic family. Opr is expressed in the entire embryonic ectoderm before gastrulation, but gradually restricted to the anterior part from the mid to late streak stage. At the beginning of neural induction, Opr is expressed throughout the anterior neural plate, but is soon restricted to the neural ridge. After neural tube closure, its expression is maintained in the dorsal part of the neural tube, except for the roof of the telencephalon. Opr is also expressed in somites and limbs.

Amino Acid Sequence↗
PubMed2000

Biotransformation of beta-myrcene by the larvae of common cutworm (Spodoptera litura).

beta-Myrcene was mixed in an artificial diet at a concentration of 1 mg/g of diet, and the diet was fed to the last instar larvae of common cutworm (Spodoptera litura). Metabolites were recovered from frass and analyzed spectroscopically. beta-Myrcene was transformed mainly to myrcene-3(10)-glycol and myrcene-1,2-glycol. Each pair of double bonds of beta-myrcene was converted to the corresponding diol by oxidation, respectively. The 3,10- and 1,2-double bonds of beta-myrcene were respectively oxidized.

Acyclic Monoterpenes↗
PubMed2000

Characterization of the 5'-flanking region of the rat TIS11 gene.

To understand the mechanism underlying the transcriptional regulation of the immediate early gene TIS11, we characterized the 5'-flanking region of the rat TIS11 gene. When fused to the luciferase reporter gene, the 5.3-kb 5'-flanking region of the rat TIS11 gene exhibited functional promoter activity in pheochromocytoma PC12 and hepatoma H4IIE cells. 5'-Deletion analyses indicated that multiple negative and positive regulatory regions were present in the 5'-flanking region, and that some of these regions functioned in a cell type-specific manner. Promoter activity of the rat TIS 11 gene was enhanced by phorbol 12-myristate 13-acetate (PMA) in both cell lines, and the PMA-responsiveness resided within the 5'-flanking region. The induction of promoter activity by PMA was completely blocked by GF109203X or PD98059, inhibitors of protein kinase C and mitogen-activated protein (MAP) kinase kinase, respectively. These results suggested that induction of the rat TIS 11 promoter by PMA is mediated by activation of the protein kinase C/MAP kinase cascade.

Animals↗
PubMed2000

Cloning and characterization of the rat TIS11 gene.

The TIS11 gene is an immediate early gene that is induced rapidly and transiently by phorbol 12-myristate 13-acetate and various growth factors. To study transcriptional regulation of the gene, a genomic clone of rat TIS11 was isolated, and the organization of exon-intron structure and transcriptional initiation site were determined. The rat TIS11 gene consisted of 2 exons spanning approximately 2.5 kb. Several canonical sequences for binding of transcriptional factors were found in the 5'-flanking region. The 5.3 kb of the 5'-flanking region fused to a luciferase reporter gene showed promoter activity when introduced into rat pheochromocytoma PC12 cells. Analyses with serial 5'-deletion mutants suggested that the major positive regulatory region is located at the region of -241 to -76, and that the minimum promoter region is within the 76-bp upstream of the transcriptional initiation site. Gel mobility shift assays revealed that PC12 cell nuclear proteins specifically bind to the major positive regulatory region of the TIS11 gene. The identified nuclear protein components may act as the positive trans-acting factors in the basal expression of the TIS11 gene in PC12 cells.

Amino Acid Sequence↗
PubMed2000

Expression of herpes simplex virus type 2 US3 affects the Cdc42/Rac pathway and attenuates c-Jun N-terminal kinase activation.

BACKGROUND: Although the US3 gene product of herpes simplex virus (HSV) has been identified as a serine/threonine protein kinase (PK), the functions are poorly understood. RESULTS: We found that US3 PK of HSV-2 induced disruption of actin filaments, cell rounding and accumulation of binucleate cells in HEp2 cells. Cell rounding was abrogated by expression of either kinase-dead forms of US3 PK or a mutant protein lacking the acidic cluster in the kinase regulatory domain. Co-expression of dominant active forms of Cdc42/Rac inhibited cell rounding, suggesting that a signal transduction pathway involving Cdc42/Rac may play a role in the morphological changes induced by the kinase. Cdc42 and Rac, members of the Rho family of small GTPases, function as molecular switches changing actin cytoskeletal organization, influencing transcription and controlling apoptotic cell death. By computed homology search, we noticed significant similarities between US3 PK and p21-activated kinase (PAK), which is activated by the Cdc42 or Rac. We also found that the expression of US3 suppressed the activation of c-Jun N-terminal kinase (JNK), a kinase that is downstream of PAK. CONCLUSIONS: These observations suggest that the US3 PK affects the Cdc42/Rac pathway and can act as an upstream suppressor of JNK in the stress-activated signalling pathway.

Actins↗
PubMed2000

Effect of suckling on NADPH-diaphorase (Nitric oxide synthase, NOS) reactivity and NOS gene expression in the paraventricular and supraoptic nuclei of lactating rats.

This study examined the effect of suckling on nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d, a histochemical marker for nitric oxide synthase, NOS) reactivity and neuronal NOS mRNA expression in the paraventricular (PVN) and supraoptic (SON) nuclei of lactating rats. Freely nursing (non-separated) dams and those separated from pups for 12 h and then reunited for 0, 15, 30, 60, 90, 120 and 180 min were used for the study. Dams separated from pups and sacrificed at time zero (without reunion) showed a significant decrease in NADPH-d staining and NADPH-d positive cells as well as in the NOS mRNA expression in the PVN and SON compared to that observed in non-separated dams. Reunion with pups and restoration of suckling significantly increased NADPH-d reactivity after 15, 30, 60 min, but not after 90, 120 and 180 min compared to non-reunited pups-deprived dams. A pattern of NADPH-d reactivity and neuronal NOS mRNA expression indistinguishable from that observed during free lactation was reinstated shortly (15 min) after the restoration of suckling stimulus, suggesting that the NADPH-d reactivity in lactation depends on the presence of the suckling stimulus. These results show that suckling stimulus may play a modulatory role in the regulation of NOS reactivity in the magnocellular neurones of the hypothalamic PVN and SON during lactation.

Animals↗
PubMed2000

The R-type pyocin of Pseudomonas aeruginosa is related to P2 phage, and the F-type is related to lambda phage.

Pseudomonas aeruginosa produces three types of bacteriocins: R-, F- and S-type pyocins. The S-type pyocin is a colicin-like protein, whereas the R-type pyocin resembles a contractile but non-flexible tail structure of bacteriophage, and the F-type a flexible but non-contractile one. As genetically related phages exist for each type, these pyocins have been thought to be variations of defective phage. In the present study, the nucleotide sequence of R2 pyocin genes, along with those for F2 pyocin, which are located downstream of the R2 gene cluster on the chromosome of P. aeruginosa PAO1, was analysed in order to elucidate the relationship between the pyocins and bacteriophages. The results clearly demonstrated that the R-type pyocin is derived from a common ancestral origin with P2 phage and the F-type from lambda phage. This notion was supported by identification of a lysis gene cassette similar to those for bacteriophages. The gene organization of the R2 and F2 pyocin gene cluster, however, suggested that both pyocins are not simple defective phages, but are phage tails that have been evolutionarily specialized as bacteriocins. A systematic polymerase chain reaction (PCR) analysis of P. aeruginosa strains that produce various subtypes of R and F pyocins revealed that the genes for every subtype are located between trpE and trpG in the same or very similar gene organization as for R2 and F2 pyocins, but with alterations in genes that determine the receptor specificity.

Amino Acid Sequence↗
PubMed2000

Granzyme B-positive primary gastric T-cell lymphoma: gastric T-cell lymphoma with the possibility of extrathymic T cell origin.

A case of primary gastric T-cell lymphoma, which was positive for granzyme B, is reported. The patient was a 47-year-old Japanese female who complained of a dull upper abdominal pain. Radiographic and endoscopic examinations revealed an ulcerative infiltrative lesion in her stomach. Following the confirmation of a high-grade malignant lymphoma, a distal gastrectomy with regional lymph nodal dissection was performed. The histology of the gastric lesion revealed a malignant lymphoma of the diffuse pleomorphic type without lymph nodal involvement. Immunohistochemistry revealed that the tumor cells were positive for LCA, CD3, TIA-1 and granzyme B, but were negative for CD4, CD8, CD56, CD30, L-26, EMA, TCR alpha/beta and TCR gamma/delta. Because the tumor cells showed T cell nature with cytotoxic activity proved by TIA-1 and granzyme B, and without evidence of further maturation of T cell, a malignant lymphoma originating from extrathymic-derived T cells was suggested.

Biomarkers, Tumor↗