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Biomedical subjects

T Kanbe

Publications and source records attributed to T Kanbe.

At least 73 records · Page 4Linked to original sources

The amino acid sequence of neocarzinostatin apoprotein deduced from the base sequence of the gene.

A segment of the neocarzinostatin apoprotein gene corresponding to T30 to A91 of the protein was amplified using a polymerase chain reaction (PCR) with total DNA from Streptomyces carzinostaticus subsp. neocarzinostaticus E-793 (ATCC 15944) as the template and with 5'- and 3'-primers synthesized in consideration of the codon usage of streptomyces. The PCR product was cloned, sequenced and confirmed to direct an amino acid sequence reasonably well matching that reported. Using the PCR product as a probe, we cloned a DNA segment (2580 bp) spanning an open reading frame (ORF) for preapoprotein (leader peptide plus apoprotein) and its upstream and downstream flanking regions. The amino acid sequence deduced from the base sequence of the DNA clearly identified those amino acid residues which had remained inconsistent among different research groups. The base sequence homology with other apoprotein genes of related antibiotics was analyzed and was found to be limited within the structural gene.

Amino Acid Sequence↗

Distinct effects of clinically used anthracycline antibiotics on ras oncogene-expressed cells.

Doxorubicin, pirarubicin, and FAD-104, but not aclarubicin or MX 2, flattened the morphology of NIH3T3 cells that had been transformed by human H-ras and K-ras. The effect appeared on almost all cells, as early as 2 d following exposure to the antibiotics at concentrations inhibiting cell growth by 50% or more. The morphological alteration accompanied other normal cell phenotypes, such as the restoration of actin stress fibers, anchorage dependence of cell growth and an increase in nucleoside diphosphate (NDP) kinase activity. NIH3T3 cells transformed by src and other tumor cell lines responded less prominently, if at all.

3T3 Cells↗

[A case of B-chronic lymphocytic leukemia/prolymphocytic leukemia (CLL/PL)].

A 59-year-old man was admitted to our hospital on May 17, 1991 because of dizziness and a sense of abdominal fullness. Physical examination on admission showed splenomegaly without hepatomegaly or lymphadenopathy, and blood examination revealed normocytic anemia, thrombocytopenia and marked leukocytosis of 16,800/microliters with 87% lymphoid cells. Prolymphocytoid cells formed 28% of the lymphoid cells. Bone marrow aspiration revealed massive infiltration of lymphoid cells. Surface marker analysis showed that the lymphoid cells were positive for anti-HLA-DR, CD 5, CD19, CD20, CD21, SmIgM and SmIgD. The patient was diagnosed as having B-CLL/PL, according to the classification advocated by Melo in 1986, and initially treated with vindesine + prednisolone + pirarubicin (VP-THP). However, the prolymphocyte count increased, so we changed to VP-THP + cyclophosphamide (VEP-THP), and remission was obtained. CLL/PL is a rare disease in Japan but we obtained a good response to chemotherapy.

Antineoplastic Combined Chemotherapy Protocols↗

Detection of intracellular forms of secretory aspartic proteinase in Candida albicans.

The extracellular proteinase (EPR) of Candida albicans was induced in a medium containing bovine serum albumin as sole nitrogen source. There were two intracellular forms in cells induced to produce EPR, a 43 kDa protein (EPR) and a 45 kDa protein (cross-reacting material of EPR; CRM-EPR); these were detected by immunoblotting using anti-EPR antiserum. The 43 kDa protein (EPR) may be the same as the extracellular form judging by molecular mass, and the 45 kDa protein (CRM-EPR) may be a precursor form of EPR. Many dense granules were observed by electron microscopy near the plasma membrane of the mother cells in EPR-producing cells. Both the 43 and 45 kDa proteins were recovered in a membrane fraction and were solubilized by Triton X-100. When the membrane fraction was further fractionated by sucrose density gradient centrifugation, the 43 and 45 kDa proteins were differentially fractionated. This suggests that they were located in different membrane-bound structures and is consistent with an assumption that the 45 kDa protein is a precursor for EPR.

Aspartic Acid Endopeptidases↗

Evidence that Candida albicans binds via a unique adhesion system on phagocytic cells in the marginal zone of the mouse spleen.

We recently demonstrated by using an ex vivo adhesion assay that Candida albicans yeast cells exhibit a unique binding affinity for the marginal zone of the spleen. This binding event provides a working model for studying mechanisms of organ dissemination of the fungus from the blood. By using the ex vivo assays reported here, we showed by bright-field and electron microscopic techniques that mouse spleen marginal zone cells capable of ingesting India ink particles are also involved in yeast cell attachment. During splenic clearance of yeast cells from the circulation in vivo, C. albicans is also associated exclusively with marginal zone cells capable of ingesting India ink. The ability to ingest the ink particles is not necessarily related to yeast cell adherence, because the fungal cells did not bind to phagocytic cells in the splenic red pulp. In fact, the marginal zone phagocytic cells appear to have a unique binding system, because yeast cells also did not bind to phagocytes in other tissues, such as the thymus and peritoneum, or to seven different myeloid cell lines. In addition, antibodies to a number of well-characterized murine adhesion molecules, such as leukocyte integrins, LECAM-1, and CD44, had no effect on binding. On the basis of these results, we propose that splenic marginal zone phagocytes express a novel adhesion system that involves either a unique adhesion molecule or previously described adhesion molecules with unique binding activities.

Adhesiveness↗

[Chronic myelomonocytic leukemia with good response to low-dose etoposide].

58-year-old man was admitted with cervical tumor and leukocytosis. Physical examination indicated splenomegaly and cervical abscess. Laboratory data showed WBC 55,000/microliters, Hb 7.9g/dl, and PLT 4.5 x 10(4)/microliters. After cure of the abscess, WBC counts were still high with 1,500-2,000/microliters monocytes, and anemia and thrombocytopenia persisted. Bone marrow aspiration showed myeloid hyperplasia and trilineage myelodysplasia. The Ph1 chromosome could not be detected. The case was diagnosed as chronic myelomonocytic leukemia and treated with oral etoposide (25mg/day). After 2 weeks, the dose was increased to 50mg, and then modified according to the blood counts. WBC counts are presently being maintained between 7,000 and 12,000/microliters, and RBC and PLT counts have gradually become normal. Splenomegaly almost disappeared and dysplastic change in bone marrow improved somewhat. At nine months following the start of chemotherapy with etoposide, remission is maintained by treatment with 25mg of etoposide on alternate days. This case suggests that low-dose etoposide is useful for treating CMMoL.

Administration, Oral↗

Improvements and important considerations of an ex vivo assay to study Candida albicans-splenic tissue interactions.

Improvements were made on an ex vivo assay to study adherence properties of Candida albicans to host internal organs. The assay is applicable to understanding mechanisms of C. albicans dissemination following a fungemia. Binding patterns of yeast forms to splenic tissue are intriguing and we found the following modifications to be especially relevant. Mice serving as spleen donors for the assay should be injected with 0.1 ml i.v. of a 10% concentration of luconyl blue, 5 min before killing. After collecting splenic sections on a glass slide, 100 microliters of a 1-2 x 10(8)/ml suspension of stationary yeast cells should be applied to the sections. The assay does not require a carbonate buffering system or serum supplements. Attachment of yeasts to host tissue occurs best if the interaction is allowed to proceed without agitation by rotation. Assessment of binding is facilitated by staining the slides with crystal violet and computer image analysis can be used for quantification of binding.

Animals↗

Evidence for expression of the C3d receptor of Candida albicans in vitro and in vivo obtained by immunofluorescence and immunoelectron microscopy.

The complement conversion product C3d binds to a receptor on the cell surface of Candida albicans. While the function of this receptor is still uncertain, we investigated whether it is expressed during a murine infection. Rabbit antiserum raised against purified receptor was used in conjunction with immunofluorescence microscopy and immunocolloidal gold electron microscopy to examine kidney tissue and peritoneal lavages from infected mice for receptor expression by C. albicans in vivo. Specificity of the antiserum was indicated by reactivity with purified receptor (55 to 60 kDa) and with a protein of similar molecular mass from whole hyphal extracts in Western blots (immunoblots). In vitro analysis by immunofluorescence microscopy showed that the antiserum reacted with both yeast and pseudohyphal forms of the organism, but reactivity was strongest with pseudohyphae. Immunocolloidal gold electron microscopy of fungal cells from peritoneal lavages revealed intense staining of mother cells of germinative forms, germ tubes, and pseudohyphae. Staining of the mother cells was heaviest at the innermost layers of the cell wall but only scant on the cell surface. In contrast, staining was observed throughout the cell walls of germ tubes and pseudohyphae. In kidney, expression of the C3d receptor was found primarily on the cell walls of hyphae and pseudohyphae, although some staining was observed in the cytoplasm. These data support that the C3d receptor of C. albicans is expressed in vivo.

Animals↗

Synthesis and antiulcer activity of optical isomers of 2-(4-chlorobenzoylamino)-3-[2(1H)-quinolinon-4-yl]propionic acid (rebamipide).

The enantiomers of 2-(4-chlorobenzoylamino)-3-[2(1H)-quinolinon-4-yl] propionic acid [(+/-)-1, rebamipide, OPC-12759], a new antiulcer agent that enhances mucosal resistance, were synthesized from optically active alpha-amino acid derivatives of 2(1H)-quinolinone. The key intermediates, alpha-amino acid derivatives, were prepared by asymmetric synthesis and optical resolution. The (+)-1 was about 1.7 times as potent as the (-)-isomer in antiulcer activity against ethanol-induced gastric ulcers.

Alanine↗

[A case of chest wall rhabdomyosarcoma].

A 56-year-old man was admitted to our hospital with right chest pain. Chest X-ray, CT scan and MRI revealed a chest wall tumor and enlarged mediastinal lymph nodes. Percutaneous lung biopsy was performed, and the pathological diagnosis of pleomorphic rhabdomyosarcoma was obtained. The only significant abnormal laboratory finding was elevation of serum NSE (24.5 ng/ml). Although chemotherapy (VAC-ADM) and radiation therapy were performed, the patient died about 7 months after admission. To our knowledge, only 17 cases of chest wall rhabdomyosarcoma have been reported in Japan.

Antineoplastic Combined Chemotherapy Protocols↗

[Liver disorders in patients with Mycoplasma pneumoniae pneumonia].

Liver disorders were reviewed in 76 patients with mycoplasma pneumoniae pneumonia (MP) admitted to Kamo Hospital from 1979 to 1989. Liver dysfunction was detected in 16 patients (21%). Liver disorder were detected in 10% of MP from 1979 to 1980, but in 20% from 1984 to 1985, and in 36% from 1988 to 1989. By multiple regression analysis CRP was found to be the most significant factor among other factors such as prevalence, age, sex, history of smoking, history of drinking, and days after onset of fever.

Adult↗

Hepatotoxicity of diethyldithiocarbamate in rats.

Hepatotoxicity of diethyldithiocarbamate (DDC) was investigated in rats. Plasma aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities were markedly elevated 24 hr after subcutaneous administration of DDC and histologically, the liver showed submassive necrosis. A sustained inhibition in the liver of Cu,Zn-superoxide dismutase (Cu-SOD) activity was observed following DDC treatment. DDC produced a significant loss in liver reduced glutathione (GSH) level after 1 hr, but the nadir was observed later than that of Cu-SOD. Catalase activity decreased gradually from 7 hr. Thiobarbituric acid reactive substances (TBARS) in the liver were significantly increased from 15 hr. Hepatic haemodynamics were scarcely changed up to 15 hr. Desferrioxamine (a chelator of iron) and piperonyl butoxide (an inhibitor of cytochrome P-450) prevented DDC-induced increases of both ALT and TBARS, but GSH did not, DDC hepatotoxicity was not changed by phenobarbital induction. Thus, we have shown that subcutaneous dose of DDC caused hepatotoxicity in rats. Although the exact sequence of its hepatotoxic factors is unproven, it seems likely that lipid peroxidation through the dysfunction of antioxidant defence factors and a toxic metabolite contribute to the formation of this liver injury.

Alanine Transaminase↗

The SLP1 gene of Saccharomyces cerevisiae is essential for vacuolar morphogenesis and function.

The SLP1 gene, which is involved in the expression of vacuolar functions in the yeast Saccharomyces cerevisiae (K. Kitamoto, K. Yoshizawa, Y. Ohsumi, and Y. Anraku, J. Bacteriol. 170:2687-2691, 1988), has been cloned from a yeast genomic library by complementation of the slp1-1 mutation. The isolated plasmid has a 7.8-kilobase BamHI-BamHI fragment that is sufficient to complement several characteristic phenotypes of the slp1-1 mutation. The fragment was integrated at the chromosomal SLP1 locus, indicating that it contains an authentic SLP1 gene. By DNA sequencing of the SLP1 gene, an open reading frame of 2,073 base pairs coding for a polypeptide of 691 amino acid residues (Mr, 79,270) was found. Gene disruption of the chromosomal SLP1 did not cause a lethal event. Vacuolar proteins in the delta slp1 mutant are not processed to vacuolar forms but remain in Golgi-modified forms. Carboxypeptidase Y in the delta slp1 mutant is localized mainly to the outsides of the cells. delta slp1 mutant cells have no prominent vacuolar structures but contain numerous vesicles in the cytoplasm, as seen by electron microscopy. Genetic and molecular biological analyses revealed that SLP1 is identical to VPS33, which is required for vacuolar protein sorting as reported by Robinson et al. (J. S. Robinson, D. J. Klionsky, L. M. Banta, and S. D. Emr, Mol. Cell. Biol. 8:4936-4948, 1988). These results indicate that the SLP1 (VPS33) gene is involved in the sorting of vacuolar proteins from the Golgi apparatus and their targeting to the vacuole and that it is required for the morphogenesis of vacuoles and subsequent expression of vacuolar functions.

Amino Acid Sequence↗

The transition of cells of the fission yeast beta-tubulin mutant nda3-311 as seen by freeze-substitution electron microscopy. Requirement of functional tubulin for spindle pole body duplication.

A previous fluorescence light-microscopic study showed that the fission yeast cold-sensitive beta-tubulin mutant nda3-311 was arrested with rod-like condensed chromosomes in a mitotic state at the restrictive temperature. Upon transfer to the permissive temperature, a spindle was formed and the nucleus was divided. In the present study, we employed freeze-substitution electron microscopy to examine the ultrastructure of arrested and released nda3-311 cells. In arrested cells, a single, displaced nucleus was seen with a single spindle pole body. Therefore, spindle pole body duplication seemed to require functional beta-tubulin. The nuclear membrane was highly deformed with a leaf-like profile in cross-section, possibly due to an interaction with the rod-like, condensed chromosomes. Upon transfer to the permissive temperature, the spindle pole duplicated and the daughter spindle pole bodies rapidly migrated to the opposite ends of the nucleus, accompanied by the formation of the mitotic spindle. Elongation of the nuclear envelope occurred with concomitant spindle extension, as in a wild-type mitosis. The deformed nuclear membrane became smooth and described a convex curve. The numerous vacuoles that are seen in the arrested cells decreased in number and increased in size. Septation was completed, leaving the two divided nuclei in one half of the cell. Hexagonally arranged microtubules, apparently forming the mitotic spindle, were observed in a cross-section of a cell after return to the permissive conditions.

Freezing↗

Synthesis and antiulcer activity of 4-substituted 8-[(2-benzimidazolyl)sulfinylmethyl]-1,2,3,4-tetrahydroquinoli nes and related compounds.

A series of 4-substituted 8-[(2-benzimidazolyl)sulfinylmethyl]-1,2,3,4-tetrahydroquinolin es was synthesized and examined for their (H+ + K+)adenosine triphosphatase (ATPase)-inhibitory and antisecretory activities against histamine-induced gastric acid secretion in rats. Many compounds tested were potent inhibitors of (H+ + K+)ATPase. Most compounds showed antisecretory activity. The antiulcer activity against water-immersion stress-induced gastric ulcer, aspirin-induced gastric ulcer and gastric necrosis induced by hydrochloric acid also were tested in the rat. Some of these compounds, in particular, 4-(N-allyl-N-methylamino)-1-ethyl-8-[(5-fluoro-6-methoxy-2-benzimidazoly l) sulfinylmethyl]-1-ethyl-1,2,3,4-tetrahydroquinoline (XVIIx) were found to have potent activity. The structure-activity relationships are discussed.

Adenosine Triphosphatases↗

High frequency variation of colony morphology and chromosome reorganization in the pathogenic yeast Candida albicans.

A clinical isolate of the pathogenic yeast Candida albicans varied in its colony morphology from smooth (o-smooth) to semi-rough type (SRT) and concomitantly lost its virulence for mice. In terms of DNA content, the smooth parent was near triploid when Saccharomyces cerevisiae strains of known ploidy were used as references. The SRT variant showed several features characteristic of polyploidy. From the SRT variant, revertant-like smooth (r-smooth) variants with recovered virulence were derived at a frequency of 5 x 10(-3). The results of pulsed-field gel electrophoresis on chromosomal DNA showed changes in patterns of chromosome-sized DNA bands in the SRT variant as well as in r-smooth variants, which correlated with these variations. Correlations between colony morphology, state of ploidy and virulence of this asporogenous yeast are considered.

Candida albicans↗