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Biomedical subjects

M Kawakami

Publications and source records attributed to M Kawakami.

At least 127 records · Page 7Linked to original sources
PubMed1999

Biochemical characterization and expression analysis of neural thrombospondin-1-like proteins NELL1 and NELL2.

Two closely related genes coding for NELL proteins (NELL1 and NELL2) have been cloned by the yeast two-hybrid screening of a rat brain cDNA library with the regulatory domain of protein kinase C betaI (PKCbetaI) as bait. The rat NELL proteins show about 55% identity with each other and contain several protein motifs assigned to a secretion signal peptide, an NH(2)-terminal thrombospondin-1 (TSP-1)-like module, five von Willebrand factor C domains, and six epidermal growth factor-like domains; the NELL proteins share many protein motifs with TSP-1. The NELL proteins expressed in COS-7 cells are homotrimeric glycoproteins and possess heparin-binding activity. Furthermore, while NELL1 and NELL2 show distinct subcellular localization in cytoplasm, they both are partially secreted into the culture medium of COS-7 cells. Although the NELL1 mRNA is faintly expressed in adult neural cells, the NELL2 mRNA is expressed abundantly, particularly in the pyramidal cells of rat hippocampus, showing neuronal high plasticity. During mouse embryogenesis, expression of the NELL2 mRNA is initiated 7-11 days postcoitum, simultaneously with neural plate formation. These results strongly suggest that the NELL2 protein, similar to but not identical with TSP-1, is involved in the growth and differentiation of neural cells. Additionally, the NELL1 and NELL2 mRNAs were found to be expressed abundantly in Burkitt's lymphoma Raji cells and colorectal adenocarcinoma SW480 cells, respectively. Thus, it is likely that the NELL proteins also participate in the growth, differentiation, and oncogenesis of cancer cell lines.

Amino Acid Sequence↗
PubMed1999

Pharmacokinetic analysis and antitumor efficacy of MKT-077, a novel antitumor agent.

MKT-077 (1-ethyl-2-[[3-ethyl-5-(3-methylbenzothiazolin-2-yliden)]-4- oxothiazolidin-2-ylidenemethyl] pyridinium chloride), a novel rhodacyanine dye in phase I/II clinical trials, may provide a new approach to cancer therapy based on the accumulation in the mitochondria of the cells of certain carcinomas, for example, those of the colon, breast and pancreas. To support the development of MKT-077 for clinical application as an intravenous (i.v.) therapy, we investigated the metabolic fate of [14C]MKT-077 in BDF1 mice as well as the distribution of MKT-077 in experimental LS174T tumor-bearing mice using a high-performance liquid chromatography (HPLC) method. The plasma levels of 14C after i.v. administration of [14C]MKT-077 declined in a triphasic manner. In the first distribution phase, the levels of 14C decreased with a T1/2 of approximately 5 min. In the second and terminal phase, the T1/2 of 14C was 2.8-4.6 h and 16.2 h, respectively. Cmax (1 min after injection) increased from 0.3 to 1.5 microg/ml linearly, but less than proportionately between the doses. The AUC(0-infinity) at 0.3, 1 and 3 mg/kg were 0.030 +/- 0.002, 0.60 +/- 0.12 and 1.73 +/- 0.25 microg x h/ml, respectively. Plasma clearance was approximately 1.8 l/h per kg (at doses of 1 and 3 mg/kg). The steady state volume of distribution (6.8 and 25.1 l/kg) indicated that MKT-077 distributed as a lipid-soluble molecule. The mean residence time (MRT) was 4.1 (at a dose of 1 mg/kg) and 14.1 h (at a dose of 3 mg/kg). In the first rapid phase (5 min after dosing), 14C radioactivity was detected in most of the tissues and organs, most strongly in the kidney cortex, and not in the central nervous system and testes. In the terminal phase (24 h after dosing), 14C contents increased in the intestinal tract, and in the kidney and liver were nearly to the background level. After i.v. bolus administration at a dose of 3 mg/kg of [14C]MKT-077, the predominant route of elimination of the radioactivity was via the feces, and recoveries of total radioactivity in urine and feces corresponded to 33.5% and 61.1%, respectively. More than 60% was recovered within 24 h and 95% within 1 week. MKT-077 was primarily excreted in unmetabolized form with five unidentified metabolites found in the urine and plasma. Intact MKT-077 was retained in the tumor tissue longer than in plasma and kidney in LS174T tumor-bearing mice receiving MKT-077 at an i.v. therapeutic dose (10 mg/kg). This accumulation decreased very slowly, suggesting that the high membrane potentials of tumor cell mitochondria may help retain the drug in tumors.

Animals↗
PubMed1999

Constitutive expression of the Wilms' tumor gene WT1 inhibits the differentiation of myeloid progenitor cells but promotes their proliferation in response to granulocyte-colony stimulating factor (G-CSF).

Bone marrow (BM) cells that were concentrated for hematopoietic progenitor cells by in vivo treatment with 5-FU were infected with a recombinant retrovirus containing a human full-sized, non-spliced type WT1 (Wilms' tumor gene 1) cDNA and then colony-assayed in the presence of granulocyte-colony stimulating factor (G-CSF). Significantly more colony-forming units granulocyte-monocyte (CFU-GM), colony-forming units granulocyte (CFU-G), and colony-forming units monocyte (CFU-M) colonies were formed in response to G-CSF from the BM cells infected with the WT1-containing retrovirus than from the control BM cells infected with an empty vector. Furthermore, FACS analysis of cell surface differentiation markers showed the inhibition of differentiation by constitutive WT1 expression resulting from the infection with the WT1-containing retrovirus. These results thus showed that the constitutive WT1 expression promoted the proliferation of myeloid progenitor cells but inhibited their differentiation in response to G-CSF, suggesting the alteration of G-CSF signaling pathway. The results also supported our hypothesis that the WT1 gene performs an oncogenic rather than a tumor suppressor gene function in hematopoietic progenitor cells, although the WT1 gene potentially performs both functions. This finding implies an important role of the WT1 gene in leukemogenesis.

Animals↗
PubMed1999

Gene structure of the P100 serine-protease component of the human Ra-reactive factor.

The Ra-reactive factor (RaRF) is a complement dependent anti-microbial factor that reacts with numerous microorganisms such as viruses, bacteria, fungi and protozoa. It is a complex of a mannan-binding lectin (MBL) and the serine protease, P100 (MASPI). P100 activates the C4 component of the complement system and its domain organization is similar to C1r and C1s. In this study, determination was made of the structure of the human P100 gene which was found longer than 67 kbp and to be comprised of 16 exons. Its non-protease region consisted of 10 exons, as in the case of C1r and C1s, and the introns were found present in the boundary separating two CUB domains, an EGF-like domain and two CCP domains and each CUB and CCP domain contained extra internal introns. The serine protease region was comprised of 6 exons in contrast to C1r and C1s, either of which consists of a single exon. The exon-intron structure was found to reflect the evolution of these molecules and P100 to have derived earlier in the stage of evolution than C1r or C1s.

Amino Acid Sequence↗
PubMed1999

Determination of MKT-077, a novel antineoplastic agent, in plasma samples by high-performance liquid chromatography and its application to pharmacokinetics in rats.

A simple high-performance liquid chromatographic method was developed for determination of a novel antineoplastic agent MKT-077 in plasma. MKT-077 was extracted from 50 microl of plasma with acetonitrile containing 1 ml trifluoroacetic acid per liter. Chromatographic separation was achieved within 13.5 min using a reverse-phase Puresil C18 analytical column. A visible detector operated at 490 nm was used. The linearity of the calibration curve was obtained (r2 = 0.99986) over the analytical range of 10-500 ng/ml(-1). The intra- and inter-assay precision was in the range of 0.9-11.1 and 0.3-4.4%, respectively. The intra- and inter-assay bias ranged from -7.3 to 11.1% and from 0.4 to 11.6%, respectively. The utility of this assay was demonstrated after the administration of a single dose of MKT-077 to rats. The plasma elimination half-life of MKT-077 was 1.8-4 h.

Animals↗
PubMed1999

The Wilms' tumor gene WT1 is a good marker for diagnosis of disease progression of myelodysplastic syndromes.

The Wilms' tumor gene, WT1, is a tumor marker for leukemic blast cells. The WT1 expression levels were examined for 57 patients with myelodysplastic syndromes (MDS) (refractory anemia (RA), 35; RA with excess of blasts (RAEB) 14; RAEB in transformation (RAEB-t), six; and MDS with fibrosis, two) and 12 patients with acute myeloid leukemia (AML) evolved from MDS. These levels significantly increased in proportion to the disease progression of MDS from RA to overt AML via RAEB and RAEB-t in both bone marrow (BM) and peripheral blood (PB). WT1 expression levels in PB significantly correlated with the evolution of RAEB or RAEB-t to overt AML within 6 months. Therefore, WT1 expression levels in PB were superior to those in BM for early prediction of the evolution to AML by means of quantitation of the WT1 expression levels. Furthermore, WT1 expression in PB of patients with overt AML evolved from MDS was significantly decreased by effective chemotherapy or allogeneic stem cell transplantation and became undetectable in long-term survivors. These results clearly showed that WT1 expression levels are a tumor marker for preleukemic or leukemic blast cells of MDS and thus reflect the disease progression of MDS. Therefore, monitoring of WT1 expression levels has made continuous assessment of the disease progression of MDS possible, as well as the prediction of the evolution of RAEB or RAEB-t to overt AML within 6 months. The results also showed that quantitation of WT1 expression levels is useful for diagnosis of minimal residual disease of MDS with high sensitivity, thus making it possible to evaluate the efficacy of treatment for MDS.

Adult↗
PubMed1999

Antibody-independent classical complement pathway activation and homologous C3 deposition in xeroderma pigmentosum cell lines.

Of human malignantly transformed cell lines, xeroderma pigmentosum (XP) cell lines were found to be highly susceptible to homologous complement (C): cells were opsonized by C3 fragments on incubation with diluted normal human serum. C3 fragment deposition on XP cells was Ca2+-dependent and occurred on live cells but not UV-irradiated apoptotic cells. (Ca2+ is required for activation of the classical C pathway via C1q and the lactin pathway via mannose binding lectin (MBL), and the surface of apoptotic cells usually activates the alternative C pathway.) In this study we tested which of the pathways participates in XP cell C3 deposition. In seven cell lines that allowed C3 deposition (i), Clq was shown to be essential but MBL played no role in C activation, (ii) Cls but not MASP bound XP cells for activation, (iii) no antibodies recognizing XP cells were required for homologous C3 deposition, and (iv) the alternative pathway barely participated in C3 deposition. Furthermore, the levels of C-regulatory proteins for host cell protection against C, decay-accelerating factor (DAF, CD55) and membrane cofactor protein (MCP, CD46), were found to be relatively low in almost all XP cell lines compared with normal cells. These results indicate that XP cells activate the classical C pathway in an antibody-independent manner through the expression of a molecule which directly attracts C1q in a C-activating form, and that relatively low levels of DAF and MCP on XP cells facilitate effective C3 deposition. The possible relationship between the pathogenesis of XP and our findings is discussed.

Antibodies↗
PubMed1999

Max protein expression is associated with survival of children with lymphoblastic lymphoma.

Overexpression of c-Myc in murine B lineage cells is associated with a polyclonal pre-B cell expansion as well as development of pre-B or B lymphoblastic lymphoma (LL) accompanied by leukemia, which mirrors the clinical features of childhood LL. Of interest, Max overexpression attenuates aberrant growth of B cells triggered by c-Myc. However, the clinical significance of Max in human lymphoid tissue remains to be clarified. In the present studies, we studied the expression of the c-Myc and Max proteins in normal lymph nodes and in childhood LL. In normal lymph nodes, c-Myc protein was expressed by the majority of cells in germinal center and marginal zone, but Max protein was expressed only by some of them. In contrast, c-Myc and Max were absent in mantle zone. Cells positive for c-Myc and Max expression in LL were 70.6 +/- 19.8% and 47.3 +/- 32.4%, respectively, determined by immunohistochemical staining using paraffin blocks from 23 cases of childhood LL. The survival of children with LL showing higher Max expression (> or = 30%) was significantly greater than that of lower expression (< 30%; P = 0.0027 using the Mantel-Cox test). These results suggest that Max protein may affect prognosis of childhood LL.

Basic Helix-Loop-Helix Leucine Zipper Transcriptio↗
PubMed1999

Evaluation of adsorption selectivity dextran sulfate bound cellulose beads for the removal of anti-DNA antibodies.

The purpose of this study was to clarify the basic adsorption selectivity characteristics of dextran sulfate (DS) columns (Selesorb, Kaneka Corporation, Osaka, Japan). Recovery rates of blood chemical components, hormones, coagulation factors, and antinuclear antibodies (anti-SS-A, SS-B, Sm, Scl-70, and RNP antibody) in vitro were assessed by mixing normal volunteers' or patients' sera with DS bound cellulose beads. For tested blood chemical components other than triglyceride and total cholesterol, the recovery rate was not changed significantly by incubation. No significant changes in hormone levels resulted from incubation. Among coagulation factors, the activities of antithrombin III, plasminogen, and factors V, VIII, IX, XI, and XII were significantly reduced by incubation. Among antinuclear antibodies tested, anti-SS-A and anti-RNP were absorbed to some extent, but not anti-SS-B, Sm, or Scl-70 antibodies. Taking into account these characteristics, apheresis therapy using a DS column should be performed.

Adsorption↗
PubMed1999

Changes in CD4+ T lymphocyte subsets in circulating blood and synovial fluid following filtration leukocytapheresis therapy in patients with rheumatoid arthritis.

The purpose of this study is to determine the changes in CD4+ T lymphocyte subsets in the circulating blood and synovial fluid following filtration leukocytapheresis (LCP) therapy for patients with rheumatoid arthritis (RA). A Cellsorba column packed with polyester fibers was used for the removal of circulating leukocytes. For patients with RA, filtration LCP or sham procedures were performed 3 times with 1 week intervals between procedures. T lymphocyte surface markers in the peripheral blood and synovial fluid were measured by flow cytometry. The proportions of activated CD4+ T cells (CD4+DR+, CD4+CD25+, and CD4+CD71+) and CD4+CD29+ T cells increased significantly in the peripheral blood, but the counts of these cells were significantly reduced in the synovial fluid after 2 treatment sessions in the LCP group. No significant changes were observed in the proportion of these cells in the control group. Our findings suggest that filtration LCP may cause a redistribution of activated T cells from affected joints into the circulating blood.

Arthritis, Rheumatoid↗
PubMed1999

Pharmacological characterization of PABSA, an orally active and highly potent endothelin-receptor antagonist.

The pharmacological characterization of a nonpeptide endothelin (ET)-receptor antagonist, PABSA [(R)-(--)-2-(benzo[1,3]dioxol-5-yl)-N-(4-isopropyl-phenylsulfon yl)-2-(6-methyl-2-propylpyridin-3-yloxy)-acetamide hydrochloride] was studied. PABSA competitively inhibited the binding of [125I]-ET-1 to A7r5 cells expressing ET(A) receptors and of [125I]-ET-3 to COS cells expressing porcine ET(B) receptors with Ki values of 0.11 and 25 nM, respectively. PABSA inhibited ET(A) receptor-mediated and ET(B) receptor-mediated vasocontraction and ET(B) receptor-mediated vasorelaxation in isolated rabbit vessels with K(b) values of 0.46, 94, and 26 nM, respectively. The antagonist potency of PABSA for ET(A) receptor-mediated vasocontraction was 63- and 87-fold more potent than those of BQ-123 and bosentan, respectively, and was similar to those of TAK-044 and SB209670. Oral administration of PABSA (1-10 mg/kg) caused dose-dependent inhibition of the pressor response to exogenous ET- 1 (0.1 nmol/kg) in conscious normotensive rats. PABSA (10-100 mg/kg, p.o.) reduced blood pressure in deoxycorticosterone acetate (DOCA)-salt hypertensive rats, spontaneously hypertensive rats (SHRs), and stroke-prone spontaneously hypertensive rats (SHRSPs). The hypotensive effect of PABSA was sustained for > or =24 h in these rats. These results suggest that PABSA is a highly potent ET(A)-receptor antagonist with weak ET(B)-receptor antagonist activity. Because PABSA has a long duration of action in vivo, this antagonist should be useful in the therapy of ET-related disease.

Administration, Oral↗
PubMed1999

Expression of the Wilms' tumor gene WT1 in solid tumors and its involvement in tumor cell growth.

To determine the role of the Wilms' tumor gene WT1 in tumorigenesis of solid tumors, expression of the WT1 gene was examined in 34 solid tumor cell lines (four gastric cancer cell lines, five colon cancer cell lines, 15 lung cancer cell lines, four breast cancer cell lines, one germ cell tumor cell line, two ovarian cancer cell lines, one uterine cancer cell line, one thyroid cancer cell line, and one hepatocellular carcinoma cell line) by means of quantitative reverse transcriptase-polymerase chain reaction. WT1 gene expression was detected in three of the four gastric cancer cell lines, all of the five colon cancer cell lines, 12 of the 15 lung cancer cell lines, two of the four breast cancer cell lines, the germ cell tumor cell line, the two ovarian cancer cell lines, the uterine cancer cell line, the thyroid cancer cell line, and the hepatocellular carcinoma cell line. Therefore, of the 34 solid tumor cell lines examined, 28 (82%) expressed WT1. Three cell lines expressing WT1 (gastric cancer cell line AZ-521, lung cancer cell line OS3, and ovarian cancer cell line TYK-nu) were further analyzed for mutations and/or deletions in the WT1 gene by means of single-strand conformation polymorphism analysis. However, no mutations or deletions were detected in the region of the WT1 gene ranging from the 3' end of exon 1 to exon 10 (the WT1 gene consists of 10 exons) in these three cell lines. Furthermore, when AZ-521, OS3, and TYK-nu cells were treated with WT1 antisense oligomers, the growth of these cells was significantly inhibited in association with a reduction in WT1 protein levels. Furthermore, constitute expression of the transfected WT1 gene in cancer cells inhibited the antisense effect of WT1 antisense oligomer on cell growth. These results indicated that the WT1 gene plays an essential role in the growth of solid tumors and performs an oncogenic rather than a tumor-suppressor gene function.

DNA-Binding Proteins↗
PubMed1999

Involvement of H-cadherin (CDH13) on 16q in the region of frequent deletion in ovarian cancer.

Ovarian cancer is one of the leading causes of mortality unique to women. Deletions within chromosome 6q are the most frequent events in high-grade invasive epithelial ovarian cancer (IEOC). While previous reports seem to indicate that there is loss of 16q sequences in IOEC, only a very small number of markers (single marker in two different reports) were used. In order to more precisely define the regions of deletions on 16q, we first analyzed LOH with 13 polymorphic markers on 16q in 10 benign, 3 low-grade and 21 high-grade invasive ovarian cancer samples. There was no loss with any of the markers with the benign ovarian samples and loss of one marker in one of three low-grade tumors. In contrast, 14 of 21 (67%) high-grade invasive ovarian tumors showed loss of one or more markers. Detailed deletion mapping revealed three distinct commonly deleted regions on this chromosomal arm: 10/21 (48%) of high-grade tumors showed loss at 16q23.1-23.2 (D16S518, D16S3049 and D16S3029). The second region of loss at 16q23.3-16q24.1 (D16S3144, D16S504, HSD17B2 and D16S507) was observed in 11/21 (52%) of the tumors. The highest frequency of loss was seen at 16q24.2-16q24.3 (D16S422, D16S402 and D16S520) in 12/21 (57%) of tumors. The genomic map of CDH13 indicates that the marker D16S422 that was lost in 5 of these 12 tumors is part of this gene. Three of these 5 tumors showed very low levels of CDH13 expression. Three tumors with LOH of other markers in this region also showed lower levels of CDH13 expression. Analysis of the methylation status of CDH13 in tumors with low levels of expression with methylation-specific PCR revealed that four of six (67%) tumors had methylation of one of the CDH13 alleles. These results suggest that a combination of hyper-methylation and deletion cause the inactivation of CDH13 in ovarian tumors.

Cadherins↗
PubMed1999

Axial symptoms and cervical alignments after cervical anterior spinal fusion for patients with cervical myelopathy.

This retrospective clinical study was designed to examine the relation between cervical alignment and axial symptoms developing after cervical anterior spinal fusion. Sixty patients with myelopathy treated with cervical anterior spinal fusion were reviewed. For radiographic evaluation, lordosis, enlargement of the fused segments and neural foramen, radiographic union, and degeneration of adjacent segment were reviewed before or after surgery or both. Twenty-three patients had axial symptoms. Only local kyphosis and narrowing of the neural foramen at the fused segment were recognized more often in patients with axial symptoms than in those without such symptoms. No less than 2 mm and < or = 5 mm in enlargement of the anterior disc space immediately after surgery resulted in maintenance of cervical lordosis. These findings suggest that > or = 2 mm and < or = 5 mm in enlargement of anterior vertebral body height during operation results in prevention of axial symptoms.

Adult↗
PubMed1999

[Two surgical cases of intrathoracic lipoma].

We experienced two cases of intrathoracic lipoma arising from the chest wall. The first case was a 55-year-old male, and another case was a 70-year-old female. In the second case, the computed tomography was highly suggestive of a pleural tumor with almost same density as the subcutaneous adipose tissue. This findings may be diagnostic in assessing intrathoracic chest wall type lipoma. Both cases had thoracotomy to remove the tumors. The tumors were easily resectable, though their capsules were partially unclear. We resected the tumors completely together with pleura, periosteum and intercostal muscle. The diagnosis of intramuscular lipoma was confirmed postoperatively by histopathologic examination in each case. When the capsule of lipoma is not clear, combined resection of the tumor and chest wall should be considered. There have been no recurrence of the tumors in our cases.

Aged↗
PubMed1999

[New classification of diabetes by ADA and WHO with special reference to pathogenesis].

New classification of diabetes mellitus was proposed by ADA international committee on the diagnosis and classification and by WHO consultation committee. The classification based on the pathogeneses of diabetes and also on the stages of diseases. Since our knowledge on causes of diabetic syndrome is not yet enough to classify diabetes in genetic bases, the classification had been carried out some sort of mixture of pathogeneses, genetic findings and clinical features of hyperglycemia. Stages of diseases based on the natural course of diabetes mellitus in reference to shortage of insulin effects and degree of chronic hyperglycemia. These two components of classification composed two dimensions of the field that expresses all the pathogeneses and metabolic conditions of all diabetic patients.

Diabetes Mellitus↗
PubMed1999

[Role of advanced glycation end products (AGE) in the development of diabetic microangiopathies and the beneficial effects of AGE inhibitors].

Glucose or other reducing sugars in plasma or tissue fluids react with proteins by a nonenzymatic, post-translational process called glycation or Maillard reaction which leads to the formation of advanced glycation end products (AGEs). AGEs are implicated in the multiorgan complications of diabetes because of their multipotent bioactivities through their receptors as well as the functional and structural derangement of the AGE-bearing proteins. AGE inhibitors such as aminoguanidine have been shown to have beneficial effects on the progression of a full range of diabetic complications in animal models. Furthermore, a recent clinical study showed that aminoguanidine therapy resulted in significant improvements in several key measurements; it reduced urinary protein, LDL-cholesterol and triglycerides as well as blood pressure. A novel class of AGE-antagonists, AGE breakers have been also developed and waiting for the clinical evaluation.

Diabetic Angiopathies↗