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Biomedical subjects

C Heidelberger

Publications and source records attributed to C Heidelberger.

At least 55 records · Page 3Linked to original sources
PubMed1980

Comparison of adriamycin- and ouabain-induced cytotoxicity and inhibition of 86rubidium transport in wild-type and ouabain-resistant C3H/10T1/2 mouse fibroblasts.

Ouabain (OUA) inhibited 86Rb uptake (50% inhibitory concentration = 0.8 X 10(-4) M) over concentration ranges close to those at which it caused a reversible cytotoxicity (50% lethal dose = 2.5 X 10(-4) M) in growing wild-type C3H/10T1/2 cells. On the other hand, Adriamycin (ADM) inhibited 86Rb uptake (50% inhibitory concentration = 2 X 10(-3) M) but at concentrations 10(4)-fold higher than those causing irreversible cytotoxicity in growing wild-type cells (50% lethal dose = 3 X 10(-8) M). While OUA inhibited 86Rb uptake more in wild-type cells than in a OUA-resistant mutant, ADM inhibited 86Rb uptake to the same extent in confluent wild-type and OUA-resistant cells. Further, three OUA-resistant mutants were not cross-resistant to ADM- or daunomycin (DM)-induced cytotoxicity during log phase or to ADM-induced cytotoxicity at confluence. In addition, ADM, DM, or 5-iminodaunomycin did not displace the cardiac glycosides digoxin or digitoxin from their respective antibody complexes. The order of potency of anthracycline derivatives in inhibiting 86Rb uptake in confluent wild-type cells was the same as their order of inhibiting the growth of wild-type cells and in detaching confluent wild-type cells (DM > ADM > 5-iminodaunomycin) but did not correlate with their cardiotoxic potentials (ADM > DM > 5-iminodaunomycin). Therefore, in this model system, ADM cytotoxicity is mediated differently from OUA cytotoxicity. Further, we find no biological evidence consistent withADM binding to the OUA site on the cell surface (Na+-K+) adenosine triphosphatase and therefore no evidence in this model system that ADM cardiotoxicity could be a digitalis-type toxicity per se.

Animals↗
PubMed1980

Effect of 12-O-tetradecanoylphorbol-13-acetate on the morphology and growth of C3H/10T1/2 mouse embryo cells.

The effects of the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) on the morphology and growth properties of C3H/10T1/2 clone 8 cells were examined. The morphology of these cells was changed within 30 min following treatment with 0.1 microgram of TPA per ml; they became smaller and refractile with long beady processes. Such changes were observed in both logarithmic and confluent cultures and lasted about 72 hr. Subsequent treatments were much less effective in inducing these changes. Scanning electron microscopy showed cell retraction and rounding as the most significant immediate effects of TPA treatment; many cells remained partially rounded 48 hr afterwards. Long-term surface modifications ascribable to TPA treatment were not detected. TPA had only minor effects on the growth of cultured C3H/10T1/2 cells in the presence of 10% fetal calf serum. Slight increases in plating efficiencies and saturation densities were generally observed in the presence of TPA but not with the related non-tumor-promoting compound phorbol. The cells grew slowly in 1% fetal calf serum and demonstrated serum batch-dependent alterations in their growth properties when exposed to TPA. Under conditions that produce doubling times of 70 hr or greater, TPA, but not phorbol, reduced the doubling time to about 50 hr. Saturation densities were also increased by TPA in 1% fetal calf serum. The effects of TPA on the growth of an oncogenically transformed variant of C3H/10T1/2 were quite different. While minimal effects of TPA were observed quite different. While minimal effects of TPA were observed when transformed cells were treated in the presence of 10% total calf serum, TPA treatment in 1% fetal calf serum significantly inhibited cell growth.

Animals↗
PubMed1979

Establishment of epithelial cell lines from adult mouse regenerating liver.

A simple technique for developing epithelial cell lines from regenerating mouse liver has been described. Twenty-one epithelial cell lines have been developed and can be divided into four groups according to their morphology. All these near diploid cell lines have the capacity to metabolize diverse classes of chemical carcinogens (3-methylcholanthrene, 2-acetylaminofluorene, dimethylnitrosamine and aflatoxin B1) to cytotoxic metabolites. It is not yet possible to determine which ones of these cell lines originated from hepatocytes. Studies are in progress to further characterize and to use these cell lines as lethally irradiated feeder layers for cell-mediated activation of various classes of chemical carcinogens and mutagens with C3H/10T1/2 mouse embryo fibroblasts as indicator cells.

2-Acetylaminofluorene↗
PubMed1979

Role of thymidylate synthetase activity in development of methotrexate cytotoxicity.

Methotrexate (MTX) inhibition of the growth of mouse or human leukemia cells in culture was partially prevented by either thymidine (dThd) or hypoxanthine. 5-Fluoro-2'-deoxyuridine (FdUrd) also decreased the growth-inhibitory potency of MTX in the presence of small concentrations of 5-formyltetrahydrofolate (citrovorum factor) and sufficient exogenous dThd to support the synthesis of thymidylate nucleotides by salvage mechanisms. In addition, citrovorum factor-induced reversal of MTX was several orders of magnitude more efficient in the presence of both FdUrd and dThd than in the presence of dThd alone or in the absence of both nucleosides. Likewise, the presence of FdUrd (3 microM) and dThd (5.6 microM) completely prevented the lethality of 0.3 mM MTX to L1210 cells in culture medium supplemented with micromolar concentrations of citrovorum factor. We propose that this protection against the cytotoxic effects of MTX by dThd, hypoxanthine, and FdUrd have a common biochemical mechanism--namely, inhibition of the de novo synthesis of thymidylate by either a direct [FdUrd; inhibition of thymidylate synthetase (thymidylate synthase; 5,10-methylenetetrahydrofolate:dUMP C-methyl-transferase, EC 2.1.1.45)] or indirect (dThd and hypoxanthine; feedback inhibition by anabolites on ribonucleotide reductase and deoxycytidylate deaminase) effect. The resultant decreased rate of loss of reduced folates due to de novo thymidylate synthesis would allow a higher degree of inhibition of dihydrofolate reductase to be endured without damage to the cell.

Animals↗
PubMed1979

Chemical carcinogens produce mutations to ouabain resistance in transformable C3H/10T1/2 Cl 8 mouse fibroblasts.

Chemical carcinogens induce mutations to ouabain resistance in the transformable mouse fibroblast cell line C3H/10T1/2 Cl 8. The mutant phenotype is stable and heritable in the absence of selective agent, and dose--response curves for mutant frequency were obtained with N-menthyl-N'-nitro-N-nitrosoguanidine, 3-methylcholanthrene, benzo[a]-pyrene, N-acetoxy-N-2-acetylaminofluorene, and the anti-7,8-diol-9,10-epoxide of benzo[a]pyrene. The ratio of the malignant transformation frequency to the mutation frequency was 12 for benzo[a]pyrene and 21 for N-acetoxy-N-2-acetylaminofluorene. The development of the mutational assay reported here allows the use of this permanent cell line for comparison of mutation and transfomration frequencies and as a screening system for xenobiotics that pose mutagenic or carcinogenic hazards to mammalian cells.

2-Acetylaminofluorene↗
PubMed1979

Biochemical determinants of tumor sensitivity to 5-fluorouracil: ultrasensitive methods for the determination of 5-fluoro-2'-deoxyuridylate, 2'-deoxyuridylate, and thymidylate synthetase.

Techniques have been developed to measure FdUMP, the active metabolite of 5-FUra; thymidylate synthetase (TMP synthase; 5,10-methylenetetrahydrofolate:dUMP C-methyltransferase, EC 2.1.1.45), the target enzyme for this antimetabolite; and dUMP, the substrate that competes with FdUMP for binding to TMP synthetase. As little as 0.02 pmol of FdUMP can be quantitated with a competitive ligand binding assay by using homogeneous Lactobacillus casei/MTX TMP synthetase as a binding protein. A new binding assay for TMP synthetase allows detection of 0.005 pmol of enzyme. The quantitative enzymatic conversion of dUMP to [methyl-(14)C]-TMP using 5,10-methylene[(14)C]tetrahydrofolate by pure L. casei TMP synthetase is used as an assay for dUMP with a sensitivity of 10 pmol. Cultured CCRF-CEM human lymphoblastic leukemia cells formed high levels of FdUMP (2.6 nmol per 10(9) cells) within 11 hr after exposure to 30 muM 5-FUra. Tumor cell TMP synthetase levels dropped, and then free FdUMP appeared. The intracellular dUMP pool was low (2-5 nmol per 10(9) cells) in logarithmically growing cultures of several tumor cell lines but expanded rapidly in CCRF-CEM cells on exposure to 5-FUra after enzyme levels decreased. The levels of dUMP found after exposure to 5-FUra are sufficient to severely retard inhibition of TMP synthetase by FdUMP.The methods described are sufficiently sensitive to allow these biochemical parameters of 5-FUra action to be measured in cell culture or in needle biopsy samples of human tumors.

Cell Line↗
PubMed1979

Determinants of 5-fluorouracil sensitivity in human tumors.

It is not apparent that advanced human carcinomas of the breast or the large bowel are conprised of at least two populations: those responding to treatment with 5-fluorouracil (approximately 20 per cent) and those unresponsive to this drug. This classification cannot be made before chemotherapy on the basis of any clinical parameter. Biochemical parameters to distinguish between responding and nonresponding tumors are being sought in this laboratory. Techniques have been developed to measure thymidylate synthetase, the target enzyme for 5-fluorouracil, 5-fluoro-2'-deoxyuridylate, the active form of the antimetabolite, and 2'-deoxyuridylate, the naturally occuring competing metabolite. These methods are sufficiently sensitive to permit analysis of these parameters in needle biopsy specimens, and do not require exposure of patients to radioisotopes.

Biopsy, Needle↗
PubMed1978

Enhancement of oncogenesis in C3H/10T1/2 mouse embryo cell cultures by saccharin.

Impure and pure samples of saccharin (2 milligrams per milliliter) did not produce oncogenic transformation of C3H/10T1/2, clone 8, mouse embryo fibroblasts. However, after treatment of the cells with a nontransforming initiating dose (0.1 microgram per milliliter) of 3-methylcholanthrene, continuous treatment with either sample of saccharin (100 micrograms per milliliter) led to significant transformation. It is concluded that in this system saccharin is a cocarginogen, probably functioning as a promoting agent that is 1000-fold less active than the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate.

Animals↗
PubMed1978

Separation of glutathione S-transferase activities with epoxides from the mouse liver h-protein, a major polycyclic hydrocarbon-binding protein.

The C3H mouse liver h-protein is a cytoplasmic protein to which metabolites of carcinogenic polycyclic hydrocarbons bind covalently following i.p. injection. It has a number of physical properties similar to those of the glutathione S-transferases (EC 2.5.1.18). These properties include molecular weight (40,000), number of subunits (2), basic isoelectric point around 8.0, sedimentation coefficient (3.5S), and subcellular localization. In this communication, we have shown that glutathione S-transferase activities with 1,2-epoxy(3-p-nitrophenoxy)propane and benz[a]anthracene 5,6-oxide as substrates were separated from the h-protein on carboxymethylcellulose and isoelectrofocusing columns. The purification of the mouse h-protein as a [3H]-7,12-dimethylbenz[a]anthracene conjugate or as the free form is also described.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1978

Enhancement of X-ray transformation by 12-O-tetradecanoyl-phorbol-13-acetate in a cloned line of C3H mouse embryo cells.

A phorbol ester promoting agent, 12-O-tetradecanoyl-phorbol-13-acetate, enhances X-ray transformation in vitro in a two-stage fashion similar to that shown previously for ultraviolet radiation and chemical carcinogens. In studies with a mouse embryo-derived cell line (C3H/1OT 1/2 clone 8), there were clear interactive effects between X-radiation and 12-O-tetradecanoylphorbol-13-acetate. These were particularly marked when a minimally transforming X-ray dose (50 or 100 rads) was followed by 12-O-tetradecanoylphorbol-13-acetate treatment beginning either immediately after the radiation exposure or 48 to 96 hr later.

Animals↗
PubMed1978

Ultraviolet light in the oncogenic transformation of cultured C3H/10T1/2 mouse embryo cells.

Skin tumors are produced in rats, mice, and humans by exposure to UV light. We developed a mouse embryo fibroblast line C3H/10T1/2, which can be transformed by chemical carcinogens, X-irradiation, UV radiation, and oncornavirus. When we irradiated these cells with 10, 25, 50, 100, 150, or 200 ergs/mm2 of UV light, neither transformation nor cytotoxicity was observed at the two lower doses. When the irradiated cells were cultured in medium containing 0.1 micrograms TPA/1 ml (a potent tumor promoter) starting from 0-120 hours after irradiation, a high frequency of transformation was always produced. When the cells were initiated with subeffective concentrations of 0.1 micrograms MCA/ml followed by UV radiation at different intervals, no transformation occurred; however, these initiated cells were transformed after TPA treatment. When we treated the cells with multiple exposures to UV light, no transformation occurred then nor when the cells were treated with TPA followed by UV irradiation at different intervals. Thus UV in our system acts as a pure initiator in the two-stage process of oncogenic transformation.

Animals↗
PubMed1977

Liver homogenate-mediated mutagenesis in chinese hamster V79 cells by polycyclic aromatic hydrocarbons and aflatoxins.

Several chemical carcinogens that require metabolic activation have been examined for their cytotoxic and mutagenic activity in Chinese hamster V79 cells. Mutagenic activity was measured as the induced frequency of 6-thioguanine-resistant colonies. Metabolic activation was provided by the 9000 g supernatant fraction of rat liver plus cofactors. The cytotoxicity and mutagenicity of aflatoxin B1 and B2, benzo(a)pyrene, 3-methylcholanthrene, 7,12-dimethylbenz(a)anthracene, dibenz(a,h)-anthracene, dibenz(a,c)anthracene, and benz(a)anthracene were examined as functions of concentration. Except for the two isomers of dibenzanthracene, the mutagenic activity in general paralleled the carcinogenic activity. An assay of this type may be useful as a prescreen for environmental chemicals that require metabolic activation.

Aflatoxins↗