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T Nishihara

Publications and source records attributed to T Nishihara.

At least 145 records · Page 8Linked to original sources
PubMed1995

Vitamin D receptor contains multiple dimerization interfaces that are functionally different.

The vitamin D receptor mediates the signal of 1 alpha, 25-dihydroxyvitamin D3 by binding to vitamin D responsive elements in DNA as a homodimer or as a heterodimer composed of one vitamin D receptor subunit and one retinoid X receptor subunit. We have mapped the dimerization interfaces of the vitamin D receptor that is involved in homo- or heterodimer formation in the absence of DNA. While deletion of the first zinc finger region of vitamin D receptor diminished homodimerization activity, it did not affect heterodimerization. In contrast, a deletion just beyond the zinc finger region affected heterodimerization with retinoid X receptor, but not homodimerization. The zinc finger region alone could form a homodimer with full-length vitamin D receptor, but not a heterodimer with retinoid X receptor. The carboxy-terminal region was also necessary for heterodimer formation. This region showed only a weak dimerization activity in the absence of ligand, but this was dramatically increased in the presence of ligand for both homo- and heterodimerization. These results suggest that the vitamin D receptor has at least three dimerization interfaces whose functions are apparently distinguishable. These are located in the first zinc finger region, the region just beyond this zinc finger and in the carboxy-terminal region.

Amino Acid Sequence↗
PubMed1995

Sequence analysis of the rat jun-D gene.

A rat jun-D genomic clone was isolated from a rat liver lambda EMBL3 library. Sequence analysis revealed a high sequence similarity with mouse jun-D, and relatively low similarity with human and chicken jun-D. In both the trans-activation and DNA-binding domains, however, the amino-acid sequences were well conserved among rat, mouse, human and chicken.

Amino Acid Sequence↗
PubMed1995

Induction of apoptosis in B lineage cells by activin A derived from macrophages.

A factor produced by P388D1 cell line murine macrophages showed a profound suppressive effect on the in vitro proliferation of B lineage cells. It was purified to homogeneity from conditioned media of P388D1 cells stimulated with phorbol 12-myristate 13-acetate for 48 h by a three-step procedure. The purified factor gave a single band of protein with a molecular mass of 16 kD on SDS-polyacrylamide gel electrophoresis. We show here that exposure of B lineage cells to this factor results in the induction of a cytotoxic effect and a significant increase in the proportion of fragmented DNA. DNA fragmentation was detected in B lineage cells after 3 h culture with the factor in the quantitative colorimetric determination. The mechanism of cell death was characterized by a ladder-like electrophoretic pattern of degraded chromosomal DNA, indicating that the factor induces apoptosis. The NH2-terminal amino acid sequence of this factor was identical with that of activin A over the 26 amino acid residues identified. We sought to determine whether apoptosis could be modulated by two kinds of inhibitor of protein kinases, H7 and HA1004, in concentrations that are below their toxicity limits. Apoptosis induced by the factor was suppressed by H7 but was relatively unaffected by HA1004. These findings suggest that the signals by protein kinases may regulate apoptotic B cell death by the factor activin A, derived from macrophages.

Activins↗
PubMed1995

Role of prostaglandin in the formation of osteoclasts induced by capsular-like polysaccharide antigen of Actinobacillus actinomycetemcomitans strain Y4.

We found no reports that capsular-like polysaccharide antigen purified from Actinobacillus actinomycetemcomitans either induces osteoclastic bone resorption in mouse organ cultures or promotes osteoclast formation in mouse marrow cultures. In contrast, capsular-like polysaccharide antigen purified from A. actinomycetemcomitans strain Y4 induced bone resorption in mouse organ culture. To examine the mechanism of bone resorption induced by A. actinomycetemcomitans, mouse bone marrow cells were cultured with A. actinomycetemcomitans strain Y4 capsular-like polysaccharide antigen. A. actinomycetemcomitans strain Y4 capsular-like polysaccharide antigen stimulated osteoclast-like cell formation in mouse bone marrow cultures. However, the polysaccharide of A. actinomycetemcomitans lipopolysaccharide did not induce the formation of osteoclast-like cells. Indomethacin inhibited osteoclast-like cell formation mediated by A. actinomycetemcomitans strain Y4 capsular-like polysaccharide antigen in a dose-dependent manner. There was a good correlation between the number of osteoclast-like cells formed in the marrow culture and the amount of prostaglandin E2 released into the culture media. When mouse bone marrow cells were cultured with prostaglandin E2 during the culture periods, many osteoclast-like cells were formed. These results indicate that prostaglandin E2 is involved in the mechanism of the formation of osteoclast-like cells mediated by A. actinomycetemcomitans strain Y4 capsular-like polysaccharide antigen. A. actinomycetemcomitans strain Y4 capsular-like polysaccharide antigen may play an important role in inflammatory bone resorption by promoting osteoclast formation in periodontal disease.

Acid Phosphatase↗
PubMed1995

Molecular and immunological characterization of a 64-kDa protein of Actinobacillus actinomycetemcomitans.

The 64-kDa protein to which about half the sera from patients with localized juvenile periodontitis and rapidly progressive periodontitis reacted strongly was purified from Actinobacillus actinomycetemcomitans Y4. Determination of the N-terminal sequence of the protein revealed that it was a GroEL-like protein. The DNA fragment containing the groEL gene of A. actinomycetemcomitans was amplified by polymerase chain reaction, and the groESL operon was cloned by using colony hybridization with the amplified fragment from A. actinomycetemcomitans chromosomal DNA. Sequence analysis revealed that structures of the operon and its products were typical in gram-negative bacteria. Rabbit polyclonal antibodies to the 64-kDa protein cross-reacted with approximately 65-kDa proteins of Haemophilus aphrophilus, Haemophilus influenzae, Haemophilus paraphrophilus, Escherichia coli and Eikenella corrodens but not with any cellular proteins of Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum. It is possible that antibodies reactive to the 64-kDa protein in periodontitis patients are induced by the cross-reactivity with the hsp60 proteins of other bacteria.

Aggregatibacter actinomycetemcomitans↗
PubMed1995

Evidence for apoptosis of murine macrophages by Actinobacillus actinomycetemcomitans infection.

The gram-negative bacterium Actinobacillus actinomycetemcomitans is considered an important etiological agent in periodontal diseases. In this study, we show that A. actinomycetemcomitans strains are cytotoxic for the murine macrophage cell line J774.1. On the other hand, Porphyromonas gingivalis strains, other gram-negative oral species implicated in adult periodontitis, showed weak cytotoxic effects. For this to occur, A. actinomycetemcomitans had to gain entry into the macrophages, since cytotoxicity was prevented by cytochalasin D. We demonstrate that cell death induced by A. actinomycetemcomitans Y4 occurs through apoptosis, as shown by changes in nuclear morphology, an increase in the proportion of fragmented DNA, and the typical ladder pattern of DNA fragmentation indicative of apoptosis. We further sought to determine whether the cytotoxicity induced by A. actinomycetemcomitans Y4 could be modulated by the protein kinase inhibitors H7 and HA1004. Apoptotic cell death induced by A. actinomycetemcomitans Y4 was suppressed by H7 but was relatively unaffected by HA1004. These findings suggest that the signals of protein kinases may regulate apoptosis induced by A. actinomycetemcomitans Y4. The ability of A. actinomycetemcomitans to promote the apoptosis of macrophages may be important for the initiation of infection and the development of periodontal diseases.

Actinobacillus Infections↗
PubMed1995

Actinobacillus actinomycetemcomitans Y4 capsular-polysaccharide-like polysaccharide promotes osteoclast-like cell formation by interleukin-1 alpha production in mouse marrow cultures.

The mechanism of osteoclast-like cell formation induced by periodontopathic bacterium Actinobacillus actinomycetemcomitans Y4 (serotype b) capsular-polysaccharide-like polysaccharide (capsular-like polysaccharide) was examined in a mouse bone marrow culture system. When mouse bone marrow cells were cultured with A. actinomycetemcomitans Y4 capsular-like polysaccharide for 9 days, many multinucleated cells were formed. The multinucleated cells showed several characteristics of osteoclasts, including tartrate-resistant acid phosphatase (TRACP) and the ability to resorb the calcified dentine. In this study, we examined the effects of antisera to interleukins on the formation of osteoclast-like cells induced by A. actinomycetemcomitans Y4 capsular-like polysaccharide. Monospecific anti-mouse recombinant interleukin-1 alpha (rIL-1 alpha) serum completely inhibited the formation of osteoclast-like cells in the presence of A. actinomycetemcomitans Y4 capsular-like polysaccharide. However, anti-mouse rIL-1 beta and anti-mouse rIL-6 sera showed no effect on osteoclast-like cell formation. IL-1 receptor antagonist significantly inhibited the osteoclast-like cell formation mediated by A. actinomycetemcomitans Y4 capsular-like polysaccharide in mouse marrow cultures. The bioactive IL-1 was detected in the culture media of mouse bone marrow cells stimulated with A. actinomycetemcomitans Y4 capsular-like polysaccharide. These results indicate that IL-1 alpha is involved in the mechanism of the formation of osteoclast-like cells induced by A. actinomycetemcomitans Y4 capsular-like polysaccharide. We sought to determine whether osteoclast-like cell formation induced by A. actinomycetemcomitans Y4 capsular-like polysaccharide could be modulated by the protein kinase inhibitors H8 and HA1004. The formation of osteoclast-like cells was suppressed by H8 and HA1004. These findings suggest that the signals by protein kinases may regulate osteoclast-like cell formation induced by A. actinomycetemcomitans Y4 capsular-like polysaccharide. Furthermore, a correlation between IL-1 alpha and prostaglandin E2 in the osteoclast recruitment induced by A. actinomycetemcomitans Y4 capsular-like polysaccharide is discussed.

Acid Phosphatase↗
PubMed1995

The potent bone-resorbing mediator of Actinobacillus actinomycetemcomitans is homologous to the molecular chaperone GroEL.

Actinobacillus actinomycetemcomitans is a Gram-negative bacterium implicated in the pathology of localized juvenile periodontitis, a condition involving rapid destruction of alveolar bone. We have established that gentle extraction of this bacterium in saline releases a proteinaceous fraction (which we have termed surface-associated material [SAM] which has potent osteolytic activity in the murine calvarial bone resorption assay. Fractionation of the SAM has now revealed that activity is associated with a 62-kD protein. This bone-resorbing activity can be blocked by a monoclonal antibody (raised to the whole bacterium) that is claimed to recognize a protein homologous to the Escherichia coli molecular chaperone GroEL. Purification of this bone-resorbing protein to homogeneity has been achieved by a combination of anion exchange, gel filtration, and ATP-affinity chromatography and the NH2-terminal sequence shows > 95% homology to E. coli GroEL. This GroEL homologue is found in the SAM of A. actinomycetemcomitans but is not found in the osteolytically active SAM from other Gram-negative or Gram-positive bacteria. The GroEL protein from E. coli, but not from Mycobacterium tuberculosis and Mycobacterium leprae, also showed activity in the bone resorption assay. We believe this to be the first observation that a molecular chaperone has the capacity to stimulate the breakdown of connective tissue.

Aggregatibacter actinomycetemcomitans↗
PubMed1995

Membrane-associated interleukin-1 on macrophages stimulated with Actinobacillus actinomycetemcomitans lipopolysaccharide induces osteoclastic bone resorption in vivo.

The effect of paraformaldehyde-fixed murine macrophage P388D1 cells stimulated with Actinobacillus actinomycetemcomitans Y4 lipopolysaccharide (membrane-associated interleukin-1) on osteoclastic bone resorption was investigated. Both the number of osteoclasts and the bone resorption surfaces increased considerably along the inside of the marrow space of mouse calvaria when membrane-associated interleukin-1 mediated by A. actinomycetemcomitans Y4 lipopolysaccharide were injected into the subcutaneous tissues overlying the central calvaria of 4-week-old C3H/HeJ mice. Membrane-associated interleukin-1 caused hypercalcaemia when injected into C3H/HeJ mice twice a day for 2 days. The calvaria of C3H-HeJ mice injected with membrane-associated interleukin-1 contained less mineral than those of the control mice. These results suggest that membrane-associated interleukin-1 on macrophages stimulated with A. actinomycetemcomitans Y4 lipopolysaccharide plays an important role in inflammatory osteoclastic bone resorption in periodontal diseases.

Aggregatibacter actinomycetemcomitans↗
PubMed1995

[Effects of separate administration of low-dose CDDP on antitumor immune reactivity for treatment of malignant glioma].

Recently, it has been reported that CDDP modifies the immune responses of tumor bearing hosts, but problem of the most appropriate treatment with CDDP to augment the host immunity remains unresolved. The purpose of this study was to determine the most advantageous administration of CDDP to increase both its cytotoxic effect and antitumor immune reactivity. Cell growth inhibition in vitro was assessed by MTT assay after treatment of U-251MG cells with various concentrations of CDDP for 24 or 120 hours. The results of this experiment showed that the growth of U-251MG cells in vitro was suppressed by CDDP in a dose-dependent as well as a time-dependent manner. Ten days after subcutaneous inoculation of human glioma cells, GL-9, in the rear flank of nude mice, they were assessed for growth suppression by CDDP. Totally, 20 ml/kg of CDDP was administered to each mouse; in one group (high-dose CDDP group), 5 ml/kg of CDDP on every fifth day, and in the other group (low-dose CDDP group), 2 ml/kg on every day. Measurement of the tumor volume in each group revealed no significant difference between the two groups in terms of the efficacy of tumor growth suppression. Twenty-one days after inoculation, we measured the splenic natural killer(NK) cell activity in each mouse. The results showed that NK cell activity was significantly increased in the low-dose cisplatin (2 ml/kg) group, and significantly decreased in the high-dose cisplatin (10 mg/ml) group, as compared to the control group. The results of our study suggest that the separate administration of low-dose CDDP is a useful treatment strategy for malignant glioma, because it increases the antitumor immune reactivity of hosts without decreasing the direct tumor cytotoxicity of CDDP.

Animals↗
PubMed1995

[Chylothorax in a patient with 9-year remission of malignant lymphoma].

We describe a 69-year-old man who developed chylothorax after a 9-year remission of malignant lymphoma. The patient was admitted to our hospital and received exploratory laparotomy for ileus in February 1986. Bulky masses in the posterior mediastinum and the retroperitoneum, and also a jejunal tumor were observed. Fibrosis of the liver was also observed. The jejunal tumor was removed and histological findings revealed diffuse large B-cell malignant lymphoma. He was treated by combination chemotherapy and remission was achieved. He was discharged in June and remained in remission, but was readmitted for right pleural effusion in October 1994. Effusion was chylous and the chylomicron level was estimated to be 181 mg/dl. Liver cirrhosis also developed but there was no chylous ascites. Chylorrhea disappeared after continuous aspiration, but recurred in December. Continuous aspiration was ineffective, therefore 10 KE of OK-432 was administered twice into the pleural cavity, and chylorrhea again disappeared. No findings suggestive of malignant lymphoma were not detected by computerized tomography and gallium scintigram. He was discharged in March 1995 and chylothorax has not recurred since. These findings suggest that the fragility of the thoracic duct which had been infiltrated by malignant lymphoma might increase, resulting in rupture, even if in remission.

Aged↗
PubMed1994

Difference and similarity of DNA sequence recognized by VDR homodimer and VDR/RXR heterodimer.

Nuclear receptors for the thyroid hormone and vitamin A and D cooperate with the retinoid X receptor (RXR) in activating the transcription. Although the hormone response elements for these receptors have been proposed in which spacing of the direct repeated motifs determine the specificity (so called 3-4-5 rule), vitamin D response elements (VDREs) in the natural context consist of often imperfect direct repeats. Vitamin D receptor (VDR) alone can bind to the mouse osteopontin (mSPP-1) VDRE, which contains a direct repeat separated by 3 nucleotides, but not to the rat osteocalcin (rOST) VDRE having inexact direct repeat. The presence of RXR not only allows the VDR to bind to the rOST VDRE, but also increases the binding affinity for the mSPP-1 VDRE. The RXR/VDR heterodimer exhibits the similar affinity constants for the mSPP-1 VDRE and the rOST VDRE, in spite of the apparently different affinities for two VDREs of the VDR homodimer. A random oligonucleotide selection procedure revealed that the consensus sequence selected by the RXR homodimer is the direct repeat spaced by one A residue. In contrast, the sequences preferentially selected by the VDR homodimer and the VDR/RXR heterodimer are similar, which are the direct repeats spaced by 3 nucleotides. The difference and similarity of DNA sequence recognition are discussed.

Animals↗
PubMed1994

Membrane-associated interleukin-1 promotes osteoclast-like cell formation in vitro.

The effect of paraformaldehyde-fixed murine macrophage P388D1 cells on osteoclast-like cell formation was investigated in mouse marrow cultures. When mouse marrow cells were cocultured for 8 days with paraformaldehyde-fixed P388D1 cells stimulated with lipopolysaccaride (LPS), many tartrate-resistant acid phosphatase (TRACP)-positive multinucleated cells were formed. Non-stimulated, and paraformaldehyde-fixed P388D1 cells did not induce the formation of TRACP-positive multinucleated cells. Salmon calcitonin and indomethacin strongly inhibited the paraformaldehyde-fixed, LPS-stimulated P388D1 cells-induced TRACP-positive multinucleated cell formation. Monospecific anti-mouse recombinant IL-1 alpha serum inhibited TRACP-positive multinucleated cell formation in the presence of paraformaldehyde-fixed P388D1 cells stimulated with LPS. These results suggest that membrane-associated IL-1 on macrophages is responsible for the resorptive effect of paraformaldehyde-fixed P388D1 cells stimulated with LPS.

3T3 Cells↗
PubMed1994

Characterization of phe B gene encoding catechol 2,3-dioxygenase.

DNA sequence of the phe B gene isolated from a chromosome of the phenol degrading bacterium Pseudomonas putida BH is identical to that of the dmp B gene from the phenol degradative plasmid, pVI150. Catechol 2,3-dioxygenase encoded by phe B showed similar substrate specificity to that of xyl E. However, Phe B has much smaller Km values than Xyl E, indicating that Phe B is useful for treatment of low concentrations of catechol derivatives in waste water.

Biodegradation, Environmental↗
PubMed1994

Mouse interleukin-1 receptor antagonist induced by Actinobacillus actinomycetemcomitans lipopolysaccharide blocks the effects of interleukin-1 on bone resorption and osteoclast-like cell formation.

We have reported that P388D1 cell line murine macrophages stimulated with lipopolysaccharide (LPS) from Actinobacillus actinomycetemcomitans release interleukin-1 (IL-1) inhibitor. The IL-1 inhibitor was purified from conditioned media of P388D1 cells stimulated with A. actinomycetemcomitans LPS for 72 h to homogeneity by a four-step procedure: acetic acid extraction from conditioned media; Bio-Gel P-60 gel filtration chromatography; DEAE-Sepharose CL-6B column chromatography; and reverse-phase high-performance liquid chromatography on a C18 hydrophobic support. The purified IL-1 inhibitor gave a single band of protein with a molecular mass of 26 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified IL-1 inhibitor was a heat- and acid-stable protein that was inactivated by digestion with trypsin and reduction with dithiothreitol. This inhibitory factor suppressed the proliferation of C3H/HeJ mouse thymocytes and the proliferation of IL-1-dependent cell lines, D10.G4.1 and RPMI 1788, induced by IL-1. However, this inhibitor did not affect the proliferation of IL-2-dependent CTLL-2 cells induced by IL-2, the proliferation of C3H/HeJ mouse thymocytes stimulated with a mitogenic dose of concanavalin A, and the proliferation of IL-6-dependent B9 cells induced by IL-6. Furthermore, the IL-1 inhibitor significantly blocked stimulation of bone resorption in organ cultures of newborn mouse calvaria and inhibited the osteoclast-like cell formation in mouse marrow cultures. A monoclonal antibody prepared against the purified IL-1 inhibitor reacted with mouse recombinant IL-1 receptor antagonist (rIL-1ra), and a polyclonal antibody to mouse rIL-1ra reacted with the IL-1 inhibitor by Western blot (immunoblot) analysis. These results indicate that the IL-1 inhibitor is an identical molecule to rIL-1ra, suggesting that the IL-1 inhibitor (IL-1ra) released by macrophages stimulated with LPS from A. actinomycetemcomitans may play an important mediative role in the development of periodontal disease.

Aggregatibacter actinomycetemcomitans↗
PubMed1994

Responses of glutathione-related enzymes in isolated rat small intestine to Fe(2+)-EDTA-mediated oxidative stress.

The activities of glutathione-related enzymes in isolated rat small intestine were investigated under oxidative stress mediated by Fe(2+)-EDTA. The isoelectric points and approximate molecular weights of the enzymes investigated were first determined. The reduced and oxidized glutathione contents, and low molecular weight thiols in isolated rat small intestine were also studied under oxidative stress. Significant reducing activities of glutathione S-transferase and lactate dehydrogenase were observed accompanied by increases in oxidized glutathione content, whereas thioltransferase, glutathione reductase, glutathione peroxidase and thioredoxin reductase retained the same levels of activity as controls. First-order inactivation of purified rat small intestine glutathione S-transferase including class-alpha, mu and pi was observed and the isozyme class-pi was inactivated at several pH's under Fe(2+)-EDTA-mediated oxidative stress. Leakage of protein and both reduced and oxidized forms of glutathione was significantly increased during incubation with Fe(2+)-EDTA. In conclusion, enzymes which were not inactivated under Fe(2+)-EDTA-mediated oxidative stress may play an important role in cellular antioxidant defenses in the small intestine. Furthermore, enzymes such as glutathione S-transferase, mainly a class-pi isozyme, and lactate dehydrogenase which were inactivated may form part of the barrier against oxidative stress similar to reduced glutathione.

Animals↗
PubMed1994

Metallothionein genes in the nematode Caenorhabditis elegans and metal inducibility in mammalian culture cells.

Genomic DNAs of metallothionein I and II in Caenorhabditis elegans (CeMT-I and CeMT-II) were isolated by YAC library/polytene filter hybridization followed by subcloning of corresponding cosmid clones. Both genes are mapped at chromosome V. Although the similarities of 5'-flanking regions and coding regions have shown only 55-58%, the introns are split at the same position in both genes, indicating that these two genes are originally from the same gene. While several metal responsive elements are conserved among eukaryotes, only one metal responsive element was found in the promoter region in CeMT-II and not in CeMT-I. Indeed, neither of 5'-flanking regions of CeMT-I nor CeMT-II connected to chloramphenicol acetyltransferase reporter gene is responsive to heavy metals in mammalian culture cells by transient transfection analysis. These results would suggest that the metal regulatory factors in C. elegans might be different from those conserved in invertebrates and vertebrates, although the MTs in C. elegans revealed the similarities to mammalian MTs in several points.

Amino Acid Sequence↗