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A Kawai

Publications and source records attributed to A Kawai.

At least 55 records · Page 3Linked to original sources
PubMed2000

Mapping of epitopes and structural analysis of antigenic sites in the nucleoprotein of rabies virus.

Linear epitopes on the rabies virus nucleoprotein (N) recognized by six MAbs raised against antigenic sites I (MAbs 6-4, 12-2 and 13-27) and IV (MAbs 6-9, 7-12 and 8-1) were investigated. Based on our previous studies on sites I and IV, 24 consecutively overlapping octapeptides and N- and C-terminal-deleted mutant N proteins were prepared. Results showed that all three site I epitopes studied and two site IV epitopes (for MAbs 8-1 and 6-9) mapped to aa 358-367, and that the other site IV epitope of MAb 7-12 mapped to aa 375-383. Tests using chimeric and truncated proteins showed that MAb 8-1 also requires the N-terminal sequence of the N protein to recognize its binding region more efficiently. Immunofluorescence studies demonstrated that all three site I-specific MAbs and one site IV-specific MAb (7-12) stained the N antigen that was diffusely distributed in the whole cytoplasm; the other two site IV-specific MAbs (6-9 and 8-1) detected only the N antigen in the cytoplasmic inclusion bodies (CIB). An antigenic site II-specific MAb (6-17) also detected CIB-associated N antigen alone. Furthermore, the level of diffuse N antigens decreased after treatment of infected cells with cycloheximide. These results suggest that epitopes at site I are expressed on the immature form of the N protein, but epitope structures of site IV MAbs 6-9 and 8-1 are created and/or exposed only after maturation of the N protein.

Amino Acid Sequence↗
PubMed2000

The rabies virion-associated 100-kDa polypeptide (VAP100) is a host-derived minor component of the viral envelope.

We investigated a minor polypeptide component of 100-kDa detected in the rabies virion (referred to as VAP100) by using a monoclonal antibody (mAb), #16743, which was shown to recognize the SDS-denatured VAP100 antigen by immunoblot analyses. Although the VAP100 antigen was hardly detectable in the cell by usual immunoblot methods with this mAb, we could detect the antigen by a luminescent immunoblot method as well as by immunoprecipitation from the metabolically radiolabeled cell lysates and virions. Fluorescent antibody (FA) staining with mAb #16743 detected the uniformly distributed antigen on the formalin-fixed normal BHK-21 cells, while slight accumulation of the antigen was also seen in the Golgi area when the cells were permeabilized by treatment with Triton X-100 after fixation. Rabies virus infection induced alteration of the behavior of VAP100 to show a spotted distribution pattern in virus-infected cells. Double FA staining with mAb #16743 and rabbit antibody against the rabies virus envelope antigen demonstrated colocalized distribution of the viral envelope antigens and VAP100 in the cell. From these results, we think that VAP100 is a membrane-associated component of the cell, and its colocalized distribution with the viral envelope antigens in the cell implicates an intimate association of the VAP100 with viral envelope protein(s) and a reflection of possible involvement in the efficient incorporation of VAP100 into the virion.

Animals↗
PubMed2000

Standardized ejection fraction as a parameter of overall ventricular pump function.

To evaluate the pump function of the ventricle, a parameter which (i) incorporates systolic and diastolic function and (ii) separates the heart from preload and afterload is needed. This study utilized ejection fraction (EF), calculated from the end-systolic (ES) and end-diastolic (ED) pressure-volume relationship (PVR) using an arbitrary set of loading conditions. Ten isolated canine hearts with a balloon placed inside the left ventricle were used to determine the ESPVR and EDPVR. An end-diastolic volume (EDV) at a pressure of 15 mmHg and an end-systolic volume (ESV) at 70 mmHg were obtained from the EDPVR and ESPVR, respectively. EF was calculated as (EDV-ESV)/EDV. With low-dose (8 microg/min) and high-dose (40 microg/min) dobutamine infusion, the EF increased from 0.25+/-0.16 to 0.33+/-0.13 and 0.57+/-0.08 (p<0.01), respectively, in conjunction with increases in end-systolic elastance from 3.11+/-0.83 to 3.48+/-1.08 and 5.38+/-1.91 mmHg/ml (p<0.01). It was thus concluded that because the estimation of EF separates the heart from preload and afterload, this method may facilitate comparing overall pump function of hearts beating under different loading conditions.

Animals↗
PubMed2000

Gait characteristics of patients after proximal femoral replacement for malignant bone tumour.

We analysed the gait characteristics of 15 patients with prosthetic reconstruction of the proximal femur after resection of a malignant bone tumour using stride analysis and measurement of oxygen consumption. Compared with normal volunteers their gait was slower, with less cadence and reduced stride length. The mean net energy cost of free walking was 141% of normal. The degree of asymmetry of the single-limb support time correlated with the free-walking velocity and the net energy cost. If they used a single cane the subjects walked with less cadence, longer stride length, and prolonged single-limb support times. The net energy cost of walking and asymmetry of the single-limb support time had a negative correlation with the strength of the hip abductor muscles. Their walking performance was better than that of six subjects who had hip disarticulation.

Adolescent↗
PubMed2000

Mutagenicity of bovine lactoferrin in reverse mutation test.

The mutagenicity of bovine lactoferrin, which is an iron-binding glycoprotein in milk, was evaluated by the Ames mutagenicity test. A total of 5 test strains including 3 base-pair substitution-type strains, Salmonella typhimurium TA100, TA1535 and Escherichia coli WP2uvrA, and 2 frameshift-type strains, TA98 and TA1537, were used in the test. The test was performed by both the direct method and the metabolic activation method with preincubation applied in each instance. The concentration range of the test solution was 0.16 to 5.00 mg/100 microliters (plate). Results of the test revealed that the number of revertant colonies at each concentration of the test solutions was less than 1.4 times that of the control group. In the test system used, bovine lactoferrin did not exhibit mutagenicity.

Animals↗
PubMed2000

A new human malignant peripheral nerve sheath tumour-cell line, HS-sch-2, harbouring p53 point mutation.

Only a few human malignant peripheral nerve sheath tumour (MPNST)-cell lines have been reported, and their characteristics have not been fully established. In this study, we established a new human cell line, HS-Sch-2, from an MPNST of the ordinary type which arose in a 54-year-old woman without von Recklinghausen's disease. This cell line was characterized by chromosome analysis, immunohistochemistry, ultrastructural examination, and direct sequencing of the p53 gene. The HS-Sch-2 cells have grown for more than 48 months in vitro, and exhibited hypotriploid karyotypes with complex chromosome abnormalities lacking a specific pattern. Histological features of the heterotranplanted nude mouse tumours were essentially the same as those of the original MPNST, with positive reactions for S-100 protein and neuron-specific enolase but not for epithelial membrane antigen, fibronectin or CD34. Ultrastructural examination in vivo revealed intricate interdigitation of long cytoplasmic processes and basal lamina-like structures. In addition, direct sequencing of the p53 gene detected a point mutation from CGT to CAT at codon 273 in exon 8. This HS-Sch-2 cell line, which exhibits distinctive morphological characteristics of MPNST and a p53 point mutation, will be useful for biological and pathological investigations of MPNST.

Animals↗
PubMed2000

Hemodialysis-related subacromial lesion: diagnostic imaging and minimally invasive treatment.

BACKGROUND: In an attempt to refine the diagnosis and surgical treatment system for spontaneous shoulder pain in supine position (SPSP) in long-term hemodialysis (LTHD) patients we reviewed shoulders of patients during a 4-year period. PATIENTS AND METHODS: Clinical findings, imaging findings and operative results (average duration of follow-up was 39 months) on both shoulders of 110 patients (Shigei Hospital, Okayama, Japan) were analyzed to identify the cause of SPSP in order to accurately differentiate it from other shoulder pains, and to select the proper treatment method. RESULTS: As the hemodialysis period lengthened, increase in the incidence and severity of bone and soft tissue changes (radiolucency, joint destruction, increase of thickness of subacromial bursa and rotator cuff, and synovial proliferation) in the shoulder induced by dialysis-related amyloidosis were noted in roentgenography, ultrasonography and magnetic resonance imaging. SPSP occurred in the LTHD patients. An increase in subacromial bursa and rotator cuff thickness correlated with SPSP. A decrease in subacromial space correlated with change in position and SPSP. An increase of subacromial pressure was judged to be the cause of SPSP, and subacromial decompression by coraco-acromial ligament release was effective for relieving SPSP. When massive synovial proliferation or invasion of granulation tissues was observed in the glenohumeral joint, arthroscopic debridement was necessary. CONCLUSION: SPSP is a sensitive indicator of a distinct subacromial lesion frequently found in LTHD patients. These lesions can be treated by minimally invasive endoscopic coraco-acromial ligament release, resulting in marked pain relief.

Arthroscopy↗
PubMed1999

Nucleocapsid formation and/or subsequent conformational change of rabies virus nucleoprotein (N) is a prerequisite step for acquiring the phosphatase-sensitive epitope of monoclonal antibody 5-2-26.

We investigated the antigenic maturation of rabies virus N protein, for which we used some conformational epitope-specific monoclonal antibodies (MAbs) and an MAb (5-2-26) against a phosphorylation-dependent linear epitope. Infected cells were lysed with a deoxycholate-free lysis buffer and separated by ultracentrifugation into the soluble top and the nucleocapsid fractions. None of the study MAbs recognized N proteins in the top fraction, whereas nucleocapsid-associated N proteins were recognized by all of the MAbs. Immunoprecipitation with polyclonal anti-N antibodies coprecipitated the P proteins from the top fraction, indicating that soluble N proteins are mostly associated with the P protein. The N proteins dissociated from both the N-P complex and nucleocapsids were recognized by none of the study MAbs, whereas the MAb 5-2-6 recognized the SDS-denatured N proteins of the nucleocapsid but not of the top fraction. In addition, the phosphorylation-deficient mutant N proteins were shown to be similarly accumulated as the wild-type N proteins into the viral inclusion bodies, defined as the virus-specific structures composed of viral nucleocapsids, that are produced in the cytoplasm of the infected cells. Based on these results, we believe that newly synthesized N proteins are not immediately phosphorylated at serine-389 (a common phosphorylation site) but are first associated with the P protein. After being used for encapsidation of the viral RNA, the N proteins undergo conformational changes, whereby epitopes for the conformation-specific MAbs are formed and become phosphorylated at serine-389.

Animals↗
PubMed1999

Double-layered membrane vesicles released from mammalian cells infected with Sendai virus expressing the matrix protein of vesicular stomatitis virus.

The matrix (M) protein of vesicular stomatitis virus (VSV) was reported to form vesicles on the cell surface and subsequently to be released into the cultured medium when expressed from cDNA by virus vectors. To further investigate VSV M activity, we generated a recombinant Sendai virus (SeV) expressing the VSV M protein (SeV-M(VSV)). When cells were infected with SeV-M(VSV), VSV M was found abundantly in the culture medium. Electron microscopy demonstrated the budding of two-membraned vesicles (>/= 0.8 microm in diameter) from the infected cells. The outer membrane of the vesicle was derived from the plasma membrane and the inner one possibly derived from the membrane of an intracellular vesicle. Immuno-gold labeling showed that VSV M was exclusively located in a double-layered region. The released membranes were divided into three parts: the VSV M vesicles with SeV F and HN glycoproteins, SeV particles, and vesicles associated with the cytosolic components. The last abundantly contained phosphorylated SeV matrix (M) protein, which is not released in a usual SeV infection. Furthermore the VSV M protein expressed without using a virus vector was efficiently released into the culture medium. These results suggest that the VSV M protein has a budding activity per se and that SeV proteins are passively involved in the release of VSV M.

Animals↗
PubMed1999

Imaging assessment of the responses of osteosarcoma patients to preoperative chemotherapy: angiography compared with thallium-201 scintigraphy.

BACKGROUND: Assessment of the responses of osteosarcoma patients to preoperative chemotherapy is of clinical importance. The purpose of this study was to estimate the accuracy of angiography and thallium-201 scintigraphy, compared with histology, in assessing the responses of patients with osteosarcoma to preoperative chemotherapy. METHODS: Nineteen patients with osteosarcoma who were diagnosed between 1992 and 1997 were studied. The findings of angiography and thallium-201 scintigraphy before and after preoperative chemotherapy were compared with the percentage of necrosis of tumor cells and the response grade as determined histologically. Quantitative analysis of the isotopic uptake by thallium-201 scintigraphy before and after chemotherapy, defined as the alteration ratio, was correlated with the percentage of tumor necrosis. RESULTS: Angiography yielded a sensitivity of 88%, a specificity of 73%, and a predictive accuracy of positive test of 70%, whereas thallium-201 scintigraphy achieved 88%, 100%, and 100%, respectively. Both angiographic and scintigraphic assessment showed a significant correlation with response grade as determined histologically (P < 0.05 and P < 0.0003, respectively). The alteration ratio of thallium-201 scintigraphy showed a strong, highly significant correlation with the percentage of tumor necrosis (P < 0.0001). CONCLUSIONS: A change in the tumor uptake of thallium-201 scintigraphy after preoperative chemotherapy can predict the tumor necrosis in osteosarcoma precisely. Thallium scintigraphy is a noninvasive technique and seems to be more useful than angiography in assessing the response of osteosarcoma to preoperative chemotherapy.

Adolescent↗
PubMed1999

Newly identified Chinese hamster ovary cell mutants defective in peroxisome assembly represent complementation group A of human peroxisome biogenesis disorders and one novel group in mammals.

We isolated peroxisome biogenesis-defective mutants from rat PEX2-transformed Chinese hamster ovary (CHO) cells, using the 9-(1'-pyrene)nonanol/ultraviolet method. A total of 18 mutant cell clones showing cytosolic localization of catalase were isolated. By complementation group (CG) analysis by means of PEX cDNA transfection and cell fusion, cell mutants, ZP124 and ZP126, were found to belong to two novel CGs of CHO mutants. Mutants, ZP135 and ZP167, were also classified to the same CG as ZP124. Further cell fusion analysis using 12 CGs fibroblasts from patients with peroxisome deficiency disorders such as Zellweger syndrome revealed that ZP124 belonged to human CG-A, the same group as CG-VIII in the United States. ZP126 could not be classified to any of human and CHO CGs. These mutants also showed typical peroxisome assembly-defective phenotypes such as severe loss of catalase latency and impaired biogenesis of peroxisomal enzymes. Collectively, ZP124 represents CG-A, and ZP126 is in a newly identified CG distinct from the 14 mammalian CGs previously characterized.

Animals↗
PubMed1999

Isolation and characterization of novel peroxisome biogenesis-defective Chinese hamster ovary cell mutants using green fluorescent protein.

We developed an improved method for isolation of peroxisome biogenesis-defective somatic animal cell mutants, using a combination of green fluorescent protein (GFP) expression and the 9-(1'-pyrene)nonanol/ultraviolet (P9OH/UV) selection method. We used TKaG1 and TKaG2 cells, the wild-type Chinese hamster ovary (CHO) cells, CHO-K1, that had been stably transfected with cDNAs each encoding rat Pex2p as well as GFP tagged at the C-terminus with peroxisome targeting signal type 1 (PTS1) or N-terminally PTS2-tagged GFP. P9OH/UV-resistant cell colonies were examined for intracellular location of GFP on unfixed cells, by fluorescence microscopy. Seven each of the mutant cell clones isolated from TKaG1 and TKaG2 showed cytosolic GFP-PTS1 and PTS2-GFP, respectively, indicating the defect in peroxisome assembly. By transfection of PEX2, PEX5, PEX6, and PEX12 cDNAs and cell fusion analysis between the CHO cell mutants, five different complementation groups (CGs) were identified. Two mutant clones, ZPG207 and ZPG208, belonged to novel CGs. Further CG analysis using fibroblasts from patients with peroxisome biogenesis disorders, including rhizomelic chondrodysplasia punctata (RCDP), revealed that ZPG208 belonged to none of human CGs. ZPG207 was classified into the same CG as RCDP. Taken together, ZPG208 is in a newly identified, the 12th, CG in peroxisome-deficient CHO mutants reported to date and represents a novel mammalian CG.

ATPases Associated with Diverse Cellular Activitie↗
PubMed1999

Studies on the structures and antigenic properties of rabies virus glycoprotein analogues produced in yeast cells.

We investigated two forms (designated as yGI and yGII) of rabies virus glycoprotein (G) analogues produced in the G cDNA-transfected yeast cells. Molecular weights of yGI and yGII were estimated as 66 and 56 kDa, respectively, according to their relative mobility in SDS-PAGE. Although being produced in large amounts, yGI was present mostly in insoluble forms and hardly extractable with non-ionic detergents. The yGI reacted with polyclonal anti-G antibodies, but did not react with our conformational epitope-specific anti-G monoclonal antibodies (G-MAbs). No protective immunity was induced by yGI in guinea pigs nor in mice. On the other hand, yGII was Triton-soluble, but was only small in amount (at most 1% of total G proteins) and was shown to lack the cytoplasmic domain. The yGII, however, reacted with the G-MAbs and induced protective immunity in guinea pigs as well. When the G-cDNA was expressed in animal cells in culture, a single form (about 66 kDa) of G protein was produced, which displayed similar behaviors as seen in its reactivity with the MAbs and intracellular distribution as seen in the virus-infected cells. These results suggest that most G protein molecules were not processed normally in yeast cells, resulting in abnormal folding and multimer formation, while only a small fraction were occasionally folded normally to have conformational epitopes but were mostly deprived of the C-terminal portion.

Animals↗
PubMed1999

Relationship between magnitude of resection, complication, and prosthetic survival after prosthetic knee reconstructions for distal femoral tumors.

BACKGROUND AND OBJECTIVES: Limb-sparing surgery has become the preferred surgical treatment of malignant bone tumors. The objective of this study was to evaluate factors that influence the morbidity and outcome of prosthetic knee replacement after resection of malignant tumors of the distal femur. METHODS: Eighty-two patients who had a resection of malignant tumor of the distal femur and implantation of a segmental knee prosthesis (minimum follow-up, 2 years) were retrospectively reviewed. RESULTS: Twenty-nine patients (35%) underwent 32 prosthetic revisions, 6 from perioperative wound complications, 13 from aseptic loosening, and 13 from other complications. The 3-, 5-, and 10-year Kaplan-Meier prosthetic survival rates were 82%, 71%, and 50%, respectively. On univariate analysis, patients who had more than 40% resection of the distal femur (P = 0.010) and those who had all their vasti muscles resected (P = 0.011) had significantly worse prosthetic survivals than the others. On multivariate analysis, resection of more than 40% of the distal femur was a significant negative prognostic factor for prosthetic survival (P = 0.017). Aseptic loosening was the primary cause of late prosthetic failure. Differences in the magnitude of resection influenced prosthetic survivorship more than prosthetic design. CONCLUSIONS: In the distal femoral endoprosthetic replacement, higher short- and long-term complications were found after extensive resections. Aseptic loosening was the primary cause of prosthetic failure.

Adolescent↗
PubMed1999

Electrophoretic and serologic characterization of 56 kDa antigen (M56) with autologous serum derived from a chondrosarcoma patient: a shared antigen of immunoresponses in cancer and autoimmune diseases.

We investigated whether antibodies specific to autologous cancer cells are produced in the peripheral blood of patients with chondrosarcoma. There have been few reports on the investigation of the immune responses, such as autologous antibody production, to chondrosarcoma. Here, tumor-associated antigens were separated by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis and detected by immunoenzymatic amplification. A 56 kDa molecule (M56) was detected in the serum from patients' peripheral blood. M56 is ubiquitously expressed in various kinds of tissue-derived cells. However, the molecule seemed to be retained mostly in the cytosolic compartment of lymphoid cells, while it was expressed on the cell surface of nonlymphoid cancer cells. Furthermore, the antibodies reactive to the 56 kDa molecule were frequently observed in sera derived from patients with other cancers and autoimmune diseases as compared to the sera from healthy control donors, suggesting that M56 is a common target molecule of immune responses in patients with various cancers and autoimmune diseases.

Animals↗
PubMed1999

Expression of CD44 variants in osteosarcoma.

The standard form of CD44 (CD44H) is a transmembranous glycoprotein, widely distributed on a variety of human lymphoid cells, epithelial cells and tumours. CD44 has many variant forms, which are generated by alternative splicing. In recent years, CD44 has been reported to be related to the degree of tumour differentiation, tumour cell invasion, and metastasis. We investigated 44 tumour specimens in 39 patients with osteosarcoma immunochemically to analyse the expression of CD44 standard (CD44H) and variant exon-encoded gene products (CD44v3, v4, v5, v6, v7, v9, and v10). Furthermore, the relationship between CD44 expression and the clinical outcome of patients with osteosarcoma was analysed. Membrane accentuation and exclusive cytoplasmic reactivity were analysed as separate staining patterns. Tumour cells and some multinucleated giant cells were markedly stained. CD44H, v3, v4, v5, v6, v7, v9, and v10 were expressed in 85%, 49%, 54%, 59%, 46%, 5%, 28%, and 10% of the specimens respectively. The cumulative 5-year metastasis-free survival was 58% in CD44v6-negative cases and 24% in CD44v6-positive cases (P=0.046). However, the cumulative 5-year metastasis-free survival was not significantly different between cases positive and negative for other variants of CD44. Multivariate analysis (Cox proportional-hazard model) with CD44v6 expression (positive or negative), chemotherapy (intensive or non-intensive), tumour site (proximal or distal), and age (at least 30 years or less than 30 years) showed that expression of CD44v6 and chemotherapy were important prognostic factors in patients with osteosarcoma. Overexpression of CD44 isoforms containing variant v6 is correlated with poor prognosis in patients with osteosarcoma.

Adolescent↗
PubMed1999

Prolonged preservation of the blood-perfused canine heart with glycolysis-promoting solution.

BACKGROUND: Prolonged ischemia and inadequate myocardial preservation remain significant perioperative risk factors in cardiac transplantation. Long-term preservation techniques that have been effective in small rodent hearts have not been as effective in larger animal models or in clinical studies. We developed a cardioplegia solution formulated to promote high-energy phosphate production from glycolysis and determined its efficacy in a blood perfused canine heart model subjected to 24 hours of ischemia. METHODS: Hearts harvested from adult dogs (n = 6 per group) were flushed with a histidine-buffered cardioplegia solution containing glucose or University of Wisconsin solution. The hearts were maintained at 4 degrees C for 24 hours then reperfused with autologous blood. After reperfusion, left ventricular pressures were measured with an intracavitary balloon at varying balloon volumes and compared with control nonischemic hearts. Predicted stroke volume and ejection fraction were calculated at an end-systolic pressure of 70 mm Hg and end-diastolic pressure of 15 mm Hg. RESULTS: Developed pressure was better preserved in the hearts that received histidine-buffered solution (93+/-9 versus 38+/-7 mm Hg, p<0.05), along with a higher end-diastolic volume at 15 mm Hg (31+/-3 versus 22+/-2 mL histidine-buffered versus University of Wisconsin solutions, respectively, p<0.05). Stroke volume and ejection fraction were also higher in the histidine group (17+/-2.5 versus 2.3+/-1.2 mL and 50%+/-3.5% versus 9% +/-4.5%, respectively) in the presence of dobutamine. CONCLUSIONS: The highly buffered glycolysis-promoting cardioplegia solution provided effective preservation of the blood perfused canine heart with superior recovery of pump performance after 24 hours of hypothermic ischemia compared with University of Wisconsin solution in this model.

Animals↗