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Biomedical subjects

Y Hashimoto

Publications and source records attributed to Y Hashimoto.

At least 1,315 records · Page 73Linked to original sources
PubMed1987

Transforming activity of human c-Ha-ras-1 proto-oncogene generated by the binding of 2-amino-6-methyl-dipyrido[1,2-a:3',2'-d]imidazole and 4-nitroquinoline N-oxide: direct evidence of cellular transformation by chemically modified DNA.

An activity that transforms NIH 3T3 cells was generated by the in vitro modification of plasmids containing the human c-Ha-ras-1 proto-oncogene with the synthesized ultimate carcinogen, 2-acetoxyamino-6-methyldipyrido[1,2-a:3',2'-d]-imidazole (N-OAc-Glu-P-1). DNAs isolated from the transformed cells were analyzed by restriction fragment length polymorphism (RFLP) assay using the restriction enzyme Msp I. Of fourteen transformants studied, six contained a mutation in the region of the CCGG sequence of the eleventh and the twelfth codons, in which GG corresponds to the first two nucleotides of the twelfth codon. Transforming activity was also generated by the chemical modification of the plasmids with 4-acetoxyaminoquinoline N-oxide (N-OAc-4AQO). The results clearly indicate that formation of DNA adducts with N-OAc-Glu-P-1 or N-OAc-4AQO causes the induction of transformation of mammalian cells.

4-Nitroquinoline-1-oxide↗
PubMed1987

Immunochemically detected nuclear envelope-associated cytochrome P-450 component(s) in rat hepatocyte culture lines.

The presence and location of cytochrome P-450 in Donryu rat hepatocyte culture lines, Ac2F cells and 3 other cell lines were assessed by indirect immunofluorescence examination using anti-cytochrome P-450 monoclonal antibodies. Ac2F cells and other hepatocyte cell lines were selectively stained at their nuclear envelope, but not the cytoplasm, with a monoclonal antibody selective to a high-spin form of cytochrome P-448 (P-448H), although this monoclonal antibody stained primary cultured normal rat hepatocytes at both cellular components and did not stain hepatoma cells of 2 transplantation lines. The results of unscheduled DNA synthesis assay with Ac2F cells using several carcinogenic aromatic amines (4-aminoazobenzene derivatives and amino acid pyrolysis products) suggested that this nuclear envelope-associated cytochrome P-450 activates a restricted portion of these aromatic amines, i.e., a tryptophan pyrolysis component and a glutamic acid pyrolysis component. These results indicate that rat hepatocyte culture lines lack (or contain a reduced amount of) the cytoplasmic cytochrome P-450 but maintain a characteristic type of cytochrome P-450, probably a kind of cytochrome P-448H in their nuclear envelope, and this may be involved in oxidative metabolism of a restricted portion of aromatic amines.

Animals↗
PubMed1987

Characteristics of mouse thymocyte-derived, interleukin 2-activated killer cells and their precursors.

Culture of hydrocortisone (HC)-resistant C57BL/6 mouse thymocytes with recombinant human interleukin 2 (IL 2) allowed the proliferation of the thymocytes and resulted in the generation of lymphokine-activated killer (LAK) cells cytotoxic to a variety of tumor cells. The cytotoxic activity of the LAK cells was greatly reduced by treatment with anti-Thy 1.2 or anti-Lyt 2.2 monoclonal antibody and complement but not with anti-asialoGM1 antibody plus complement. Fractionation of IL 2-stimulated thymocytes by means of Percoll density gradient centrifugation revealed that both cytotoxic activity and binding capacity to target cells were greater in the cells with lower density and larger size than in the cells with higher density and smaller size. These IL 2-activated thymocytes expressed higher levels of both Thy 1, Lyt 2 and lymphokine-activated cell-associated (LAA) antigens than unstimulated thymocytes, as indicated by a flow cytometric analysis. The frequency of LAK precursor cells was found to be 7.5 times greater in the HC-resistant thymocyte population than in total thymocytes, as determined by means of a limiting dilution method. The LAK precursor cells in HC-resistant thymocytes appeared to be Lyt1- (or dull Lyt 1+), L3T4-, Lyt 2-, asialoGM1-T cells, because elimination of bright Lyt 1+, Lyt 2+ or L3T4+ T cells from HC-resistant thymocytes had no effect on the generation of LAK cells. These results indicate that LAK cells from mouse thymocytes are Lyt 2+ T cells which are inducible from HC-resistant Lyt 2- thymocytes.

Animals↗
PubMed1987

Altered expression of immunohistochemically detected cytochrome P-450 component(s) in nitrosamine-induced rat urinary bladder lesion.

Male ACI/N rats were treated with N-butyl-N-(4-hydroxybutyl)nitrosamine (BBN) in the drinking water, and in conjunction with histological examination, the changes of the expressed cytochrome P-450 components in the urothelium and other tissues (liver, kidney, esophagus, intestines) were examined by means of immunohistochemistry. Frozen tissue sections were prepared and immunostained with anti-rat cytochrome P-450 monoclonal antibodies and an avidin-biotin-peroxidase complex. Monoclonal antibodies used were APH-3 and APH-8 raised against a high-spin form of cytochrome P-448, APL-1 and APL-2 against a low-spin form of cytochrome P-448, and APF-3 against cytochrome P-450. BBN-induced qualitative and quantitative changes of cytochrome P-450 components recognized by these monoclonal antibodies were not observed in tissues other than the bladder. Untreated rat bladder epithelium was not stained with any of these 5 monoclonal antibodies. The treatment with BBN for more than 3 weeks, however, resulted in the expression of cytochrome P-450 component(s) recognized by APH-8 antibody. This cytochrome P-450 component increased with the advance of carcinogenic changes in the urothelium. The component reactive with AHP-8 was also detected in the cancer tissues of transplantation lines of rat bladder cancers. In contrast, the cytochrome P-450 components recognized by APL-1, APL-2 or APF-3 were undetectable or present at low levels throughout the BBN carcinogenesis. These results suggest that a certain cytochrome P-450 component(s), probably a high-spin form of cytochrome P-448, is selectively induced in urothelium in association with neoplastic bladder lesion.

Animals↗
PubMed1987

The precursor cells of mouse lymphokine-activated killer (LAK) cells.

Culture of mouse spleen cells with recombinant human interleukin 2 (r-IL 2) resulted in the generation of lymphokine-activated killer (LAK) cells, which could lyse a variety of tumor cells. Negative selection study using various kinds of antibodies and complement indicated that LAK precursor cells existed in both mature spleen lymphocytes and immature spleen null cells. LAK cells were also induced from lymph node cells but not from unfractionated thymocytes. However, hydrocortison (HC)-resistant thymocytes or PNA- thymocyte subpopulations were highly responsive to r-IL 2 and maturated into LAK cells after 5 day-culture with r-IL 2. Moreover, it was demonstrated that r-IL 2 allowed the induction of LAK cells from nude mouse spleen cells, but not from bone marrow cells and fetal liver cells.

Animals↗
PubMed1987

[Properties of glycoprotein 125, a proliferation-associated cell surface antigen].

A cell surface antigen expressed in association with cell proliferation was detected and identified as a glycoprotein of molecular weight of 125,000 (gp 125) by using monoclonal antibodies against human or rat bladder cancer cells. We have found that appearance of the antigen in stimulated lymphocytes precedes the emergence of interleukin 2 receptor and that it is regulated by Ca ion and protein kinase-C.

Animals↗
PubMed1987

[Analysis of bladder cancer-associated antigens and their application to diagnosis].

Hybridomas were established from cell fusions between mouse myeloma cells and mouse or rat spleen cells immune to human and rat bladder tumors. Hybridoma supernatants were screened for their immunospecificities against human bladder tumor tissues and urine samples from bladder cancer patients. Eight monoclonal antibodies including 2 rat antibodies were selected and their diagnostic values were assessed. Four mouse monoclonal antibodies termed IMb1, IMb4, IMb9 and PMe were shown to react with 40 to 80% of human bladder tumor tissues examined in this investigation. No relationship was found between the antigen expression and clinical stage of the disease. Four other antibodies termed S31, S85, HBP-1 and RBM-1 reacted with antigens in urine from patients with bladder tumor and other types of uropathy, in contrast to their low reactivities against urine from healthy donors.

Animals↗
PubMed1987

Basic approach to application of chemoimmunoliposomes for cancer therapy.

Liposomes were coated with subunits of murine IgM monoclonal antibody and used as the carrier of an anti-cancer drug. The antibody-coated liposomes containing an anti-cancer drug (chemoimmunoliposomes, CIL) were prepared by using dactinomycin and monoclonal antibodies against a human bladder cancer or a mouse mammary cancer. These CIL exhibited antigen-specific binding to the target cancer cells, and there was more efficient killing of the target cancer cells than with free dactinomycin. In the mouse mammary cancer model, the CIL showed a therapeutic effect in both local and systemic applications. Relevance of this CIL model to cancer therapy and the problems related to its development are discussed.

Animals↗
PubMed1986

Genetic control of glycolipid expression.

A polymorphic variation of sialic acid species of sialosyllactosylceramide was found in dog erythrocytes. The analysis of the glycolipids in the erythrocytes of the individual dogs in a family of a Japanese breed of dog, Shiba-Inu, showed that the expression of sialosyllactosylceramide containing N-glycolylneuraminic acid was an autosomal dominant trait over the expression of that containing N-acetylneuraminic acid. Polymorphic variations of major liver gangliosides were also found in various strains of inbred mice. The strains were classified into three groups; the first group possessed only II3 NeuGc-LacCer, the second group possessed II3NeuGc-GgOse3Cer in addition to II3NeuGc-LacCer and the third group possessed II3NeuGc-GgOse4Cer and II3NeuGc,IV3NeuGc-GgOse4Cer as well as the above two gangliosides. By subjecting mice of these three groups to genetic analysis, the strain of the first group (WHT/Ht mice) was demonstrated to be a recessive homozygote which had a single autosomal defective gene making it unable to express N-acetylgalactosaminyltransferase activity to produce II3NeuGc-GgOse3Cer. The strains of the second group (BALB/c and C57BL/10 mice) were also demonstrated to be recessive homozygotes which had a single autosomal defective gene making them unable to express high enough level of galactosyltransferase activity to produce II3NeuGc-GgOse4Cer. By the analysis of gangliosides and the enzyme activity of H-2 congenic mice and mice produced by a mating, this defective gene controlling the expression of II3NeuGc-GgOse4-Cer through the regulation of the transferase activity was demonstrated to be linked to H-2 complex on chromosome 17.

Animals↗
PubMed1986

Molecular cloning and nucleotide sequence of the lipase gene from Pseudomonas fragi.

The gene coding for the lipase of Pseudomonas fragi was cloned into Escherichia coli JM83 by inserting Sau3A-generated DNA fragments into the BamH I site of pUC9. The plasmid isolated, pKKO, was restriction mapped and the position of the lipase gene on the 2.0 kb insert was pinpointed by subcloning. DNA sequencing revealed that the open reading frame comprises 405 nucleotides and gives a preprotein of 135 amino acids with a predicted Mr of 14643. By comparing the putative lipase amino acid sequence with porcine pancreatic, rat lingual and Staphylococcus hyicus lipases the amino acid sequence around the reactive serine was found to be common among the types of lipase which have been reported.

Amino Acid Sequence↗
PubMed1986

Induction of a high spin form of microsomal cytochrome P-448 in rat liver by 4-aminoazobenzene derivatives.

Male Sprague-Dawley rats were treated with 4-aminoazobenzene derivatives or other drug metabolizing enzyme inducers such as phenobarbital, 3-methylcholanthrene and isosafrole. The expression of hepatic microsomal cytochrome P-450 of the rats, principally that of a high spin form of cytochrome P-448 (cytochrome P-448H), was assessed by a bacterial mutation test and by immunological methods. The results of the mutation test with use of Salmonella typhimurium TA9 and 3 aromatic amine substrates showed that 2-methoxyl, 3-methoxyl and 2',3-dimethoxyl derivatives of AAB and methyl derivatives of AAB such as o-aminoazotoluene, N-methyl-4-aminoazobenzene and N,N-dimethyl-4-aminoazobenzene have a large capacity for the selective induction of cytochrome P-448H. Activity of the cytochrome increased by 6 hr after an azo dye treatment, reached a maximum after 24 hr, and then declined. In contrast, 4'-methoxy-AAB has a small, and AAB has no, capacity for the cytochrome induction. The aminoazo dye-induced enzymes differ in their substrate specificities from those induced with 3-methylcholanthrene or phenobarbital, and the induced enzyme was identified to be cytochrome P-448H, as determined by an enzyme-linked immunosorbent assay and immunoblotting with use of anti-cytochrome P-448 monoclonal antibodies. These observations indicate that several methoxyl and methyl derivatives of 4-aminoazobenzene are potent and selective inducers of cytochrome P-448H in the rat.

Animals↗
PubMed1986

Functional analogues of bleomycin: DNA cleavage by bleomycin and hemin-intercalators.

New hemin-intercalators (Hem-G's) that cleave DNA were synthesized, on the basis of 2-amino-6-methyldipyrido[1,2-alpha:3',2'-d]imidazole (Glu-P-1) as an intercalator moiety. Hem-G's, which possess an intramolecular ligand of the ferrous ion (a histidine or imidazole moiety), cleave DNA very efficiently and act at guanine-pyrimidine sequences preferentially. Bleomycin (BLM) also cleaved DNA with the same base-sequence selectivity shown by Hem-G's. The 5'-terminus of the DNA fragments cleaved by Hem-G's or by BLM is a phosphoryl group, while the 3'-terminus of the cleaved DNA fragments does not possess a 3'-phosphoryl group. There are more than three kinds of 5'-end 32P-labeled DNA fragments, which can be substrates of terminal deoxynucleotidyl transferase (TdT). One of the 3'-termini of the cleaved DNA fragments is a 3'-hydroxy group. The mobility of the 3'-end 32P-labeled DNA fragment cleaved by Hem-G's or by BLM corresponds to the removal of pyrimidine bases having guanine at the 5'-side. The mobility of one kind of the cleaved 5'-end 32P-labeled DNA fragments corresponds to the removal of guanine having pyrimidine at the 3'-side, followed by 3'-dephosphorylation. We propose that there exist plural mechanisms for DNA cleavage by Hem-G's or by BLM. The deduced structures of the cleaved DNA fragments suggest that one of the mechanisms involves deletion of two nucleotide units from DNA.

Base Sequence↗
PubMed1986

Mouse liver gangliosides.

The major gangliosides from mouse liver were purified and characterized by t.l.c., g.l.c., sialidase treatment, and a methylation study. GM3(NeuAc), GM3(NeuGc), GM2(NeuGc), GM1(NeuGc), and GDla(NeuGc, NeuGc) were identified. The structural identification of three of the gangliosides, GM2(NeuGc), GM1(NeuGc), and GDla(NeuGc, NeuGc), was supported by the results of 1H-n.m.r. analysis, and the structures of GM3(NeuGc), GM2(NeuGc), and GM1(NeuGc) were further confirmed by negative-ion fast-atom bombardment mass spectrometry. Ganglioside mapping showed that there was polymorphic variation of gangliosides in the liver of inbred strains of mice and that the major gangliosides were GM3(NeuGc) in WHT/Ht, GM2(NeuGc) in BALB/c and C3H/He, and GM2(NeuGc), GM1(NeuGc), and GDla(NeuGc, NeuGc) in ICR mice. Gangliosides containing N-acetylneuraminic acid, except for GM3(NeuAc), were not detected as major gangliosides in the strains of mice we analyzed.

Animals↗
PubMed1986

Effect of serum on inhibition of DNA synthesis in leukemia cells by cis- and trans-(Pt (NH3) 2C1(2)).

We examined the inhibition of DNA synthesis by cis- and trans-diamminedichloroplatinum (II) (cis- and trans-Pt) in leukemia cells, YAC-1 and RADA1. The degree of inhibition by trans-Pt was about the same as that by cis-Pt in vitro in the absence of serum, but the former was much lower than the latter in vivo or in the presence of serum in vitro. Atomic absorption studies showed that the amount of trans-Pt trapped by the serum in vitro is much larger than that of cis-Pt. Therefore, the amount of trans-Pt bound to DNA in vivo must be considerably smaller than that of cis-Pt, which eventually results in the antitumor-inactive nature of trans-Pt.

Animals↗
PubMed1986

Administration of slowly released recombinant interleukin 2. Augmentation of the efficacy of adoptive immunotherapy with lymphokine-activated killer (LAK) cells.

When recombinant human interleukin 2 (r-IL-2) was given to mice by single subcutaneous (s.c.) injection it rapidly disappeared from the blood. However, administration of slowly released r-IL-2 using mini-osmotic pumps caused a significant prolongation of serum levels of IL-2. Using a method for assaying IL-2 in vivo, it was also demonstrated that both the viability and the cytotoxicity of lymphokine-activated killer (LAK) cells could be maintained at a high level in vivo by administration of slowly released r-IL-2 rather than by a single injection of r-IL-2. In addition, we successfully treated EL4-bearing mice by combination therapy consisting of LAK cells and slowly released r-IL-2.

Animals↗