DNA synthesis and content in the nuclei of epidermal cells of mice during their differentiation and specialization

Synthesis and content of DNA in the nuclei of differentiating cells of mouse skin epidermis was studied by using cytomorphometric, autoradiographic and cytophotometric methods. It has been shown that the cells of the keratinoid series divide only in the basal layer and contain 2-4c DNA. Keratinocytes of the thorny layer are mostly tetraploid, 2c cells are lacking. H4c and 8c cells comprise 12% of the population. In the keratinocytes of the granular layer DNA content is somewhat lower due to nuclei break down and conversion of cells into anucleate scale. Part of the melanocytes of the basal layer also contain 4c DNA. Highly specialized element of the basal layer Merkel and Langerhans cells are polyploid. Conclusion is drawn that DNA hyper-replication by multiplication of the whole genome is part of the development program of the population.     

Cell population kinetics and DNA content during thyroid carcinogenesis

The proliferation kinetics and DNA content of thyroid follicular cells in rats were studied by autoradiography and cytophotometry. Continuous treatment of animals with methylthiouracil (MTU) results in hyperplasia followed by tumour growth in the thyroid gland. The mitotic index (MI) increases from 0.006 +/- 0.002% in controls to 0.13 +/- 0.06% in hyperplasia and to 0.09 +/- 0.03% in malignant cells. The same is true for the labelling index (LI) which rises from 0.08 +/- 0.003% in controls to 1.4 +/- 1.1% in hyperplasia and to 1.0 +/- 0.6% in follicular adenomas. The S-phase duration (TS) is shortened from 8.0 +/- 1.2 hr in controls to 6.0 +/- 1.4 hr in animals treated for 9 months with MTU and prolonged to 15.4 +/- 2.1 hr in papillary carcinomas. In all MTU-treated animals a decrease in the value of the potential population doubling time (TPD) and thyroid weight doubling time (TD) was observed. The cell loss factor (phi) decreases in animals treated for 3 months with MTU and increases during the stage of tumour growth in the gland (animals treated 12-15 months with MTU). DNA measurements in the nuclei of hyperplastic and neoplastic thyroid tissues reveal cells with values exceeding that in control animals. However, no difference was found in the DNA content between thyroid adenomas and carcinomas, nor between thyroid hyperplasia and neoplasia.     

DNA content and synthesis in the nuclei of rat cerebellar cells in organotypic cultures

Using cytophotometric and autoradiographic methods, it has been shown for the first time that in condition of an organotypic culture the replicative synthesis of DNA is induced in the Purkinje neurons of the cerebellum of newborn rats completing their terminal differentiation. This synthesis is accompanied by polyploidization of the initially diploid population of these cells (4c, much more rarely 8c, and a single 16c cell appear) rather than by cell division. In constant, the granular cells mostly retain their diploid state and only a few of them synthesize DNA to H2c values. The glial cells divide actively. Hence, evidence is presented that neurons, at least those of cerebellum, retain their potential of replicative synthesis of DNA in the organotypic culture. The important point is that DNA synthesis in their nuclei proceeds simultaneously with processes of differentiation.     

Effect of azidothymidine on reactivation of DNA synthesis in macrophage nuclei contained in heterokaryons

EIn heterokaryons, DNA synthesis is reactivated in macrophage nuclei only in the case of fusion with immortal cells. Assuming that telomerase is responsible for reactivation, the effect of its inhibitor azidothymidine (AZT) was studied in heterokaryons of mouse resident peritoneal macrophages and immortal 3T3 Swiss cells. AZT suppressed reactivation of DNA synthesis in macrophage nuclei and had no effect on DNA synthesis in 3T3 Swiss cell nuclei, suggesting an altered telomere structure in normal mouse macrophages.     

Changes in numbers of pancreatic acinar cell nuclei and in DNA content during raw soya flour feeding in mice

Nuclei of pancreatic cells were isolated by trypsin-detergent digestion of fresh tissue and stained with propidium iodide, and nuclear DNA was measured by flow cytometry. Samples were isolated from mice fed either chow or raw soya flour (RSF) for periods ranging from 1 day to 48 weeks, beginning at 4 weeks of age. In chow-fed mice, the pancreas contained about 80% diploid (2N) and 20% tetraploid (4N) cells at the start of the study, but tetraploidy gradually increased to about 40% 2 weeks later (6 weeks of age) and remained at this level from that time onwards. Low levels of octaploid nuclei (8N) were also present in some animals after 2 weeks. In RSF-fed mice, about 20% tetraploid nuclei were also present for 1 and 2 days after starting RSF, but by 4 days tetraploidy had increased significantly to 40% and by 14 days had further increased to 50%. This level was significantly higher than that seen in chow-fed animals and was maintained for up to 48 weeks. Significantly higher numbers of octaploid nuclei were also present in the RSF-fed animals. In both chow- and RSF-fed mice, most cells were mononuclear, averaging 70% in chow-fed and 64% in RSF-fed animals. This difference was significant. This study shows that the mouse pancreas differs from the rat pancreas in the absence of a large population of binucleate acinar cells and the presence of considerable nuclear tetraploidy. Raw soya flour feeding leads to significant changes in these features, but in this species these changes do not appear to predispose to neoplasia.     

Multiple forms of DNA methylases in hepatic cell nuclei of animals in the normal state and in pathology

Using hydrophobic chromatography multiple nature of DNA-methylating enzymes have been revealed. It was established that the most active enzymatic fraction of rats' liver and that of chicken are completely identical.     

DNA content in the mouse spermatogonia B and spermatid nuclei during treatment with insecticides

The DNA content in nuclei of germ cells: spermatogonia B and spermatides in the maturation phase of male mice receiving commercial insecticide preparations commonly used in this country was cytophotometrically measured. The following insecticides were tested: 1% Metox-30 solution, containing 30% of methoxychlorine (p,p-dimethoxydiphenyltrichlorethane), 0,3% solution of Sadofos-30 containing 30% of malathione (1,2-dicarboethoxyethyl-0,0-dimethyldithiophosphate) and 0,3% solution of Foschlor-50 containing about 50% trichlorfon (2,2,2-trichloro-1-hydroxyethyl-0,0-dimethyl-diphosphonate). It was found that a statistically significant number of spermatogonium B and spermatide nuclei in the phase of maturation of the animals treated with the insecticides exhibited an abnormal DNA content as compared with the nuclei of control animals. The chromatin of these nuclei is also more sensitive to acid hydrolysis in Feulgen reaction.     

Suppression of DNA synthesis in normal, but not neoplastic, nuclei by cytoplasmic extracts from resting cells

We have previously shown that a heat-stable protein in cytoplasmic extracts from human quiescent peripheral blood lymphocytes (PBL) is capable of inhibiting the induction of DNA synthesis in isolated resting nuclei. We now report that these cytoplasmic extracts are also capable of suppressing DNA synthetic activity in replicative nuclei isolated from mitogen-activated PBL. PBL extracts had little or no inhibitory effect, however, on replicative nuclei derived from several transformed lymphoblastoid cell lines. These results suggest that the growth of normal lymphocytes may be negatively controlled by cytoplasmic inhibitory factors. Furthermore, the relative resistance of tumor cell nuclei to these inhibitory signals provides a possible explanation for the loss of growth control in neoplastic cells.     

Changes in DNA content of nuclei in rust uredospore germlings during the start of differentiation

Digital video microscopy in conjunction with the DNA-binding fluorescent probe 4′,6-diamidino-2-phenylindole was used to determine the relative DNA content of single nuclei in germ tubes of uredospores of the bean rust fungus (Uromyces phaseoli), a parasite of bean plants (Phaseolus vulgaris). The uredospores develop a series of infection structures in response to contact stimuli, and the first structure to appear, the appressorium, occupies the stomatal opening. This study was made to determine the time after the start of germination on an inductive surface that DNA replication and mitosis occur. It was found that the start of DNA replication detected by increased nuclear fluorescence power of some individual nuclei in the population was nearly coincident with mitosis between 2.0 and 2.5 h after the start of germination and that the appressorium was completed about 30 min later. The fluoresence power of nuclei was about the same before DNA replication began as at the end of mitosis; hence the germ tube nuclei were in the G₁ phase of the cell cycle before mitosis.     

DNA/nuclear protein content in the evaluation of cell cycle modifications during colon carcinogenesis

OBJECTIVE: To investigate the colorectal adenomacarcinoma sequence by biparametric DNA/nuclear protein flow cytometry with the aim of evaluating cell cycle modifications during carcinogenesis. STUDY DESIGN: Paraffin-embedded specimens of 27 adenomas with mild/moderate dysplasia, 20 adenomas with severe dysplasia/intramucosal adenocarcinomas, 28 adenocarcinomas and 14 normal colon mucosa specimens were analyzed by biparametric DNA/nuclear protein content flow cytometric analysis in order to evaluate cell cycle modifications during colorectal carcinogenesis. RESULTS: The mean G0-G1A fraction of the cell cycle was 50.6% (SD +/- 17.2), 25.7% (SD +/- 15.1), 27.8% (SD +/- 11.7) and 29% (SD +/- 13.8) for normal mucosa, adenomas with mild/moderate dysplasia, adenomas with severe dysplasia and adenocarcinomas, respectively. The difference between normal mucosa and the other groups was statistically significant (P < .05), while no significant differences were detectable between adenomas with different degrees of dysplasia and adenocarcinomas. CONCLUSION: Our results show a decrease in G0-G1A in adenomas with mild/moderate dysplasia, suggesting that modification of the cell cycle may represent an early step in colon carcinogenesis, and they support the hypothesis that disregulation of cell cycle-controlling genes is an early event in the adenoma-carcinoma sequence.     

Semisynthetic derivatives of daunorubicin and carminomycin: inhibition of DNA synthesis after intravenous administration to mice

Inhibition of DNA synthesis in the liver, kidneys, spleen and heart of mice after intravenous administration of 0.1 and 0.3 LD50 of semisynthetic derivatives of rubomycin (daunorubicin) and carminomycin was studied. The level of DNA synthesis inhibition was estimated by a decrease in incorporation of (methyl-3H) thymidine. Under the action of 13-trebutoxycarbonyl hydrazone and 14-salicyloiloxy derivatives of rubomycin and carminomycin maximum inhibition of DNA synthesis was reached later while its recovery started earlier as compared to the initial antibiotics.