Study of the mammary stem cells / progenitors committed to the development of the bovine mammary gland

Milk production is highly dependent on the extensive expansion of the mammary epithelium which largely occurs during puberty. Within the mammary epithelium, the Mammary Stem Cells (MaSC) and their progeny play a key role in driving the development of the future functional mammary gland. Later, these cells contribute to regenerate the secretory epithelium in order to ensure future lactations. To elucidate the epithelial cell lineage involved in the establishment of the functional gland, we investigated the MaSC/progenitors populations in the bovine mammary tissue at various physiological stages. We used flow cytometry to phenotype epithelial cells on the basis of CD49f and CD24 expression in animals at three contrasted physiologic stages: puberty (heifers of 17 months of age), lactation (multiparous cows producing 35 kg of milk) and drying off (cows at 6 years after drying off). Comparison of the cytometric profiles highlighted the presence of a distinct epithelial population of CD49flowCD24pos cells at puberty (23.7%±3.6) which substantially decreased in the others stages (4.1%±1 at lactation and 6%±1 at drying off). These CD49flowCD24pos cells had features of a dual lineage, with luminal and basal characteristics (CD10, ALDH1 and keratin 7 expression) and were considered to be early common progenitors. The MaSC fraction was recovered in the sub-population of CD49fhighCD24pos cells. When cultured in vitro (in a 3D culture system with matrigel), these cells formed mammospheres, a functional property of MaSC. At puberty, total mammary cells contained 2.8%±0.3 of MaSC. This proportion of MaSC was significantly reduced at lactation (1.1±0.3%) and at drying off (0.4±0.3%). Further phenotypic characterization of the CD49fhighCD24pos cells showed that they expressed CD10 and keratin 14 at any stages. This in-depth characterization of the epithelial sub-populations at various physiological stages provide new insights into the epithelial cell hierarchy in the bovine mammary gland from puberty to drying off.