Figure 6

Parity-identified mammary epithelial cells (PI-MECs) remain lineage-restricted during pregnancy. (A) Immunofluorescence on paraffin sections from the 7-day second pregnancy time point, showing alveoli in which all cells are yellow fluorescent protein-positive (YFP^pos), except for rare luminal cells expressing estrogen receptor alpha (ERα) (red, yellow arrows) or basal (CK8-negative, white arrowhead) cells. (B) Alveolar clusters at day 14 of the first pregnancy that already consist mostly of YFP^pos cells also contain hormone-sensing (HS) cells (labeled by an antibody against the progesterone receptor, red) which do not express YFP. Scale bar is 10 μm. (C) Mammary epithelial cells from 7-day pregnant glands (second pregnancy) labeled with CD24 and CD49f to segregate luminal (blue gate) from basal (red gate) single-cell populations. (D) YFP^neg cells and YFP^pos cells (PI-MECs) are shown separately on density/contour plots for CD24 and CD49f. (E) Sca1 and CD49b plots of the luminal cell population at 7 days of pregnancy, showing the HS cell population (purple gate) and alveolar cells (orange gate). (F) Distribution of YFP^neg cells and YFP^pos PI-MECs within populations of HS cells and of alveolar cells. Note that the combined density/contour plot shows the relative distribution of the population on display and therefore the HS population appears larger in (D) than in (C) because the YFP^pos population is plotted separately in (D). (G) Distribution of YFP^pos cells across the different populations. Inset: magnified basal and HS cell bars. (H) Percentage of YFP^pos cells observed within the basal, HS, and alveolar cell populations. Error bars indicate standard deviation of four independent mice. (I) A small percentage of ER⁺ cells are also YFP^pos (arrowhead), although most HS cells are YFP^neg (arrow). Scale bar is 10 μm.