Cell density dependence of focus formation in the C3H/10T1/2 transformation assay

Some of the dynamics of neoplastic transformation in vitro have been studied with the use of benzo(a)pyrene as the carcinogen in the C3H/10T1/2 morphological transformation assay. Experiments that involved the dsipersion of cells into new culture dishes at various times after carcinogen treatment have shown that no change in the fraction of potentially transformed cells occurs while cultures grow to form a confluent monolayer, that little or no change in the fraction of potentially transformed cells occurs for approximately 3 weeks after confluence is attained, and that this fraction increases rapidly some 7 weeks after BP treatment. When confluent benzo(a)pyrene-treated cultures are dispersed in new culture dishes prior to the onset of growth toward focus formation, the formation of transformed foci is suppresssed at high cell densities of seeding. This phenomenon is independent of the total number of divisions undergone by cells after treatment. We suggest that phenotypic expression of morphological transformation is dependent on colony interactions in the C3H/10T1/2 system, which we do not yet understand, but which are independent of time posttreatment either in cell generations or absolute time.