The molecular characterization of human mammary myoepithelial cells is incomplete, hindering our understanding of its importance in breast physiology and pathology. Because data on the precursors of this cell lineage remain scarce and often contradictory, basal epithelial cells of second trimester fetal breasts were studied by light microscopy (LM) and immunohistochemistry (IHC). Up to 20 wk of gestational age, the mammary rudiments only comprised roundish primary outgrowths, “primary buds,” more likely to represent immature nipples than true mammary tissue. At 21 wk secondary outgrowths, “projections,” extended from enlarged primary buds into well-vascularized layers of dense mesenchyme. Basal projection cells had a partial myoepithelial-like phenotype: they reacted with CD29, CD49f, CD104, keratin 14, vimentin, S100β protein, and p63; furthermore, many became positive for keratin 17, α-smooth muscle actin, and CD10 (but not for keratin 19) between wk 21 and 25. The continuous basement membrane associated with the fetal mammary rudiments was strongly positive for collagens type IV and VII, and for laminin 5. Consistently strong and basally polarized staining for hemidesmosomal components suggested that although incompletely differentiated, most second trimester myoepithelial precursors might already mediate local epithelial-mesenchymal interactions, i.e., complex signaling pathways which are crucial for both orderly growth during development and maintenance of homeostasis during adult life. Because they are likely implicated in the phenomenon of menstrual cycle-related growth spurts in the adult resting breast, the strategically positioned cells of the myoepithelial lineage might constitute critical protagonists in defective epithelial-mesenchymal signaling associated with cancer progression.