Characterization of rat mammary epithelial cell subpopulations by peanut lectin and anti-Thy-1.1 antibody and study of flow-sorted cells in vivo.

A cell separation method was developed for studies of the growth kinetics of rat mammary epithelial cell (RMEC) subpopulations in grafts and in culture in vitro. By flow cytometry of RMEC stained with fluorescein isothiocyanate-peanut agglutinin and phycoerythrin-anti-Thy-1.1 monoclonal antibody, we could distinguish four cell subpopulations from primary cultures of 7- to 8-week-old F344 female rat mammary glands: both negative (B-), PNA+, Thy-1.1+, and both positive (B+). We studied the growth patterns of these subpopulations in vitro for 1 to 14 days in complete hormone medium (CHM) containing 10% fetal bovine serum and prolactin, 17 beta-estradiol, cortisol, progesterone, and insulin. The fractions of PNA+ and B- cells steadily decreased with time in culture. The fraction of Thy-1.1+ cells steadily increased with time in culture. There were small numbers of B+ cells. We grafted RMEC in hyperprolactinemic recipient rats. The mean numbers of transplanted cells required to produce at least one alveolar unit in 50% of the graft sites (AD50 values) are inversely related to the clonogenic fractions. AD50s of RMEC are as follows: unsorted mammary cells cultured in CHM for 1 to 4 days, approximately 220; unsorted cells cultured in CHM for 7 days, approximately 550; sorted PNA+ RMEC from outgrowths of 3-day cultures, approximately 82; B+, approximately 350; B-, approximately 545; Thy-1.1+, approximately 9372. We conclude that the PNA+ cell subpopulation includes most of the clonogenic cells. Thy-1.1+ cells appear to be terminally differentiated; they are likely to be myoepithelial cells.