Bovine mammary myoepithelial cells. 1. Isolation, culture, and characterization

The objective of this study was to isolate, purify, culture, and characterize myoepithelial cells from bovine mammary glands. Myoepithelial cells were separated from other mammary and blood cells after collagenase digestion and centrifugation using metrizoate-ficoll gradients. Myoepithelial cells were identified by their characteristic morphology and cloned using selective detachment. They contained many densely packed myofilaments, very few cytoplasmic organelles, elongated surface projections, and a dense, irregularly shaped nuclei. Some cells were as large as 1.2 mm in culture. Myoepithelial cells contained an extensive network of cytoskeletal proteins, including alpha-smooth muscle actin, alpha-actinin, and vimentin. When cultured, they tended to repel one another and never grew as closely associated cells. The myoepithelial nature of these cells was verified by showing that they contracted in response to oxytocin, bound oxytocin, and did not produce casein. Myoepithelial cells from fetal and lactating glands grew very well in culture. Active division of myoepithelial cells could be maintained for at least 3 mo, and cells could be serially subcultured at least seven times. The successful isolation and culture of bovine mammary myoepithelial cells make utilization of these cells possible in order to study their role in mammary growth and differentiation and milk ejection.