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Biomedical subjects

H Yoshida

Publications and source records attributed to H Yoshida.

At least 829 records · Page 46Linked to original sources
PubMed1997

Phosphorylation of A170 stress protein by casein kinase II-like activity in macrophages.

A170 is an oxidative stress-inducible protein having a Zinc finger domain, two PEST sequences, and many potential phosphorylation sites for serine/threonine kinases. These structural features suggest that the phosphorylation of A170 affects its function and degradation. We have found that A170 is phosphorylated in cultured murine peritoneal macrophages. In addition, using recombinant A170 proteins, we found two proteins of 40 and 44 kDa with kinase activity in cell extracts using an in-gel kinase assay. We compared the properties of the intrinsic A170 kinases with those of mitogen-activated protein kinase (ERK 2), protein kinase A (PKA), casein kinase II (CK II), and protein kinase C, since their catalytic subunits have molecular masses similar to A170 kinases. ERK 2, CK II, and PKA phosphorylated recombinant A170 as a substrate. The 40 and 44 kDa kinases present in the macrophage extract were similar to alpha and alpha' subunits of CK II in respect to substrate specificity, pharmacological properties, immuno-reactivities, and ubiquitous expression in tissues.

Adaptor Proteins, Signal Transducing↗
PubMed1997

Laparoscopic assisted fusion of the lumbosacral spine. A biomechanical and histologic analysis of the open versus laparoscopic technique in an animal model.

STUDY DESIGN: An animal model for laparoscopic lumbosacral fusion. OBJECTIVES: To compare the biomechanical and histologic results of open to laparoscopic lumbosacral discectomy and fusion in an animal model. BACKGROUND DATA: Early clinical reports of laparoscopic lumbosacral fusions are encouraging, but animal experiments have not been reported. METHODS: Ten pigs (50-80 kg) were divided into two groups. Group 1 underwent an open anterior lumbosacral discectomy and fusion at L7-S1 using autologous bone graft and a titanium MOSS (DePuy Motech) cage. Group 2 was identical to Group 1 except that a laparoscopic technique was used. The animals were killed at 3 months, and the lumbosacral spines were harvested for biomechanical and histologic testing. RESULTS: Estimated blood loss and average length of operation, respectively, for the two groups were: Group 1, 50 mL, 2 hours 50 minutes; and Group 2, 40 mL, 3 hours 40 minutes. There were no perioperative or postoperative complications in either group. Motion analysis results showed less motion in lateral bending, flexion, and extension than in the intact specimen in both groups. Tensile testing showed that the stiffness was significantly greater in the open group than in the laparoscopic group (P < 0.004). Histologic examination showed a less extensive discectomy and less bone growth in the implant in the laparoscopic group. Inadequate decortication of end-plates occurred in two animals who underwent laparoscopy. CONCLUSIONS: Although lumbosacral discectomy and implant insertion can be performed using the laparoscopic technique, the construct may not have the same biomechanical strength as that attained with the open procedure. Laparoscopic-assisted lumbosacral fusion surgery requires additional investigation before it is widely used in clinical situations.

Animals↗
PubMed1997

Emergence of osteoblast-like cells in a neoplastic human salivary cancer cell line after treatment with 22-oxa-1alpha, 25-dihydroxyvitamin D3.

A neoplastic clonal cell line, which was prepared by 5-azacytidine treatment of a neoplastic human salivary intercalated duct cell line, was cultivated in the presence of 22-oxa-1alpha, 25-dihydroxyvitamin D3 and 3 mM beta-glycerophosphate. Major alterations, such as expression of type I collagen and alkaline phosphatase as well as of human osteopontin and osteonectin, were observed in these cells with a phenotype similar to osteoblasts. In addition, formation of bone nodule was observed in the cultured cells. The tumors produced by transplantation into nude mice of the clonal cells were treated with 22-oxa-1alpha, 25-dihydroxyvitamin D3 and examined for tumor growth and morphology. Consequently, growth of the treated tumor was significantly suppressed. Moreover, it was found that bone formation was induced in the treated tumor, in which the tumor cells around bone formation expressed human osteopontin and osteonectin mRNA as could be detected by in situ hybridization. The above findings indicate that the emergence of osteoblast-like cells in the human salivary cancer cells occurs in the presence of 22-oxa-1alpha, 25-dihydroxyvitamin D3 and beta-glycerophosphate.

Animals↗
PubMed1997

Serine phosphorylation of Cbl induced by phorbol ester enhances its association with 14-3-3 proteins in T cells via a novel serine-rich 14-3-3-binding motif.

Stimulation of the T cell antigen receptor (TCR).CD3 complex induces rapid tyrosine phosphorylation of Cbl, a protooncogene product which has been implicated in intracellular signaling pathways via its interaction with several signaling molecules. We found recently that Cbl associates directly with a member of the 14-3-3 protein family (14-3-3tau) in T cells and that the association is increased as a consequence of anti-CD3-mediated T cell activation. We report here that phorbol 12-myristate 13-acetate stimulation of T cells also enhanced the interaction between Cbl and two 14-3-3 isoforms (tau and zeta). Tyrosine phosphorylation of Cbl was not sufficient or required for this increased interaction. Thus, cotransfection of COS cells with Cbl plus Lck and/or Syk family protein-tyrosine kinases caused a marked increase in the phosphotyrosine content of Cbl without a concomitant enhancement of its association with 14-3-3. Phorbol 12-myristate 13-acetate stimulation induced serine phosphorylation of Cbl, and dephosphorylation of immunoprecipitated Cbl by a Ser/Thr phosphatase disrupted its interaction with 14-3-3. By using successive carboxyl-terminal deletion mutants of Cbl, the 14-3-3-binding domain was mapped to a serine-rich 30-amino acid region (residues 615-644) of Cbl. Mutation of serine residues in this region further defined a binding motif distinct from the consensus sequence RSXSXP, which was recently identified as a 14-3-3-binding motif. These results suggest that TCR stimulation induces both tyrosine and serine phosphorylation of Cbl. These phosphorylation events allow Cbl to recruit distinct signaling elements that participate in TCR-mediated signal transduction pathways.

14-3-3 Proteins↗
PubMed1997

Stepwise requirement of c-kit tyrosine kinase in mouse ovarian follicle development.

Ovarian follicle development is controlled by the cycling variation of gonadotrophins derived from the central nervous system. Intragonadal signals are also required, especially in the autonomous development of small follicles. Receptor tyrosine kinase c-kit and its ligand SLF (Steel factor) are expressed on the surface of specific populations of follicle-forming cells in a contiguous manner and are thought to have important roles in follicular development. We blocked the interaction of c-kit and its ligand by administering the function-blocking antibody ACK2 to developing mice at various times after birth and monitored ovarian follicle development. A blockade of c-kit function disturbed the onset of primordial follicle development, primary follicle growth, follicular fluid formation of preantral follicles, and penultimate-stage ovarian follicle maturation before ovulation. Ovarian follicle growth was dependent on c-kit during the first 5 days after birth when the functional FSH receptor is not yet expressed in mouse ovary. In contrast, primordial follicle formation and survival, small preantral or antral follicle development, ovulation, and luteinization of the ovulated follicle were not affected by this antibody. These findings indicate the stepwise requirement of c-kit and its ligand interaction system in the developing ovarian follicle and that c-kit with its ligand supports the autonomous development of ovarian follicle independent of gonadotrophins.

Animals↗
PubMed1997

Low micromolar levels of hydrogen peroxide and proteasome inhibitors induce the 60-kDa A170 stress protein in murine peritoneal macrophages.

We previously reported cDNA cloning of a novel oxidative stress protein termed A170 from murine macrophages. Further experiments have demonstrated that exposure of the cells to low levels of H2O2 produced by glucose/glucose oxidase markedly induced the 60-kDa A170 protein. This result suggests that the level of A170 protein can also be controlled at posttranscriptional levels, because we showed previously that H2O2 hardly increased the level of A170 mRNA. We have found that proteasome inhibitors markedly induced the A170 protein after 2 to 8 h similarly to glucose/glucose oxidase, suggesting rapid degradation of the A170 protein by proteasome under normal conditions. Activation of cellular signaling pathways either by epidermal growth factor, lipopolysaccharide or tumor necrosis factor-alpha did not enhance the level of the A170 protein. The levels of glucose oxidase-induced A170 protein did not decrease after the addition of cycloheximide. These results suggest that low levels of H2O2 may stabilize the A170 protein, allowing it to accumulate within cells.

Adaptor Proteins, Signal Transducing↗
PubMed1997

Local injection of OK432 can augment the TH1-type T-cell response in tumor-draining lymph node cells and increase their immunotherapeutical potential.

The effect of local injections with streptococcal preparation OK432 on the therapeutical potential of tumor-draining lymph node (LN) cells was investigated in mice. Peritumoral injections with OK432 on days 2, 4, 6, 8 and 10 showed no effect on the in vivo growth of s.c. inoculated B16F10 melanoma. The B16F10-draining OK432-treated LN cells, however, showed a high level of anti-B16F10 cytolytic activity after an in vitro culture first with both anti-CD3 monoclonal antibody (MAb) and activated B cell blasts, and subsequently with interleukin (IL)-2 without in vitro restimulation. Such in vitro expanded LN cells showed a remarkable antitumor effect against pulmonary metastasis of B16F10 melanoma, even without the concurrent administration of IL-2. In addition, the therapeutical protocol was also found to be moderately effective against poorly immunogenic MCA fibrosarcoma, and the in vivo antitumor effect was specific to the tumor from which the LNs were harvested. Interestingly, 2 kinds of comparative analyses of the cytokines revealed that the B16F10-bearing state induced the draining LN cells to develop a Th2-type response. However, the OK432 treatment was able to effectively augment their Th1-type response. Collectively, our results suggest that peritumoral injections with OK432 significantly increased the therapeutical potential of the tumor-draining LN cells by augmenting their Th1-type response.

Animals↗
PubMed1997

Potent and long-lasting vasodilatory effects of adrenomedullin in humans. Comparisons between normal subjects and patients with chronic heart failure.

BACKGROUND: Adrenomedullin (ADM) is a recently discovered hypotensive peptide that has been isolated from human pheochromocytoma cells. Observations that ADM is produced from cardiovascular tissue and is found in plasma suggest that it may be important in the regulation of regional vascular resistance. METHODS AND RESULTS: Limb vascular responses to ADM were examined in 10 healthy subjects and compared with those in 18 patients with chronic heart failure (CHF). The peptide increased forearm blood flow (FBF) from 2.7 +/- 0.3 to 11.8 +/- 0.9 mL.min-1.100 mL-1 in the control group and from 2.4 +/- 0.3 to 6.5 +/- 0.7 mL.min-1.100 mL-1 in the CHF group. The ADM-induced FBF increase was significantly impaired in the CHF group (P < .01). After cessation of the infusion, an increased FBF level was sustained for > 60 minutes in the control group, whereas in the CHF group the response returned to the baseline in < 30 minutes. The ADM infusion increased forearm skin blood flow in both groups (P < .05), whereas the skin blood flow response was impaired in the CHF group (P < .01). The role of nitric oxide in ADM-induced vasorelaxation was also studied in 11 healthy subjects and 6 patients with CHF. FBF and skin blood flow responses during ADM administration were significantly attenuated by NG-monomethyl-L-arginine administration in healthy control subjects (P < .05), whereas both flow responses remained the same in the CHF group. CONCLUSIONS: These observations demonstrate that ADM exerts a potent and long-lasting vasodilatory effect on skeletal muscle arteries with involvement of nitric oxide-dependent mechanisms in normal human peripheral vasculature and that these vascular effects are significantly attenuated in patients with CHF, in part because of impaired production of nitric oxide in the forearm resistance vessels.

Adrenomedullin↗
PubMed1997

Structural differences and the presence of unsubstituted amino groups in heparan sulphates from different tissues and species.

This study presents a comparison of heparan sulphate chains isolated from various porcine and bovine tissues. 1H-NMR spectroscopy (500 MHz) was applied for structural and compositional studies on intact heparan sulphate chains. After enzymic digestion of heparan sulphate using heparin lyase I (EC 4.2.2.7) II and III (EC 4.2.2.8), the compositions of unsaturated disaccharides obtained were determined by analytical capillary electrophoresis. Correlations between the N-sulphated glucosamine residues and O-sulphation and between iduronic acid content and total sulphation were discovered using the data obtained by NMR and disaccharide analysis. Heparan sulphate chains could be classified into two groups based on the sulphation degree and the iduronic acid content. Heparan sulphate chains with a high degree of sulphation possessed also a significant number of iduronic acid residues and were isolated exclusively from porcine brain, liver and kidney medulla. The presence and amount of N-unsubstituted glucosamine residues (GlcNp) was established in all of the heparan sulphates examined. The structural context in which this residue occurs was demonstrated to be: high sulphation domain --> 4)-beta-D-GlcAp-(1 --> 4)-alpha-D-GlcNp-(1 --> 4)-beta-D-GlcAp-(1 --> low sulphation domain (where GlcNp is 2-amino-2-deoxyglucopyranose, and GlcAp is glucopyranosyluronic acid), based on the isolation and characterization of a novel, heparin lyase III-derived, GlcNp containing tetrasaccharide and hexasaccharide. The results presented suggest that structural differences may play a role in important biological events controlled by heparan sulphate in different tissues.

Amino Sugars↗
PubMed1997

An alternative pathway for STAT activation that is mediated by the direct interaction between JAK and STAT.

JAK is believed to be an essential tyrosine kinase that mediates signals from the cytokine receptor to its downstream events. JAK associates with the cytoplasmic domain of the type I cytokine receptor superfamily and upon the ligand stimulation it can be activated, resulting in the receptor phosphorylation. In signaling from gp130, a common signal transducer for the IL-6 family cytokines, STAT3, a transcription factor that contains an SH2 domain, is recruited by phosphotyrosines on gp130 and is subsequently phosphorylated by gp130-associated JAKs. In this study, we attempted to find a new target for JAK that is directly activated by JAK, independent of gp130 tyrosine phosphorylation, by using a yeast two-hybrid system. In the process we found that the JH2 domain of JAK1, JAK2 or JAK3 could specifically associate with the carboxy-terminal portion of STAT5, but not with STAT3 or STAT1. The interaction was confirmed using both a transient expression system in a cell line and a GST-fusion protein binding assay. Furthermore, we showed that the activation of STAT5 via gp130 did not need any phosphotyrosines on gp130 while that of STAT3 strictly depended on phosphotyrosines on gp130. Mutations of STAT5 that eliminated the interaction with JAK1 reduced the activation of STAT5 upon the gp130 stimulation, although such mutants could be still activated through erythropoietin receptor. These results indicate that STATs are activated through cytokine receptors by two distinct mechanisms, one dependent on receptor tyrosine phosphorylation and the other mediated by the JAK-STAT direct interaction.

Animals↗
PubMed1997

Lumbar nerve root compression caused by lumbar intraspinal gas. Report of three cases.

STUDY DESIGN: Three cases of lumbar nerve root compression associated with intradiscal gas formation and its migration are reported. OBJECTIVES: To establish the pathogenic mechanism by which gas is formed in the spinal canal. SUMMARY OF BACKGROUND DATA: Few such cases have been reported of patients with clinical symptoms of lumbar radiculopathy resulting from gas in the spinal canal. METHODS: Surgery with needle aspiration of the gas was performed in all cases. RESULTS: The presence of the gas was detected in the extradural space in one patient and within the nerve sheath in two patients. After surgery, all patients recovered well, and there were no obvious remnant neurologic abnormalities. One year later, all patients were completely free of symptoms. CONCLUSIONS: Surgery with needle aspiration of the gas resulted in clinical improvement in all cases, confirming that intraspinal gas is an important cause of lumbar radiculopathy. The composition of the gas in one patient was analyzed by gas chromatography, which revealed an overwhelming preponderance of nitrogen. A relationship between a gas-containing pseudocyst in the spinal canal and a degenerated intervertebral disc was identified, a key finding for understanding the precise nature of this disorder. Intradiscal gas formation and its outward migration as a sequel of intervertebral disc degeneration also has been addressed in this report.

Aged↗
PubMed1997

Induction of cyclin-dependent kinase inhibitor, p21WAF1, by treatment with 3,4-dihydro-6-[4-(3,4)-dimethoxybenzoyl)-1-piperazinyl]-2(1H)-quinoline (vesnarinone) in a human salivary cancer cell line with mutant p53 gene.

It has been found by PCR-SSCP analysis and direct DNA sequencing that a human salivary adenosquamous carcinoma-forming cell line, TYS, has a mutant p53 gene at codon 281Asp-->His. When TYS cells were treated with a differentiation-inducing agent, vesnarinone, cellular proliferation was significantly inhibited on the basis of MTT assay. In addition, it has been found by Northern blotting and/or immunoblotting that expression of p21WAF1 and transforming growth factor-beta (TGF-beta) is up-regulated by treating TYS cells with vesnarinone. TGF-beta 1 alone also induced p21WAF1 expression in TYS cells. Moreover, it has been shown by ELISA that the treatment of TYS cells with vesnarinone results in the enhanced generation of latent TGF-beta 1. The expression of TGF-beta receptor (T beta R), including T beta R-I, T beta R-II and T beta R-III, on TYS cells was detected by affinity cross-linking using 125I-TGF-beta 1 and addition of active TGF-beta 1 into serum-free culture medium inhibited the growth of TYS cells in a concentration-dependent manner. These findings suggest that vesnarinone might directly induce expression of p21WAF1 gene in TYS cells, the product of which may be associated with the inhibition of cell growth and induce differentiation.

Antineoplastic Agents↗
PubMed1997

Mechanistic studies on metabolic chiral inversion of 4-(4-methylphenyl)-2-methylthiomethyl-4-oxobutanoic acid (KE-748), an active metabolite of the new anti-rheumatic agent 2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298), in rats.

The chiral inversion properties of 4-(4-methylphenyl)-2-methylthiomethyl-4-oxobutanoic acid (KE-748), an active metabolite of 2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298), were compared with those of ibuprofen in rats. After administration of R(-)-[2 alpha-2H]KE-748, S(+)-KE-748 was present in the rat plasma, and the deuterium atoms of the S(+)-enantiomer were almost all replaced by hydrogen atoms. After administration of S(+)-[2 alpha-2H]KE-748, the deuterium content of S(+)-KE-748 in the plasma remained intact. In the in vitro study, using a cell-free system and rat liver homogenates, the chiral inversion of ibuprofen was apparent when both CoA and ATP were present; however, KE-748 was not inverted. In the study on isolated rat hepatocytes, the unidirectional chiral inversion from R(-)-to S(+)-enantiomer was observed for both ibuprofen and KE-748. When R(-)-ibuprofen was incubated with medium and long chain fatty acids (carbon chain length C6 to C16), using isolated hepatocytes, the chiral inversion decreased significantly. On the other hand, when R(-)-KE-748 was incubated with short and medium chain fatty acids (carbon chain length C3 to C8), chiral inversion was inhibited markedly. To induce hepatic microsomal long chain fatty acid CoA ligase, rats were treated with clofibric acid (CF rats). In both in vitro and in vivo experiments on CF rats, chiral inversion from R(-)-to S(+)-ibuprofen was enhanced significantly compared with that in controls, whereas the enhancement was not observed in the case of R(-)-KE-748. There was no influence of benzoic acid, a typical substrate on medium chain fatty acid CoA ligase in the mitochondrial matrix, on chiral inversion of R(-)-ibuprofen, using, isolated hepatocytes. In contrast, the chiral inversion from R(-)-to S(+)-KE-748 was strongly inhibited in the presence of benzoic acid. These results indicate that chiral inversion of R(-)-KE-748 may proceed via formation of the CoA-thioester intermediate with loss of the 2 alpha-methine proton, in a manner similar to that seem with R(-)-ibuprofen. However, the enzymes needed to form CoA-thioester of R(-)-KE-748 differ from those for R(-)-ibuprofen.

Animals↗
PubMed1997

Possible contribution of active MMP2 to lymph-node metastasis and secreted cathepsin L to bone invasion of newly established human oral-squamous-cancer cell lines.

We have established human oral-squamous-cancer cell lines, BHY and HN, derived from non-metastatic cancer and metastatic cancer respectively. We examined the expression of matrix-degrading enzymes and their inhibitors in these cell lines. Both cell lines expressed pro-matrix metalloproteinase (MMP)1, proMMP2, proMMP9, membrane-type MMP and urokinase-type plasminogen activator. In addition to these enzymes, BHY cells secreted proMMP7 and procathepsin L, while HN cells secreted a large amount of active MMP2. BHY cells secreted a tissue inhibitor of matrix metalloproteinase, TIMP2, but only a trace level of TIMP1. Contrary to BHY cells, HN cells secreted TIMP1, but only a trace level of TIMP2. When we inoculated these cells into the masseter muscle of nude mice, both types of cell formed solid tumors, whose microscopic appearance was identical to that of the original tumors. BHY tumors were highly differentiated squamous-cell carcinomas, and invasive to the masseter muscle and the mandibular bone. Despite their local aggressiveness, BHY tumors did not metastasize to any distant organs. HN tumors were poorly differentiated squamous-cell carcinomas, weakly invasive to the muscle, but not to the mandibular bone. However, HN tumors frequently metastasized to cervical lymph nodes. These results suggest that the net activity of MMP2 (active MMP2/TIMP2) and cathepsin L secreted from cancer cells may contribute respectively to lymph-node metastasis and to bone invasion by oral cancer cells.

Animals↗
PubMed1997

Chromosomal rearrangement t(11;22) in extraskeletal Ewing's sarcoma and primitive neuroectodermal tumour analysed by fluorescence in situ hybridization using paraffin-embedded tissue.

The clonal chromosomal rearrangement t(11;22) has been reported by karyotypic analysis to be specific for Ewing's sarcoma of bone and soft tissue origin as well as primitive neuroectodermal tumour. In this report, immunohistological analysis of MIC 2 expression and fluorescence in situ hybridization (FISH) were performed using paraffin-embedded tissues. We examined t(11;22) in the nuclei isolated from two Ewing's sarcomas, four primitive neuroectodermal tumours, and three neuroblastomas, which served as negative controls by FISH with an alpha-satellite DNA probe for chromosome 11, a chromosome 22 marker probe, and whole chromosome painting probes for both chromosomes 11 and 22. Both cases of Ewing's sarcoma and the four primitive neuroectodermal tumour specimens were immunoreactive for MIC 2. Both Ewing's sarcomas and three of the four primitive neuroectodermal tumours contained the tumour-specific t(11;22), but the three neuroblastomas did not show this translocation. Based on the cytogenetic results and on the immunohistological investigation of MIC 2 expression, Ewing's sarcoma is suggested to be related closely to primitive neuroectodermal tumour. FISH is a useful aid in determining the tumour type of Ewing's sarcoma and putative related tumours.

12E7 Antigen↗
PubMed1997

Involvement of Fas antigen in ovarian follicular atresia and luteolysis.

The Fas antigen (Fas) is a cell-surface receptor protein that mediates apoptosis-inducing signals and plays an important role in the immune system. Significant amounts of Fas mRNA can be detected not only in lymphoid organs but also in the liver, heart, and ovary. In the ovary, apoptosis is thought to cause follicular atresia and luteolysis. We have investigated the involvement of Fas in these events. Here we report that Fas protein is expressed on granulosa and luteal cells but not on oocytes in the ovary. An injection of anti-Fas monoclonal antibody with apoptosis-inducing activity into adult mice enhanced follicular atresia and luteolysis. After the injection, the corpora lutea disappeared and the number of follicles containing pyknotic granulosa cells increased. There were also fewer ovulated ova and lower levels of luteal cell-produced progesterone. Furthermore, as the result of a non-functional Fas/Fas ligand system, mature ovaries from the mouse mutant/pr (lymphoproliferation) were histologically abnormal in terms of follicular development, in that the number of secondary follicles significantly increased. These results suggested that Fas plays an important role in follicular atresia and luteolysis in the ovarian physiology of adult mice.

Animals↗