Etude cytologique et cytochimique des effets de l'actinomycine D sur des fibroblastes de poulet ou de rat, cultivés in vitro

Abstract

The effects of actinomycin D on chick or rat fibroblasts cultivated in vitro have been studied by several cytological and cytochemical methods. A few hours after the beginning of treatment, the size of the nucleoli diminishes considerably as already shown by others for other materials. Living treated cells have been observed by phase contrast microscopy. The duration of metaphase is increased and the separation of daughter-cells at telophase is often inhibited. The mitotic activity is strongly depressed. The DNA content has been measured by cytophotometry after Feulgen reaction in individual cells during interphase or mitosis. The diploid DNA content is not modified by actinomycin D. The latter inhibits premitotic DNA synthesis as also shown by others. Furthermore we have shown that some cells become tetraploid as far as the DNA content is concerned but are blocked as this stage (G 2 or post-synthesis). Due to the treatment, some nuclei become granulo-filamentous; this nuclear degeneration can start at any moment of the cellular cycle. The nuclear total protein content and the nucleolar total dry mass per nucleus have been measured by micro-interferometry in rat fibroblasts. The DNA content has then been measured in the same cells by cytophotometry after Feulgen reaction. Premitotic nuclear protein and DNA synthesis is inhibited but not completely; some cells reach the preprophase (tetraploid nuclear total protein and DNA content). However, the majority of these cells cannot enter into morphological mitosis probably because the nucleoli are altered. In this respect, we have shown that the nucleolar total dry mass per nucleus is strongly reduced. By histo-autoradiography, we have observed that actinomycin D inhibits DNA, RNA and protein synthesis, as also shown by others. According to our results, in cells treated by low concentrations of actinomycin D, nucleic acids synthesis can be inhibited but protein synthesis remains normal. We have also shown by histoautoradiography that tritiated actinomycin D is linked to DNA in the nucleus. According to the results of experiments now in progress, cells treated by actinomycin D can recover under some conditions.