Abstract
The study objective was to evaluate the effect of endocrine and mechanical (gel release) signaling on bovine mammary epithelial cell ultrastructure and gene expression. Cultures receiving only one stimulus demonstrated partially differentiated ultrastructure, which included abundant polysomes, limited rough endoplasmic reticulum, and absence of secretory products, whereas the 2 stimuli together induced a more complete lactogenic phenotype that included increased rough endoplasmic reticulum, abundant lipid droplets, and secretory vesicles containing casein micelles. The structural data indicated that although synthesis of milk components was initiated, the copious synthesis and secretion associated with stage II lactogenesis was not evident. Microarray analysis revealed that both prolactin and gel release independently regulated several genes linked to a wide array of cellular activities. In combination, they regulated fewer genes targeted to lactogenesis. Genes regulated by the combination treatment included claudin 7, multiple caseins, xanthine oxidoreductase, and several protein synthesis, packaging, and transport genes. Genes related to structural activity including keratin 15 (morphogenesis), α-spectrin (cell shape via actin cytoskeleton), and chitinase-like protein 1 (tissue remodeling) were up-regulated by the combination treatment as was the transcription factor Kruppel-like factor 2 (KLF-2). However, Snail 2, which down-regulates and inhibits tight junction components, was repressed in response to the combination treatment. These results suggest coordination between endocrine and physical signals at the genomic level that produces a more specific and targeted transcriptional response associated with stage I lactogenesis. A molecular pathway analysis of the differentially expressed genes revealed that genes regulating cell signaling were linked to those regulating cell structure and adhesion.
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