Growth and morphogenesis of epithelial cell organoids from peripheral and medial mammary parenchyma of prepubertal heifers

Collagen gel cell cultures were used to test the hypothesis that cells from peripheral parenchymal zones of the developing bovine mammary gland have greater proliferative and morphogenic potential than cells from the medial parenchymal mass. Tritiated thymidine was incorporated to assess cell proliferation. Dose responses for insulin-like growth factor one (IGF-I, 0 to 50 ng/ml) and transforming growth factor beta one (0 to 5 ng/ml) were determined for each cell population. Cells from the peripheral regions were two to three times more sensitive to the mitogenic action of IGF-I or serum. Transforming growth factor beta one concentrations of 12.5 to 500 pg/ml stimulated cell proliferation, but concentrations above 1 ng/ml were inhibitory. Peripheral cells showed a distinct biphasic response to addition of transforming growth factor beta one. Morphology of organoids was evaluated daily during culture and by histologic examination at the end of culture. Peripheral zone cells formed multi-layered cell structures that resembled native mammary parenchyma with serum or mammary gland extract. Our results indicate that epithelial cells of the peripheral and medial parenchymal zones in the ruminant mammary gland can differ in their proliferative and morphogenetic response to growth factor treatment in vitro. These differences likely reflect local tissue regulation necessary for sequential ductular and lobuloalveolar development in vivo.