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Biomedical subjects

Y Hashimoto

Publications and source records attributed to Y Hashimoto.

At least 1,153 records · Page 64Linked to original sources
PubMed1989

Improved method for amino acid analysis of stained collagen bands from polyacrylamide gels using high-performance liquid chromatography.

A procedure is described for the determination of the amino acid composition of stained collagen bands separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis using high-performance liquid chromatography and post-labelling with o-phthalaldehyde. Six alpha-chain bands from collagens I, III and V were analysed directly after gel electrophoresis, Coomassie blue staining and destaining. It was shown that good accuracy and reproducibility can be attained using this method, which makes it possible to determine simultaneously nineteen amino acids: Asx, Thr, Ser, Glx, Pro, Cys, Gly, Ala, Val, Met, Ile, Leu, Tyr, Phe, His, Lys and Arg together with 4-Hyp and Hyl, specific amino acids found in collagen.

Amino Acid Sequence↗
PubMed1989

A tumor-associated antigen in the scirrhous gastric carcinoma cell line MK-01 defined by monoclonal antibody S202.

A monoclonal antibody (MAb) S202, with an IgG1 isotype, that reacted strongly with the scirrhous gastric carcinoma cell line MK-01 was established. MAb S202 reacted with the colonic cancer cell line, SW1116, and the pancreatic cancer cell line, PK-1, when tested by indirect immunofluorescence. The S202 reactive antigen was expressed in the majority of acetone-fixed fresh frozen cancer tissues. Eighty to 100 per cent of the paraffin-embedded sections of stomach, colon and pancreatic adenocarcinoma were positive for the S202 antigen, with diffuse cytoplasmic staining, whereas esophageal and breast cancers demonstrated markedly less immunostaining. Supplemented serum-free medium collected from 7 day old tumor cell cultures were assayed for the presence of antibody-defined antigens. Antigens detected by MAb S202 were released by the cell lines SW1116 and PK-1. The binding of MAb S202 to the colonic adenocarcinoma sections was reduced after treatment with sodium periodate which suggests that respective antigenic determinants are of carbohydrate nature without sialic acid residues.

Adenocarcinoma, Scirrhous↗
PubMed1989

Anti-idiotype antibody reactive with a target molecule for mouse lymphokine-activated killer cells.

A rat monoclonal antibody (MoAb), termed KBA, against mouse lymphokine-activated killer (LAK) cells recognizes a LAK cell surface molecule termed LAA responsible for the binding between LAK and target cells. In order to identify a target molecule of LAK cells, we prepared anti-KBA idiotype antibodies (anti-KBA-Id) from rabbit anti-KBA sera. Immunoglobulins were separated by ammonium sulfate precipitation followed by sequential affinity column chromatographies using Affi-gel coupled with rat MoAbs other than KBA and KBA-coupled gel. An immunoglobulin(s) in a KBA-gel-bound fraction showed the selective reactivity to KBA, comprising anti-KBA-Id character. This anti-KBA-Id inhibited the binding of KBA to LAK. Moreover, it bound with a portion of mouse leukemia cells sensitive to LAK cells, but not with normal mouse cells, and inhibited the binding of LAK cells to a target leukemia. These findings indicate that the anti-KBA-Id contain anti-Id which possess a three-dimensional structure that mimics a mirror image of the antigen (LAA)-combining site in KBA or the structure of LAA. The antigen reactive with anti-KBA-Id was characterized as a glycoprotein.

Animals↗
PubMed1989

Reversal of enzymic phenotype of thymidine metabolism in induced differentiation of HL-60 cells.

Exposure of HL-60 cells to 12-O-tetradecanoylphorbol-13-acetate (TPA) resulted in specific alterations in thymidine (TdR) metabolism. Within 12 h after treatment with 1.62 nM TPA, the reciprocal alteration in the activities of opposing pathways of TdR metabolism observed during normal culture cell growth was reversed. In TPA-treated cells, the activities of anabolic enzymes, TdR kinase (TK; EC 2.7.1.21) and thymidylate synthase (TS; EC 2.1.1.45), declined to 15% and 18% of those of untreated cells by 96 h. Incorporation of 3H-TdR and 3H-deoxyuridine also decreased in parallel with decline in enzyme activities. In contrast, the activities of catabolic enzymes, TdR phosphorylase (TP; EC 2.4.2.4) and dihydrothymine dehydrogenase (DHT DH; EC 1.3.1.2), increased to 399% and 318% by 96 h. Immunotitration of DHT DH with monoclonal antibody showed that the rise in activity in the differentiated cells was due to the increase in protein amount. Kinetic properties of the enzymes were not altered during differentiation. These metabolic alterations were accompanied by an accumulation of the cells in G1 at the expense of S-phase. Present data indicate that induced differentiation of HL-60 cells results in a reversal of enzymic phenotype of TdR metabolism due to a consequence of decreased proliferation and suggest that emergence of TdR metabolic imbalance may serve as early markers of differentiation of these cells.

Cell Cycle↗
PubMed1989

Zinc ion enhances the blocking potency of synthetic analogs of spider toxin (JSTX) on the glutamate receptor.

The effect of synthetic spider toxin analogs containing aza-crown in combination with zinc and copper was studied on the lobster neuromuscular synapse. The suppressive action of N4-type spider toxin analog (N4) on excitatory postsynaptic potentials (EPSPs) was markedly enhanced in the presence of 10(-5)-10(-4) M Zn2+. Cu2+ (10(-4) M) also had a potentiating effect on the suppression of EPSPs by N4 but to a lesser degree than Zn2+. While N6-type spider toxin analog (N6) suppressed EPSPs more effectively than N4, additional suppression was not pronounced in the presence of Zn2+ or Cu2+. The results suggest that chelatable divalent metal ions play an important role in the blocking mechanism of glutamate receptor by the spider toxin.

Action Potentials↗
PubMed1989

Retinobenzoic acids. 2. Structure-activity relationships of chalcone-4-carboxylic acids and flavone-4'-carboxylic acids.

The structure-activity relationships of (E)-chalcone-4-carboxylic acids, which are retinoidal benzoic acids represented by R-Ph-X-Ph-COOH (4, X = -COCH = CH-), are discussed on the basis of differentiation-inducing activity on human promyelocytic leukemia cells HL-60. The activity was increased by the substitution of a bulky alkyl group(s) (R), and among such compounds, (E)-4-[3-(3,5-di-tert-butylphenyl)-3-oxo-1-propenyl]benzoic acid (Ch55) and (E)-4-[3-oxo-3-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1 -propenyl]benzoic acid (Ch80) are several times more active than retinoic acid. Though the stable conformer of chalcone derivatives is linear (s-cis form), the conformationally restricted analogue 4-(6,7,8,9-tetrahydro-6,6,9,9-tetramethyl-4H-4-oxonaphtho[2,3-b]py ran-2-yl)benzoic acid (Fv80) is more active than Ch80. While the effect of introduction of an oxygen atom varied, 4-[1-hydroxy-3-oxo-3-(5,6,7,8-tetrahydro-3-hydroxy-5,5,8,8-tetramethyl-2 - naphthalenyl)-1-propenyl]benzoic acid (Re80), regarded as a derivative of Ch80 with two additional hydroxyl groups, has very strong activity.

Carboxylic Acids↗
PubMed1989

Retinobenzoic acids. 3. Structure-activity relationships of retinoidal azobenzene-4-carboxylic acids and stilbene-4-carboxylic acids.

Alkyl-substituted azobenzene-4-carboxylic acids are potent differentiation inducers of human promyelocytic leukemia cell line HL-60 to mature granulocytes. Their structure-activity relationships are very similar to those of other retinoidal benzoic acids which are generally represented by 4 and named retinobenzoic acids. The structure-activity relationships of azobenzenecarboxylic acids can also be applied to the known retinoid TTNPB (3). Thus, (E)-4-[2-(3,4-diisopropylphenyl)-1-propenyl]benzoic acid (St30 (28] and (E)-4-[2-(3-tert-butylphenyl)ethenyl]benzoic acid (St40 (29], the acyclic alkyl analogues of TTNPB, are nearly as active as retinoic acid. Among the oxidatively derived compounds (Az90, Ep series and Ox series) of azobenzene- or stilbenecarboxylic acids, Az90 (71) and Ep80 (61) have strong activities. However, all the bishydroxylated derivatives of TTNPB are inactive, while a diketo analogue Ox580 (69) has only weak potency. The activities of conformationally restricted compounds of TTNPB offer some information on the stereochemistry of the active form of these retinoidal compounds.

Benzoates↗
PubMed1989

Hepatocarcinogenic heterocyclic aromatic amines that induce cytochrome P-448 isozymes, mainly cytochrome P-448H (P-450IA2), responsible for mutagenic activation of the carcinogens in rat liver.

Male F344 rats were treated with hepatocarcinogenic heterocyclic aromatic amines such as amino acid- and protein-pyrolysate components (Trp P-1, Trp P-2, Glu P-1, Glu P-2, A alpha C, MeA alpha C, IQ and MeIQx) and changes in microsomal cytochrome P-450 isozymes in the livers were examined by means of immuno-Western blotting using anti-rat cytochrome P-450 monoclonal antibodies. The results suggested that all chemicals tested induce cytochrome P-448 isozymes, particularly cytochrome P-448H (P-450IA2), which efficiently mediate mutagenic activation of the carcinogens. This was substantiated by the enzymatic analyses with the substrates showing different characters to rat cytochrome P-450 isozyme-mediated mutagenesis.

Amines↗
PubMed1989

Further studies on polymorphic expression of GM1 and GD1a in mouse liver. The presence of a third allele on the Ggm-1 locus.

C57BL/10(H-2b) and A/J(H-2a) mice have different phenotypes as to the age dependent changes in the expression of GM1 and GD1a in the liver, i.e., the contents of these two components in total gangliosides in C57BL/10 and A/J mice were 1 and 20% at 4 wk and had decreased to almost 0 and 10%, respectively, at 8 wk [Y. Nakamura et al. (1988) J. Biochem. 103, 393-396]. To analyze the mode of inheritance of these phenotypes, C57BL/10 mice were mated with A/J or A/Wy(H-2a) mice. The phenotype of A/J or A/Wy mice was found to be inherited by the F1 hybrids, indicating that the phenotype of A mice is dominant over that of C57BL/10 mice. The (C57BL/10 x A/Wy)F1 hybrids were then backcrossed with one of the parent strains, C57BL/10 mice. The phenotype of A/Wy mice was demonstrated to be inherited by 23 out of 35 mice and all of the 23 had the H-2a haplotype. These results suggest that the two phenotypes of C57BL/10 and A mice are controlled by a pair of allelic genes on a locus which is closely linked to the H-2 complex on chromosome 17. Unexpectedly, B10. A(H-2a) mice were found to have significantly higher amounts of GM1 and GD1a than both C57BL/10 and A mice, although they were a H-2 congenic strain carrying the H-2 genes transferred from A/Wy mice and should have the same genes on loci next to H-2 as one of the two strains. It remains to be determined how B10.A mice have higher amounts of GM1 and GD1a than C57BL/10 and A/Wy mice.

Alleles↗
PubMed1989

A beta-Galactosidase from Radish (Raphanus sativus L.) Seeds.

A basic beta-galactosidase (beta-Galase) has been purified 281-fold from imbibed radish (Raphanus sativus L.) seeds by conventional purification procedures. The purified enzyme is an electrophoretically homogeneous protein consisting of a single polypeptide with an apparent molecular mass of 45 kilodaltons and pl values of 8.6 to 8.8. The enzyme was maximally active at pH 4.0 on p-nitrophenyl beta-d-galactoside and beta-1,3-linked galactobiose. The enzyme activity was inhibited strongly by Hg(2+) and 4-chloromercuribenzoate. d-Galactono-(1-->4)-lactone and d-galactal acted as potent competitive inhibitors. Using galactooligosaccharides differing in the types of linkage as the substrates, it was demonstrated that radish seed beta-Galase specifically split off beta-1,3- and beta-1,6-linked d-galactosyl residues from the nonreducing ends, and their rates of hydrolysis increased with increasing chain lengths. Radish seed and leaf arabino-3,6-galactan-proteins were resistant to the beta-galase alone but could be partially degraded by the enzyme after the treatment with a fungal alpha-l-arabinofuranosidase leaving some oligosaccharides consisting of d-galactose, uronic acid, l-arabinose, and other minor sugar components besides d-galactose as the main product.

Journal Article↗
PubMed1989

A murine monoclonal antibody that recognizes an extracellular domain of the human c-erbB-2 protooncogene product.

A murine IgM monoclonal antibody, designated SV2-61, was generated against human c-erbB-2 gene-transfected NIH-3T3 (SV11) cells. SV2-61 defined a 185-kDa molecule present on the surface of SV11 cells, another line of c-erbB-2 gene-transfected NIH-3T3 (A4-15) cells, and MKN-7 human gastric cancer cell line carrying an amplified human c-erbB-2 gene. The SV2-61-defined antigen was found to show protein kinase activity in vitro. The SV2-61 was reactive with human c-erbB-2 gene-transfected NIH-3T3 cell lines but not with transfectants carrying c-erbB-2 gene mutants which lack a coding region for the extracellular domain. It was reactive with a portion of human epithelial cell lines but not with native NIH-3T3, TGF-alpha-coding gene-, activated c-raf gene- or Ha-ras gene-transfected NIH-3T3 cells, or non-epithelial human cells. These results indicate that the SV2-61 is an antibody which recognizes an extracellular domain of the c-erbB-2 gene product, 185-kDa protein.

Animals↗
PubMed1989

Strain differences in susceptibility to 2-acetylaminofluorene and phenobarbital promotion of hepatocarcinogenesis: immunohistochemical analysis of cytochrome P-450 isozyme induction by 2-acetylaminofluorene and phenobarbital.

Strain differences in the expression of cytochrome P-450 isoenzymes (P-450s) during enhancement of hepatocarcinogenesis by 2-acetylaminofluorene (2-AAF) and phenobarbital (PB) were investigated immunohistochemically using monoclonal antibodies against phenobarbital (PB) (APF3) or 3-methylcholanthrene (3-MC) (APH8) inducible P-450s. LEW, SD, WBN, F344, SHR, NAR, Wistar and ODS rats were studied, the first five strains proving to be less susceptible to 2-AAF induction of APH8 while responding strongly to the promoting influence of this chemical, as reported previously. The other three strains, NAR, Wistar and ODS, demonstrated greater inducibility, this correlating with an observed resistance to promotion by 2-AAF. PB administration was not associated with any strain difference in APF3 cytochrome P-450 inducibility except in the ODS rat, in which its effects were minimal. The results provide direct evidence that differential expression of cytochrome P-450 species plays a major role in determining responsiveness to hepatocarcinogenesis-promoting agents such as 2-AAF.

2-Acetylaminofluorene↗
PubMed1989

Changes in the quantity and activity of cytochrome P-450 isozymes in primary cultured rat hepatocytes.

Hepatocytes from male Sprague-Dawley rats pretreated with a cytochrome P-450 inducer, 3-methoxy-4-aminoazobenzene (3-MeO-AAB), 3-methylcholanthrene (MC) or phenobarbital (PB), were cultured in vitro, and changes in the quantity and activity of microsomal cytochrome P-450 isozymes in the cells were determined by means of immunochemical methods and a bacterial mutation test, respectively. The results of enzyme-linked immunosorbent assay using monoclonal antibodies against rat P-450 isozymes revealed that the amount of cytochrome P-450d induced by 3-MeO-AAB or MC declined rapidly during culture and fell to 10 to 15% of the initial value after 24 h. A similar tendency was observed with PB-induced cytochrome P-450b/e. By contrast, cytochrome P-450c in MC-induced hepatocytes declined more slowly than cytochrome P-450d and remained at 45 to 60% of the initial value after 24 h. Similar quantitative changes of the individual cytochrome P-450 isozymes in culture were also observed by immunoblotting using the anti-cytochrome P-450 monoclonal antibodies. Changes in the activities of individual cytochrome P-450 isozymes in hepatocytes by culture were in accordance with the quantitative changes of the cytochromes, as determined by a mutation test using Salmonella typhimurium TA 98 and carcinogenic aromatic amines. These results indicate that microsomal cytochrome P-450c in primary cultured rat hepatocytes is more stable in culture, in terms of both quantity and activity, than cytochrome P-450d and P-450b/e.

Animals↗
PubMed1989

In vitro targeting and cytotoxicity of adriamycin in liposomes bearing monoclonal antibody against rat or human gp125 cell proliferation-associated antigen.

Chemoimmunoliposomes (CIL) were prepared by entrapping adriamycin in monoclonal antibody (mAb)-coated liposomes and examined for their binding capacity and cytotoxicity to relevant target tumor cells. Sonicated unilamellar liposomes were coated with B3 and HBJ127 mouse, mAbs, which recognize a rat and a homologous human cell proliferation-associated surface antigen, gp125, respectively, and then adriamycin was entrapped in the liposomes by means of transmembrane Na+/K+ gradients using valinomycin. These CIL selectively bound with relevant target tumor cells bearing the corresponding gp125 antigen, such as BC47 rat bladder cancer, FTL-13 rat thymic lymphoma, T24 human bladder cancer and Molt-4 human leukemia cells, although the binding capacities of the CIL to bladder cancer cells were relatively larger than those to lymphoma cells in both rat and human systems. This difference in the target cell binding was found to be attributable to the amount of gp125 antigen expressed on each target tumor cell, as determined by a Scatchard plot analysis. In accordance with the target cell binding capacities of CIL preparations, the CIL displayed much higher cytotoxic activity to bladder cancers than to lymphomas in both rat and human systems. In conjuction with our previous finding that gp125 antigen is expressed on tumor cells but not on resting normal cells, these findings indicate that CIL composed of anti-gp125 mAb will be useful for tumor therapy and that the antitumor efficacy is dependent upon the extent of the antigen expression on target tumor cells.

Animals↗