Expression of initiated and promoted stages of irradiation carcinogenesis in vitro.

This study was initiated with the objective of separating and characterizing two or more nuclear subfractions, which could then be compared with respect to their relative propensity for binding carcinogenic polycyclic aromatic hydrocarbons. Nuclei were isolated from cloned AKR-2B mouse embryo cells, which are susceptible to transformation by chemical carcinogens. The nuclei were mechanically sheared and subfractions were separated by sedimentation through a 0.17 to 1.7 m sucrose gradient. When the cells were treated with [3H]uridine for 30 min, most of the label incorporated into RNA was recovered in the top region of the gradients, which represented Nuclear Subfraction I. The majority of the chromatin DNA, however, was localized in the bottom region (Subfraction II) and the pellet (Subfraction III). Precipitation (with CaCl2) of the rapidly labeled RNA of Subfraction I along with the chromatin DNA suggested that the label was present in nascent RNA chains still attached to the chromatin. Thus, the transcriptionally active chromatin seemed to be localized in Nuclear Subfraction I.

The chromatin of Subfraction I was also the best template for RNA synthesis in vitro with exogenous bacterial polymerase. The protein and RNA content of Subfraction I was greater than that of the other two subfractions and whole chromatin. Electron microscopy revealed the presence of membrane material in Subfractions I and II, with little such material in Subfraction III. Subfraction I differed from Subfractions II and III and whole chromatin with respect to thermal denaturation of the DNA and histone composition (as determined by gel electrophoresis). The acidic protein composition (as determined by gel electrophoresis) differed for the chromatin of all three nuclear subfractions.

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Separation and Characterization of Transcriptionally Active and Inactive Nuclear Subfractions of AKR Mouse Embryo Cells

The treatment of rat embryo secondary cultures with DMBA or DMBA-3H for 5, 9, or 24 h resulted in chromosome damage consisting mainly of chromatid type aberrations. There was an increase in the percentage of labelled nuclei and metaphases with increasing length of exposure. In terms of incidence of chromatid lesions, the largest telocentric chromosome (No. 2) was the most susceptible of the autosomes. Banding pattern analysis demonstrated that the region associated with negative band 2q24 of the No. 2 chromosome had the highest number of lesions. An increased accumulation of DMBA-3H label occurred in approximately the same chromatid area of a small fraction of cells exposed for either 5 or 9 h prior to mitosis. The complete loss of DMBA-3H chromosomal labelling after DNAse treatment suggests that the visible grains represent carcinogen-bound DNA. After DMBA and BrdUrd, there was an increase in the number of sister chromatid exchanges compared to controls treated with BrdUrd only; the location of the exchange points on chromosome No. 2 was similar in samples treated with either DMBA and BrdUrd or BrdUrd alone. Additional experiments with thymidine-3H showed that the non-random chromatid lesions on chromosome No. 2 may result from endogenous radiation from the incorporated tritium. These studies demonstrate that a specific chromosome may be affected by diverse agents and that chromatid lesions frequently occur at the site of sister chromatid exchanges.

Vulnerability of specific rat chromosomes to in vitro chemically induced damage.

Chloramphenicol and its isomer dextramycine decrease considerably the toxic effect of the carcinogenic hydrocarbons 7,12-dimethylbenz(a)anthracene or benz(a)pyrene on mouse embryo fibroblast-like cells grown in vitro.

Effect of chloramphenicol and dextramycine on the toxicity of carcinogenic hydrocarbons.

Cytologic detection of virus-specific DNA by in situ hybridization in syrian hamster embryo cells transformed by viral or combined chemical and viral treatment

Radioautography of Incorporation of 7,12-Dimethylbenz[a]anthracene-3H Label Into Syrian Hamster Embryo Cells During Exposure to Inhibitors of Protein Synthesis

N. C. Popescu

Papers

Radioautography of Incorporation of 7,12-Dimethylbenz[a]anthracene-3H Label Into Syrian Hamster Embryo Cells During Exposure to Inhibitors of Protein Synthesis