Flow cytometry analysis of cells dispersed from the MtTF4 tumor whose growth is inhibited by estradiol treatment

The aim of this work was to determine whether treatments of rats with estradiol (E) in conditions known to decrease the proliferation rate, the mitotic index and the thymidine incorporation into the DNA of the MtTF4 tumor act at a specific point in the cell cycle. Two weeks after grafting a piece of tumor under the kidney capsule, adult male Fischer rats were treated or not treated with E. Tumors were collected between 12 h and 11 days later. Cells were dispersed by collagenase-DNAse treatment and fixed with ethanol. DNA content, cell size, cell granularity and protein content were analyzed, alone or in combination with a flow cytometer. E treatments did not apparently modify the distribution of cells according to their DNA content whereas they did increase dramatically cell size, cell granularity and cell protein content. Simultaneous analysis of DNA content and light scattering or protein content allowed us to demonstrate that there was an increase of a population of large granular and protein-rich cells regardless of the phase of the division cycle considered. These effects are time-dependent, dose-dependent and hormone-specific. This work shows both the interest of flow cytometry to describe the consequences of E treatment at any phase of the cycle of cells dispersed from a solid tumor and the limits of this method in the conditions used to specify the E target points: at the present time, it cannot be decided whether E acts at one or several points of the cell cycle for inhibiting tumor growth.