Cell growth and differentiation in vitro in mouse macrophages transformed by a tsA mutant of simian virus 40. II. Changes in the distribution of DNA content during the reversible transition between macrophage and nonmacrophage states in the cultures of tsA640-transformed macrophages.

Cultures of mouse macrophage cell lines transformed by wild-type or the tsA640 mutant of simian virus 40 (SV40) show a reversible phenotypic transition between the nonmacrophage (proliferating phase) and the macrophage (stationary phase) states (Takayama, 1980; Tanigawa et al., 1983). Distribution of DNA content in the cultures of the tsA640-transformed macrophage lines in the process of the phenotypic transition was determined by flow cytometry. Taking the mean DNA content of mouse peritoneal macrophages as 1 unit in the scale of fluorescence intensity in the flow cytogram, the transformed macrophages showed, at 33 degrees C, two peaks, one located around the 1.0-unit position (peak 1.0) and the other around the 1.6-unit position (peak 1.6), and a plateau distribution continuing to 3.2 units. Peak 1.0 was predominant in the stationary-phase culture, whereas peak 1.6 was predominant in the proliferating-phase culture. Almost the entire population of the strictly resting culture, which was obtained by culturing the stationary-phase culture for a further 5 days at nonpermissive temperature (39 degrees C), was phagocytic, and had accumulated at peak 1.0. Cells in peak 1.0 moved to peak 1.6 and to higher positions, after the strictly resting culture was sparsely reseeded and incubated at 33 degrees C. In contrast, the DNA content distribution of the successively proliferating cells, which were obtained by repeated passage of an extensively proliferating culture and none of which were phagocytic, was similar to that of proliferating hypotetraploid BALB/c3T3 fibroblasts with a G1 peak at 1.6 unit followed by a plateau containing S- and G2-phase cells. The peak 1.0 cell population appeared from the recloned population of the successively proliferating cells in company with the restoration of the culture condition-dependent phagocytic ability when cocultured with primary macrophages. Each peak in the flow cytogram reflected fairly well DNA content per cell as determined by other methods.