Biomedical subjects

N Kitamura

Publications and source records attributed to N Kitamura.

At least 73 records · Page 4Linked to original sources

PubMed2000

Hepatocyte growth factor activator inhibitor type 1 is a specific cell surface binding protein of hepatocyte growth factor activator (HGFA) and regulates HGFA activity in the pericellular microenvironment.

Hepatocyte growth factor activator (HGFA) is responsible for proteolytic activation of the precursor form of hepatocyte growth factor in injured tissues. To date, two specific inhibitors of HGFA have been identified, namely HGFA inhibitor type 1 (HAI-1) and type 2 (HAI-2)/placental bikunin (PB). Both inhibitors are first synthesized as integral membrane proteins having two Kunitz domains and a transmembrane domain, and are subsequently released from cell surface by shedding. Here we show that an active form of HGFA is specifically complexed with membrane-form HAI-1, but not with HAI-2/PB, on the surface of epithelial cells expressing both inhibitors. This binding required the enzyme activity of HGFA. The selective binding of HGFA to the cell surface HAI-1 was further confirmed in an engineered system using Chinese hamster ovary cells, in which only the cells expressing HAI-1 retained exogenous HGFA. The binding of HGFA to HAI-1 was reversible, and no irreversible modifications affecting the enzyme activity occurred during the binding. Importantly, HAI-1 and the HGFA.HAI-1 complex were quickly released from the cell surface by treatment with phorbol 12-myristate 13-acetate or interleukin 1beta accompanying the generation of 58-kDa fragments of HAI-1, which are less potent against HGFA, as well as significant recovery of HGFA activity in the culture supernatant. This regulated shedding was completely inhibited by BB3103, a synthetic zinc-metalloproteinase inhibitor. We conclude that HAI-1 is not only an inhibitor but also a specific acceptor of active HGFA, acting as a reservoir of this enzyme on the cell surface. The latter property appears to ensure the concentrated pericellular HGFA activity in certain cellular conditions, such as tissue injury and inflammation, via the up-regulated shedding of HGFA.HAI-1 complex. These findings shed light on a novel function of the integral membrane Kunitz-type inhibitor in the regulation of pericellular proteinase activity.

Animals↗

PubMed2000

A deubiquitinating enzyme UBPY interacts with the Src homology 3 domain of Hrs-binding protein via a novel binding motif PX(V/I)(D/N)RXXKP.

Hrs-binding protein (Hbp) is a Src homology 3 (SH3) domain-containing protein that tightly associates with Hrs. Hbp together with Hrs is thought to play a regulatory role in endocytic trafficking of growth factor-receptor complexes through early endosomes. Association of Hbp with a binding partner(s) via the SH3 domain seems to be essential for Hbp to exert its function. In this study, we searched for Hbp-binding proteins by a far Western screening and isolated a mouse cDNA clone encoding a deubiquitinating enzyme mUBPY as an Hbp SH3-binding protein. mUBPY has two Hbp-SH3 domain binding sites. Mutagenic analysis identified a consensus sequence PX(V/I)(D/N)RXXKP as the Hbp-SH3 domain binding motif. It is a novel SH3-binding motif and does not contain the canonical proline-rich consensus binding motif, PXXP. Ubiquitination of growth factor receptors is thought to regulate their intracellular degradation. Thus, UBPY may play a regulatory role in the degradation by interaction with the SH3 domain of Hbp via the novel SH3-binding motif.

3T3 Cells↗

PubMed2000

Involvement of hrs binding protein in IgE receptor-triggered exocytosis in RBL-2H3 mast cells.

Hrs binding protein (Hbp) tightly associated with Hrs is thought to play a regulatory role in vesicular trafficking during endocytosis and exocytosis. In this study, we have expressed dominant-negative mutants of Hbp to evaluate their effects on the degranulation of secretory granules in RBL-2H3 mast cells. The dominant-negative mutants of Hbp significantly inhibited IgE receptor (FcepsilonRI)-triggered secretory response as tested by beta-hexosaminidase release. These results suggest that Hbp functions as a regulator in the FcepsilonRI-triggered degranulation of secretory granules in mast cells.

3T3 Cells↗

PubMed2000

Activation of hepatocyte growth factor/scatter factor in colorectal carcinoma.

Activation of hepatocyte growth factor/scatter factor (HGF/SF) in the extracellular milieu is a critical limiting step in the HGF/SF-induced signaling pathway mediated by Met receptor tyrosine kinase, which has potentially important roles in tumor biology and progression. However, little is known concerning the regulation of HGF/SF activation in tumors. Immunoblot analysis revealed that the activation of HGF/SF was enhanced significantly in colorectal carcinoma tissues compared with the corresponding normal mucosa. Serum-free conditioned media of cultured human colorectal carcinoma cell lines contained HGF/SF-activating activity, and the addition of a single-chain precursor form of HGF/SF to the serum-free culture of these cells resulted in HGF/SF-dependent modulation of cellular phenotypes, such as increased scattering and enhanced secretion of vascular endothelial growth factor. This processing activity was enhanced by thrombin treatment but was inhibited significantly by a neutralizing antibody against HGF activator (HGFA), a factor XIIa-like serine proteinase believed to be expressed mainly in the liver. The activity was also inhibited by recombinant HGFA inhibitor type 1 (HAI-1). The presence of HGFA mRNA and secretion of HGFA protein were confirmed in the cell lines. Therefore, extrahepatic expression of HGFA in the colorectal carcinoma cells could be responsible for the single-chain HGF/SF-processing activity of the cells. We examined the expression of HGFA and HAI-1 in human colorectal mucosa and adenoma-carcinoma sequence. Immunohistochemically, HGFA was stained weakly in the normal enterocytes, and immunoreactivity was increased modestly in the neoplastic differentiation. The subcellular localization of HGFA immunoreactivity was altered in carcinoma cells showing basal or cell-stroma interface staining patterns, compared with normal and adenoma cells with a supranuclear or apical staining pattern. In contrast to HGFA, the expression of HAI-1 decreased significantly in carcinoma cells relative to the adjacent normal or adenoma cells, indicating that the net balance between HGFA and HAI-1 shifts in favor of HGFA in carcinomas. In fact, pro-HGFA and the active form of HGFA proteins increased in carcinoma tissue compared with the corresponding normal mucosa. It was concluded that HGFA is expressed in colorectal mucosa and tumors and could be involved in the activation of HGF/SF in colorectal carcinomas. Therefore, the balance between HGFA and HAI-1 could play an important role in the regulation of HGF/SF activity in colorectal carcinomas.

Adenocarcinoma↗

PubMed2000

Early endosomal localization of hrs requires a sequence within the proline- and glutamine-rich region but not the FYVE finger.

Hrs is an early endosomal protein that is tyrosine-phosphorylated in cells stimulated with growth factors. Hrs is thought to play a regulatory role in endocytosis of growth factor-receptor complexes through early endosomes. Early endosomal localization of Hrs seems to be essential for Hrs to exert its function in the endocytosis. Hrs has a FYVE finger domain that binds specifically to phosphatidylinositol 3-phosphate in vitro. The FYVE finger is a likely domain that mediates membrane association of endosomal proteins. In this study, we examined whether the FYVE finger participates in early endosomal targeting of Hrs. Hrs with a zinc binding-defective FYVE finger was still localized to early endosomes. In addition, the N-terminal FYVE finger-containing fragment of Hrs showed a cytosolic distribution in mammalian cells. These results indicate that the FYVE finger is not required for the localization of Hrs to early endosomes. Furthermore, by analyzing a series of deletion mutants of Hrs, we identified a sequence of about 100 amino acids within the C-terminal proline- and glutamine-rich region as a domain essential for the targeting of Hrs to early endosomes.

Animals↗

PubMed2000

NESK, a member of the germinal center kinase family that activates the c-Jun N-terminal kinase pathway and is expressed during the late stages of embryogenesis.

The c-Jun N-terminal kinase (JNK) signaling pathway plays a crucial role in cellular responses stimulated by stress-inducing agents and proinflammatory cytokines. The group I germinal center kinase family members selectively activate the JNK pathway. In this study, we have isolated a mouse cDNA encoding a protein kinase homologous to Nck-interacting kinase (NIK), a member of the group I germinal center kinase family. This protein kinase is expressed during the late stages of embryogenesis, but not in adult tissues, and thus named NESK (NIK-like embryo-specific kinase). NESK selectively activated the JNK pathway when overexpressed in HEK 293 cells but did not stimulate the p38 kinase or extracellular signal-regulated kinase (ERK) pathways. NESK-induced JNK activation was inhibited by the dominant negative mutants of MEKK1 and MKK4. Tumor necrosis factor (TNF)-alpha or TNF receptor-associated factor 2 (TRAF2) stimulated the NESK activity. Furthermore, the dominant negative NESK mutant inhibited the JNK activation induced by TNF-alpha or TRAF2. These results suggest that NESK, a novel activator of the JNK pathway, functions in coupling TRAF2 to the MEKK1 --> MKK4 --> JNK kinase cascade during the late stages of mammalian embryogenesis.

Amino Acid Sequence↗

PubMed2000

NE-dlg, a mammalian homolog of Drosophila dlg tumor suppressor, induces growth suppression and impairment of cell adhesion: possible involvement of down-regulation of beta-catenin by NE-dlg expression.

Membrane-associated guanylate kinases (MAGUKs) are known to function as scaffolds for forming multiprotein complexes at the synaptic junctions of neuronal cells and at sites of epithelial cell-cell contact. In Drosophila, mutations of the lethal (1)-discs large (dlg) gene, which encodes a MAGUK protein, leads to post-synaptic structure defects in neuronal cells and neoplastic overgrowth of epithelial cells. We previously showed that NE-dlg (neuronal and endocrine dlg), a human homolog of the dlg, plays a crucial role in formation of synaptic structure in human neuronal cells. Here we demonstrate that NE-dlg, similar to Drosophila dlg, is involved in regulation of cell cycle progression and adhesive ability of non-neuronal cells. Overexpression of NE-dlg in proliferating cells including various cancer cell lines induced growth suppression and impairment of cell adhesive ability. Furthermore, NE-dlg overexpression caused the down-regulation of beta-catenin in cancer cells regardless of mutations in the APC (adenomatous polyposis coli) gene. The PDZ domains of NE-dlg were found to be essential for the growth suppression, loss of adhesive property and down-regulation of beta-catenin. We propose that NE-dlg regulates the cell growth and adhesive ability by controlling the level of beta-catenin through an APC-independent pathway. Inactivation of NE-dlg may therefore contribute to development and/or progression of human neoplasms.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Requirement of regulated activation of Ras for response of MDCK cells to hepatocyte growth factor/scatter factor.

Hepatocyte growth factor/scatter factor (HGF/SF) induces cell scattering, migration, and branching tubule formation of MDCK cells. To examine the role of the Ras protein in the HGF/SF-induced responses, we constructed MDCK cell clones expressing either inducible dominant-negative Ras or constitutively activated Ras and analyzed their effects on responses of cells to HGF/SF. Induced expression of dominant-negative Ras prevented cell dissociation required for cell scattering, migration, and cystic formation as well as branching morphology required for branching tubule formation. Constitutively activated Ras induced cell dissociation, but not a scattered fibroblastic morphology even in the presence of HGF/SF. MDCK cells expressing constitutively activated Ras migrated at a level similar to that of wild-type MDCK cells stimulated by HGF/SF. MDCK cells expressing constitutively activated Ras showed disorganized growth in three-dimensional culture and did not form the branching tubule structures. These results indicate that activation of the Ras protein is essential for the cell scattering, migration, and branching tubule formation of MDCK cells induced by HGF/SF, and a properly regulated activation is required for some stages of the HGF/SF-induced responses of MDCK cells.

Animals↗

PubMed2000

Mouse hepatocyte growth factor activator gene: its expression not only in the liver but also in the gastrointestinal tract.

A cDNA encoding mouse hepatocyte growth factor activator (HGFA) has been cloned by RT-PCR, based on the screening result from the database of expressed sequence tags. Subsequently, its gene was cloned from a mouse genomic bacterial artificial chromosome library using the cDNA as a probe. Sequencing analysis revealed that mouse HGFA protein deduced from the cDNA, similar to its human and rat counterparts, has two epidermal growth factor-like domains, type 1 and 2 fibronectin homology domains, a single kringle domain and a catalytic domain of serine proteinase, and the gene consists of 14 exon spanning approximately 7.5 kb. Interestingly, mouse HGFA mRNA was detected not only in the liver but also in the gastrointestinal tract by RNA blot analysis. Since hepatocyte growth factor (HGF) is up-regulated in the damaged gastrointestinal mucosa, our present data suggest that HGFA might activate proHGF directly in the gastrointestinal mucosa and play an important role in wound repair throughout the gastrointestinal tract.

Amino Acid Sequence↗

PubMed2000

Emission and metal- and ligand-centered-redox characteristics of the hexarhenium(III) clusters trans- and cis-[Re6(mu 3-S)8Cl4(L)2]2-, where L is a pyridine derivative or pyrazine.

Preparations of a series of face-capped octahedral hexarhenium(III) clusters having two N-heterocyclic ligands, [Bu4N]2[trans-[Re6(mu 3-S)8Cl4(L)2]] (Bu4N+ = tetra-n-butylammonium cation; L = pyrazine (1a), 4,4'-bipyridine (3a), 4-methylpyridine (5a), 4-(dimethylamino)pyridine (6a)) and their cis analogues (1b, 3b, 5b, and 6b, respectively), and their electrochemical and photophysical properties have been reported. An X-ray crystal structure determination has been carried out for 1a to confirm the trans configuration (C40H80N6S8Cl4Re6, orthorhombic, space group Cmca (No. 64), a = 19.560(5) A, b = 19.494(4) A, c = 18.592(4) A, beta = 115.76(2) degrees, Z = 4). The redox potential of the reversible ReIII6/ReIII5ReIV process of these complexes and previously reported [Bu4N]2[trans- and cis-[Re6(mu 3-S)8Cl4(4-cyanopyridine)2]] (2a and 2b, respectively) and [Bu4N]2[trans- and cis-[Re6(mu 3-S)8Cl4(pyridine)2]] (4a and 4b, respectively) in acetonitrile depends linearly on the pKa of the N-heterocyclic ligands, with the potentials being more negative with basic ligands. The ligand-centered-redox waves for 1a, 1b, 2a, and 2b were observed as split waves (delta E1/2 = 90-140 mV), the extent of the splitting being larger for the cis isomer and largest for the pyrazine complexes. Electronic interaction between the two ligands through the [Re6(mu 3-S)8]2+ core has been suggested. The second ligand-reduction wave was also observed for 3a and 3b, the potential being shifted positively to coalesce with the first reduction wave on addition of the weak proton donor imidazole. This is accounted for by the proton-coupled redox reaction at the free pyridyl site of the 4,4'-bipyridine ligands. All of the complexes show luminescence in acetonitrile at room temperature. While the complexes of pyridine and 4-methylpyridine show photophysical characteristics (lambda em 740-750 nm, phi em 0.031-0.057, tau em 4.2-6.2 microseconds) similar to those (770 nm, 0.039, and 6.3 microseconds, respectively) of [Re6(mu 3-S)8Cl6]4-, emissions of other complexes are significantly weak with lambda em, phi em, and tau em values in the ranges 763-785 nm, 0.0010-0.0017, and 0.013-0.029 microsecond, respectively. Suggestions are given for the excited states localized on the cluster core and the ligand pi* orbitals.

Journal Article↗

PubMed2000

Conserved expression of hepatocyte growth factor activator inhibitor type-2/placental bikunin in human colorectal carcinomas.

Hepatocyte growth factor activator inhibitor type 2 (HAI-2) was recently identified as a potent inhibitor of hepatocyte growth factor activator. It was also independently reported as placental bikunin (PB) and as a protein over-expressed in pancreatic cancer. The expression of HAI-2/PB was analyzed in human normal colon mucosa, adenomas, and carcinomas. HAI-2/PB mRNA was consistently expressed in the colorectal mucosa. The expression was conserved in the neoplastic colorectal mucosa, and no relationship was found between HAI-2/PB mRNA levels and tumor stages. Moreover, 13 out of 14 colorectal carcinoma cell lines expressed HAI-2/PB mRNA. Immunohistochemically, HAI-2/PB proteins were predominantly stained beneath the apical surface of normal enterocytes. In tumor tissues, rather disarranged intracytoplasmic granular staining was observed. The HAI-2/PB immunoreactivity was well conserved in the colonic adenoma-carcinoma sequence, and this protein may have important unknown function in the intestinal mucosa.

Adult↗

PubMed2000

A study on liquid-liquid distribution based on single picoliter droplets and in situ electrochemical measurements.

A microelectrode technique combined with the microcapillary injection/manipulation of a single organic droplet in water was developed. The technique was applied to the study of the distribution of a ferrocene derivative across a single-picoliter-nitrobenzene-droplet/water interface and to the simultaneous in situ electrochemical determination of the distributed solute in the picomole-femtomole range. The liquid-liquid distribution processes were discussed in terms of droplet size and solute concentration (in water) dependencies of the interfacial mass transfer rate.

Electrochemistry↗

PubMed2000

Mediastinal growing teratoma syndrome.

A 27-year-old man had undergone orchiectomy and chemotherapy for testicular cancer. Despite normalization of raised tumor marker levels after postoperative chemotherapy, computed tomographic scanning demonstrated multiple swellings of the para-aortic lymph nodes with extension from beneath the aortic arch to the bifurcation of the descending aorta. Open biopsies of the para-aortic lymph nodes disclosed mature teraroma without malignant cells. The patient presented the typical features of mediastinal and retroperitoneal growing teratoma syndrome. A two stage resection of the tumors was performed via laparotomy and left thoracotomy. Histological examination of the resected specimens revealed a mature teratoma component without malignant cells. Upon follow-up sixteen months later, the patient was well and without recurrence.

Adult↗

PubMed2000

Oblique plication for repair of the atrialized ventricle and tricuspid incompetence of Ebstein's anomaly.

An 8-year-old girl who weighed 42 kg presented Ebstein's anomaly with severe tricuspid incompetence, and mild systemic cyanosis during exercise. A new reconstructive procedure for this complex anomaly was used. Oblique transference of the displaced posterior leaflet was performed, which resulted in plication of the atrialized ventricle and reduction in the tricuspid annular diameter. This procedure requires neither detachment nor closure of the tricuspid valve.

Child↗

PubMed2000

Repair of partial anomalous pulmonary venous connection with a minimal atriotomy.

We present an alternative surgical technique for the repair of a partial anomalous pulmonary venous connection to the higher segment of the superior vena cava. Although the atriotomy is limited in this technique, a sufficiently large systemic venous chamber overlapping to the outside of the superior vena cava can be created.

Adolescent↗

PubMed2000

Comparison of Sepsis-related Organ Failure Assessment (SOFA) score and CIS (cellular injury score) for scoring of severity for patients with multiple organ dysfunction syndrome (MODS).

OBJECTIVE: To evaluate the usefulness of cellular injury score (CIS) and Sepsis-related Organ Failure Assessment (SOFA) score for determination of the severity of multiple organ dysfunction syndrome (MODS). DESIGN: A prospective observational study. SETTING: A medical and surgical intensive care unit (ICU) of a teaching hospital. PATIENTS: Forty-seven consecutive MODS patients. MEASUREMENTS AND RESULTS: SOFA score and CIS were measured every day for 12 months for 47 MODS patients. Comparison was made of the SOFA score and CIS for usefulness in the scoring of severity of MODS in 26 survivors and 21 non-survivors. In addition, receiver operating characteristics (ROC) analysis was used to determine the usefulness of these two indexes as predictors of prognosis. No significant differences were found on admission between the survivors and non-survivors, but significant differences between the two subgroups (p < 0.001) were found in maximum value within 1 week after admission and maximum value during the course of treatment for both indexes. Analysis of changes after admission indicated that significant differences between survivors and non-survivors began to appear on day 3 of admission for both indexes; at that time SOFA score began to deteriorate in the non-survivors while CIS began to improve in the survivors. ROC analysis demonstrated that the area under the ROC curve was 0.769 for SOFA scores and 0.760 for CIS. CONCLUSIONS: Both SOFA score and CIS sequentially reflected the severity of MODS. Furthermore, they were comparable in diagnostic value as predictors of prognosis. These findings may indicate the possibility that MODS is a summation of effects of cellular injury. In addition, sequential evaluation of both SOFA score and CIS would provide a more accurate prediction of prognosis than conventional methods.

Aged↗

PubMed2000

Altered plasma antigen levels of tissue factor pathway inhibitor during open-heart surgery.

Tissue factor pathway inhibitor (TFPI) is a potent inhibitor of the extrinsic pathway of the coagulation cascade that is activated by cardiopulmonary bypass (CPB) during open-heart surgery. In the present study, we investigated whether the plasma TFPI antigen increases during CPB in 12 patients who underwent bypass procedures. The plasma levels of free and total TFPI antigens were measured using an enzyme-linked immunosorbent assay, and heparin concentrations were measured using a chromogenic substrate assay. We found that changes in the total plasma TFPI antigen level were significantly dependent on the level of free TFPI antigen (r = 0.96, P < 0.0001) which increased significantly after heparin injection (P < 0.0001), and increased further during the bypass period (P < 0.005). The increased free TFPI antigen level during CPB correlated with the duration of bypass (r = 0.65, P = 0.02). When heparin was neutralized by protamine, the free TFPI antigen level decreased immediately, but remained higher than the preoperative level (P < 0.005). These results suggest that plasma TFPI antigen levels increase during CPB.

Anticoagulants↗

PubMed2000

A new technique for debridement in rheumatic valvular disease: the rasping procedure.

BACKGROUND: Valve repair is superior to valve replacement in terms of postoperative risk. However, the conventional techniques used for valve repair in patients with rheumatic valvular disease have not resulted in a good long-term outcome. METHODS: We developed a novel "rasping procedure" using an electric rasper for debridement in rheumatic valvular diseases. Between April 1986 and December 1996, the rasping procedure was performed on the aortic valve (A-rasping) of 24 patients who exhibited moderate stenosis with mild regurgitation. Between June 1992 and December 1996, this procedure was performed on the mitral valve (M-rasping) of 12 patients who exhibited mitral stenosis with mild regurgitation. RESULTS: Among the 24 patients on whom A-rasping was performed, 4 patients did not show any improvement after the A-rasping procedure, and required valve replacement. In each of the remaining 20 cases, the transvalvular pressure gradient decreased, and regurgitation disappeared, or was reduced to a trivial level. In all 12 patients on whom M-rasping was performed, the transvalvular pressure gradient significantly decreased, orifice area significantly increased, and the regurgitation disappeared. Postoperative echocardiographic examinations were performed during a mean follow-up period of 110 months in the patients who received A-rasping, and that of 50 months in the patients who received M-rasping. A significant change in the valve requiring subsequent surgery was not seen in any of the patients. CONCLUSIONS: The rasping procedure is an effective technique for excising rheumatic valvular hypertrophic lesions. It resulted in good intermediate to long-term outcome.

Adult↗