Selection and Characterization in Culture of Mammary Tumor Cells with Distinctive Growth Properties in Vivo

Abstract Two epithelial subpopulations with different growth capabilities have been isolated from a mammary tumor cell line. The parent cell line, WAZ-2T, was derived from an adenocarcinoma which developed spontaneously from hyperplastic BALB/c mouse mammary tissue. The subpopulations were isolated using agarose suspension cultures from which anchorage-independent (+SA) and -dependent (-SA) WAZ-2T cells were cultured. Both +SA and -SA sublines grew rapidly in monolayer culture and exhibited stable epithelial morphologies similar to that of the parent line. The cloning efficiency of +SA cells in agarose was 35% while -SA cells did not grow in suspension culture even when plated at high cell density (10 6 cells/25-sq cm flask). Population doubling times in vitro were 17 and 20 hr for +SA and -SA cells, respectively. Production of plasminogen activator was approximately, 2-fold greater in +SA cultures as compared to -SA cultures. Both +SA and -SA cells were tumorigenic in BALB/c mice when inoculated s.c. or i.v. However, the latency period for s.c. tumor development was markedly greater for the -SA subline. Similarly, after 4 weeks but not after 7 weeks, significantly more lung nodules developed from i.v. injection of +SA cells than -SA cells. The greater number of +SA lung nodules was not due to increased cell adhesion in the lung. We conclude that established cell lines from mammary tumors can contain widely heterogeneous subpopulations and that these subpopulations provide excellent material for establishing the significance of correlations between in vivo and in vitro behavior.