Chinese Hamster Cells In Culture Research Articles

Mutagenicity studies on alcohol extracts from gamma-irradiated potatoes.

The alcohol extracts freshly prepared from gamma-irradiated potatoes were examined for their mutagenic activity in bacterial and mammalian cell systems. Negative results were obtained from all following test systems: Mutation assays with Salmonella typhimurium His- strains such as TA 100, TA 98, TA 1535, TA 1537, and streptomycin-dependent mutant (SMd) strain, TA 100-10, inductests with Escherichia coli strains, K 12 GY 5027 and K 12 C600, chromosomal aberration tests with Chinese hamster cells in culture, as well as micronucleus tests in mice. In addition, no difference in the mutagenic activities was found between extracts prepared from the irradiated and the unirradiated potatoes, suggesting that no mutagenic substance was produced in potatoes following gamma-irradiation.

Effects of 5-thio-D-glucose on cellular adenosine triphosphate levels and deoxyribonucleic acid rejoining hy hypoxic and aerobic Chinese hamster cells.

Intracellular adenosine triphosphate (ATP) levels were measured in both hypoxic and aerobic cultures of V79 Chinese hamster cells treated with 5-thio-D-glucose (5-SH-D-Glc). This glucose analog, a known inhibitor of D-glucose transport and metabolism, reduced ATP in cell cultures allowed to become hypoxic by cell metabolism, but not in aerobic cultures treated similarly. Cells depleted of ATP were unable to rejoin x-ray-induced deoxyribonucleic acid (DNA) strand breaks as measured by the alkaline surcrose gradient sedimentation technique. Results show that 5-SH-D-Glc leads to reduced energy metabolism in cells dependent on glycolysis for ATP production. The inference for radiation therapy is that inhibition glucose metabolism selectively depletes energy reserves in hypoxic cells, rendering these cells more radiosensitive and leading to a more effective tumor treatment.

Mutation of Chinese hamster cells by near-UV activation of promutagens

A tissue-culture assay for mutagenesis and cytotoxicity incorporating near ultraviolet (NUV) light activation of polyaromatic hydrocarbons (PAH) has been developed. Cultures of Chinese hamster cells (line CHO) growing in suspension culture were inoculated with benzo[ a]pyrene (B[ a]P), 7,12-dimethylbenzanthracene (DMBA) or shale-oil retort-water and exposed to light from a high-pressure mercury lamp fitted with a Corning NUV bandpass filter. This light source both permitted activation of PAH and the shale-oil water and preculded detectable damage to DNA. Neither the PAH nor the NUV alone had any effect on cell survival or mutation frequencies but the chemicals plus NUV were extremely effective in producing mutations to 6-thioguanine resistance ( hgprt gene).

Mutagenic activity of 3 diazoacetylglycine derivatives on V79 Chinese hamster cells

3 diazoacetylglycine derivatives, diazoacetylglycine amide (DGA), diazoacetylglycine ethyl ester (DGE) and diazoacetylglycine hydrazide (DGH), known as antitumour agents, and shown to be mutagenic in bacteria, were studied for their mutagenic activity in the HGPRT system of V79 Chinese hamster cells in culture (V79/HGPRT system). All 3 drugs were highly effective in inducing 6-thioguanine-resistant mutants at concentrations that were not significantly cytotoxic.

Effects of a tumor promoter and an anti-promoter on spontaneous and UV-induced 6-thioguanine-resistant mutations and sister-chromatid exchanges in V79 Chinese hamster cells

The effects of a tumor promoter 12- O-tetradecanoylphorbol-13-acetate (TPA) and/or an anti-promotor antipain (protease inhibitor) on spontaneous and ultraviolet-induced sister-chromatid exchanges (SCEs) and 6-thioguanine- resistant (6TG r) recessive mutations were examined in V79 Chinese hamster cells in culture. TPA and/or antipain neither significantly altered base-line and UV-induced immediate SCE frequencies, nor decreased the level of delayed SCEs which persisted 6–7 days after irradiation. TPA and/or antipain appeared to enhance the recovery of UV-induced 6TG r colonies at the plateau expression phase despite non-mutagenicity by themselves and unaltered metabolic co- operation. Thus, the results conceivably imply that the 6TG r-recessive mutation expression, but not fixation, can be modulated at the cell level by the TPA and/or antipain. Our results, together with the recent results of Loveday and Latt, may argue against the notion that TPA enhances the antipain-suppressible SCEs as an index of mitotic recombination in relevance with a tumor-promotion mechanism.

Mutagenicity of K 2Cr 2O 7 in Chinese hamster cells in culture

Mutagenicity of K 2Cr 2O 7 in Chinese hamster cells in culture

Resistance to methyl mercaptopurine riboside in cultured hamster cells: Preliminary characterization of resistant cells and conditions affecting their selection in quantitative mutation studies

Cells resistant to high concentrations of methyl mercaptopurine riboside (MMPR), an analogue of adenosine, were found at a frequency of about 6 × 10 −6 per viable cell in untreated cultures of V79 Chinese hamster cells. Resistant cells were selected less efficiently if purines were added to the MMPR-medium, but high cell densities had little effect upon selection. 6 independently-isolated spontaneous MMPR-resistant sublines were characterized by their resistance to the toxicity of different purines, rate of purine excretion, incorporation of radioactive adenosine, electrophoresis of cell extracts, and expression of resistance in hybrids to an MMPR-sensitive line. 5 of these sublines showed recessive expression of MMPR-resistance in hybrids and had characteristics consistent with loss of adenosine kinase activity, while the remaining subline was much less resistant to MMPR and showed semi-dominant expression of resistance without loss of adenosine kinase activity. Cells with high resistance to MMPR were not found in a “tetraphoid” (hybrid) V79 line or in freshly-isolated human cell cultures, but occurred at a comparable frequency to V79 in another commonly-used aneuploid hamster line, CHO-K1. The frequency of MMPR-resistant cells in V79 cultures was increased to a similar extent by treatment with γ-rays or with ethyl methanesulphonate providing a suitable post-treatment interval was allowed for the expression of resistance. A genetic interpretation of these data is given in which it is proposed that resistance most usually arises through mutation of an autosomally-linked gene of which one copy has been inactivated or lost in V79 and in CHO-K1 cells. In comparison to published data on the selection of “mutants” resistant to 6-thiguanine, it is argued that MMPR could be as useful a selective agent as thioguanine and may select a different range of types of mutagenic event.

Enhancement of excision-repair efficiency by conditioned medium from density-inhibited cultures in V79 Chinese hamster cells: evidence for excision repair as an error-free repair process.

Conditioned medium from density-inhibited V79 Chinese hamster cell cultures, given as a post-treatment to UV-irradiated homologous cells, was demonstrated to reduce the lethal action of ultraviolet light by temporarily blocking DNA replication. Since the increased survival was not affected by various non-toxic concentrations of caffeine, such protective effect would be attributable to the prolonged intervention of excision repair before DNA replication during the post-treatment period. The influence of conditioned medium on the UV-induced mutation at the ouabain-resistance locus was also examined and a significant decrease in mutation frequency was noted. The observed reduction in killing and mutation as a result of post-incubation in conditioned medium, which delays DNA replication, would be interpreted as evidence that conditioned medium provides a longer period of time for an error-free excision-repair process, leaving lesion in DNA available for error-prone post-replication repair.

Effect of inhibition of synthesis in the S-period on the subsequent course of mitosis in a synchronized chinese hamster cell culture

Hemopoietic cells including CFUs could be washed off from the organ culture of fetal liver periodically for 4 weeks. Under the cultivation conditions employed this treatment did not reduce the CFUs content of the culture essentially; thus, the washings off could be used to elevate the CFUs yield per culture.

Effects of cytostatics on proliferating and stationary cultures of mammalian cells

The cytotoxic effects of dibromodulcitol (DBD), dianhydrodulcitol (DAD), vincristine (VCR), N-formyl-leurosine (F-LEU) and bleomycin (BLM) were studied on logarithmically growing and plateau-phase cultures of Chinese hamster cells. One hour treatment with DBD yielded sigmoid type survival curves. Synchronized cells were sensitive to the drug throughout the cell cycle, especially at late G 1 . DBD killed rapidly growing cells at a higher rate than plateau-phase cells. DAD proved to be equally toxic to both types of cells. At DAD concentrations above 15 μg/ml plateauphase cells were more sensitive than logarithmically growing cultures. The dosedependent survival curves for DAD were of the same type as the DBD curves. VCR and F-LEU produced survival curves characteristic of phase-specific drugs, killing specifically S-phase cells in synchronous cultures. Plateau cells were more sensitive to VCR than log cells, while F-LEU especially killed log cells. BLM produced biphasic survival curves, damaging plateau-phase cells almost 18 times more effectively than log cells.

Late changes in the course of mitosis in a synchronized Chinese hamster cell culture after inhibition of RNA and protein synthesis

The effect of inhibition of synthesis of various types of RNA and proteins at different periods of interphase on the course of late mitosis was studied in a synchronized culture of Chinese hamster cells. Analysis of the mitotic index and forms of pathology of division showed that the action of different doses of actinomycin D and puromycin in the first half of interphase produces an identical effect: C-mitoses in the immediate and late waves of cell division. Suppression of synthesis of total cell RNA and proteins in the second half of interphase was accompanied by delay of the cells in metaphase, with scattering of the chromosomes, evidence of a disturbance of the synthesis of the components of the division spindle. It is suggested that proteins (tubulins) and RNA participate as a reserve pool in the organization of the division spindle of late mitosis.

Effect of inhibitors of RNA and protein synthesis on the course of mitosis in a synchronized culture of Chinese hamster cells

The synthesis period of various types of RNA and proteins during the interphase related to the immediate after synchronization mitosis was investigated on teh synchronized culture of Chinese hamster cells with inhibitors. Analysis of MI and forms of mitosis pathology indicated the proteins functionally related to the cell division to be mainly synthesized during the second half of the interphase. The most important action of pyromycin during this time is the induction of C-mitosis on account of suppression of proteins synthesis responsible for the spiralization of chromosomes during the prophase. Inhibition of the r RNA transcription at any stage influence the cytotomy and reconstruction of daughter nuclei, this being accompanied by the delay of the cell exit from the mitosis. In difference from pyromycin, a high dose of AMD as an inhibitor of the whole cell RNA synthesis was most effective during the first half of the interphase. It is suggested that during this period at least three kinds of RNA are synthesized which are functionally related to the immediate mitosis in a different way: RNA for genome reduplication, RNA for the tubulin synthesis, and RNA which is the division spindle component.

Liver cell-mediated mutagenesis of mammalian cells by liver carcinogens.

A cell-mediated mutagenesis assay using primary cultures of rat liver cells and V79 Chinese hamster cells has been developed. Liver carcinogens and their structural analogues were studied. Mutations in the V79 cells were characterized by resistance to ouabain. Cocultivation of the liver cells and V79 cells in the presence of the carcinogens N-nitrosodimethylamine, N-nitrosodiethylamine, and aflatoxin B1 caused the induction of ouabain-resistant mutants of V79 cells. In the absence of liver cells, the carcinogens did not induce ouabain resistance. The analogues N-nitrosomethyl-tert-butylamine and aflatoxin G2 were not mutagenic. The carcinogens exhibited a dose-dependent enhancement of mutation frequency. The mutation frequency also increased with increasing numbers of liver cells seeded. It is suggested that such an experimental system may be useful for screening for chemical carcinogens.

Post-treatment with caffeine and the induction of gene mutations by ultraviolet irradiation and ethyl methanesulphonate in V-79 Chinese hamster cells in culture

The effect of caffeine on V-79 Chinese hamster cells after ultraviolet irradiation or treated with ethyl methanesulphonate was investigated. Caffeine strongly potentiated the killing of both agents, but it had no effect on the induction of mutations at the hypoxanthine-guanine phosphoribosyl transferase locus. The results are consistent with the notion that caffeine slows down an error-prone post-replicative repair mechanism without changing the mutation frequency.

Modified method of differential staining of sister chromatids

A modified method of obtaining differential staining of sister chromatids is described. It is simple, rapid, and effective, and at the same time inexpensive and accessible, since it allows one to use available reagents. When 5-bromdeoxyuridine was administered 24 hours before fixation into the Chinese hamster cell culture the percentage of metaphases with a differential chromatid staining constituted 95--98, and when this substance was administered 28 hours before fixation into the human lymphocyte culture this percentage varied from 75 to 92, depending on the individual. The mean number of sister chromatid exchanges in human lymphocytes failed to depend on the time of fixation.

The Use of Hypoxic Cell Radiosensitizers with Mixed X and α Radiations

When used alone, radiations of mixed LET will not completely eliminate the radioresistance of hypoxic cells. Hypoxic cell sensitizers also may not completely eliminate the problem because of toxic effects of the compounds at higher concentrations. Experiments were performed using a mixed LET 85 percent dose of x rays and a 15 percent dose of ..cap alpha.. particles from plutonium) and a 0.4 mM concentration of the hypoxic cell sensitizer Roche-07-0582. Cultures of both human cells (line T-1) and Chinese hamster (line CHO) cells were used in this investigation. The results indicate that the radioresistance of hypoxic cells can be dealt with more effectively by a combination of high-LET radiation and a hypoxic cell sensitizer than by either of these alone.

The effect of aflatoxins on Chinese hamster cells in culture

The effect of aflatoxins on Chinese hamster cells in culture

Inhibition by caffeine of post-replication repair in Chinese hamster cells treated with 7-bromomethylbenz(a)anthracene: Enhancement of toxicity, chromosome damage and inhibition of ligation of newly-synthesized DNA

The lethal and chromosome damaging effects of 7-bromomethylbenz(a)anthracene (7-BMBA) on Chinese hamster cells in culture were enhanced by posttreatment incubation in non-toxic concentrations of caffeine. 7-BMBA produced a marked dose-dependent depression in the rate of DNA synthesis seen as a dose-dependent delay in the time of appearance of the peak rate of DNA synthesis in G 1-treated synchronous cultures. Posttreatment incubation in medium containing caffeine partially reversed the hydrocarbon-induced depression of DNA synthesis. The rate of ligation of DNA molecules was reduced in 7-BMBA-treated cells relative to untreated of caffeine-only treated cells. If the time allowed for ligation of DNA was extended to compensate for the reduced rate of DNA synthesis in 7-BMBA-treated cells then the size of DNA attained that in control cells. Thus no evidence for so-called “gaps” in newly-synthesized DNA was obtained. Posttreatment incubation in the presence of caffeine produced a very marked further decrease in the rate of ligation of DNA molecules despite the increase in rate of DNA synthesis. The size of the small molecular weight nascent DNA synthesized in the presence of caffeine during 3 h after treatment approximated to the distance between arylalkylations in the template DNA. These effects of newly-synthesized DNA are discussed in terms of a model which permits replication to proceed past unexcised lesions in DNA by a process of replication repair which can be inhibited by incubation of treated cells in a non-toxic concentration of caffeine. The relevance of these observations to the modification of DNA by carcinogenic compounds in general is discussed.

Comparative radiobiology of fast neutrons: Relevance to radiotherapy and basic studies

Comparative neutron radiobiological properties relevant to radiation therapy are being investigated using V79 Chinese hamster cells in culture. Three neutron beams are being used: linear accelerator-produced neutrons, p + → Be, at the Cancer Therapy Facility of the Fermi National Accelerator Laboratory (Fermi); cyclotron-produced neutrons, d + → Be, at the Franklin McLean Research Institute, University of Chicago (FMI): and fission-produced neutrons at the JANUS Reactor of the Argonne National Laboratory (JANUS). The mean neutron energies of the Fermi, FMI and JANUS beams are 25 MeV, 3.6 MeV and 0.85 MeV, respectively. The RBE values of cell survival relative to 250 kVp X-rays, at a given surviving fraction, decreased in the order JANUS, FMI, Fermi, a trend opposite to the mean neutron energy. The OER's measured with Fermi and JANUS neutrons were essentially the same, however. Other radiobiological studies have been initiated with JANUS neutrons. Results from two-dose fractionation experiments showed that very little repair of neturon-induced sublethal damage occurred for incubation at 37°C for intervals up to 5 hr between exposures. Data from postirradiation growth kinetics indicated that neutron radiation produces cell division delays that increase linearly with dose. An RBE for cell division delay, relative to 55 kV X-rays, was calculated to be approximately 3.5, which is close to the upper limit of the RBE values for survivals. Hyperthermic treatment, immediately following neutron irradiation, enhanced cell killing. Postirradiation treatments of cells with hypo- and hyper-tonicity also resulted in enhanced cell killing.

Carbohydrate energy sources for Chinese hamster cells in culture.

Three Chinese hamster cell lines (CHO-K1, V79, and CHS) were tested to see which carbohydrates would support their growth in culture. There were differences between lines (mannose supported growth of CHO-K1 only), and glucose was the only carbohydrate that supported growth of all three strains. CHO-K1 cells possess enzymes for hydrolyzing some disac-charides which do not support growth. Fetal calf serum contains enzymes that hydrolyze maltose, glycogen, and a-mannosides.