Morphogenesis by dissociated immature rat testicular cells in primary culture

Dissociated cells from immature (15-day) rat testes maintained in primary culture were shown to undergo morphogenesis. Dissociated cells (1-4 X 10(6)) were cultured in 2 ml medium NCTC-135 containing 10% rat serum at 32.5 degrees C under an air; CO2 atmosphere (95%:5%). During the initial culture period (1-3 days) the dissociated cells attached to the culture dish and formed a monolayer. After 4-5 days of culture, aggregates of cells formed at discrete foci within the monolayer; these structures averaged 50 micrometer in diameter and numbered 3-4 X 10(3)/dish. Within 5-6 days of culture some of these aggregates (2-3 X 10(2)) released their contact with the substratum. Histological examination of released aggregates indicated a more regular, tissue-like organization with increasing time in culture. Aggregates at earlier time periods (4-5 days) exhibited concentric rings of cell while by day 6 some aggregates exhibited a vesicular appearance with a single layer of columnar epithelium surrounding a fluid-filled lumen. The type of structure formed varied with plating density; at densities less than 2 X 10(6)/dish the structures were spheroid while at greater densities tubes were formed. Time-lapse observations indicated that spheres formed by movements of cells into an aggregating center while tubes formed by a rolling from the edge of the monolayer. The cells comprising the vesicle epithelium appeared to be predominantly one cell type as determined by light microscopy. These results therefore indicate that dissociated testicular cells have the ability to reform tissue-like associations in culture.