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Regulation of mammary gland remodeling during the lactation cycle in cattle still remains unclear. The present study focused on the role of TGF-ß1 and somatotropic pathways proteins in control of the switch between survival and death of bovine mammary epithelial cells (MEC). Expressions of TGF-ß1, TGF-ßRII, IGF-IRα, IGF-IRß, GH-R, IGFBP-3, -4, and -5 in mammary tissue explants in Holstein-Fresian heifers (n=7) and cows (n=23) in early lactation (1-100 day), late lactation (200-260 day) and drying off (280-340 day) were compared with biochemical indices of apoptosis (caspase 3, 89 kDa fragment of PARP) and autophagy (Beclinl). The results revealed that an increase in apoptosis during the dry period was accompanied by highly significant increases in TGF-ß1 and TGF-ßRII expression. Beside biochemical markers, typical morphological features of apoptosis, such as cell shrinkage, separation from the neighboring cells and condensation of chromatin were observed. TGF-ß1 expression and induction of apoptosis was facilitated by the suppression of somatotropic pathway during drying off, manifested with down-regulation of GH-R and IGF-IRα, and up-regula- tion of IGFBP-4 and -5. This is the first report describing autophagy in the bovine mammary gland. Similarly to apoptosis, the intensity of autophagy was the highest in the dry period, as shown by increased expression of Beclinl and morphological features, e.g. autophagosomes, autophagic vacuoles. Autophagy observed in the involuting mammary tissue could be the natural cell defense against transient undernourishment and action of apoptogenic peptides (e.g. TGF-ß1, IGFBPs), thus maintaining cellular homeostasis in the dry period.
Our earlier studies carried out in silico demonstrated that different transcription factors have their putative binding sites in the 5’-flanking regions of bovine milk protein genes. Now we extended our study to include the experimental analysis of these transcription factors. This study on electrophoretic mobility shift assay (EMSA) of nuclear proteins derived from bovine mammary glands showed for the first time the presence there of CREB, NF-κB, Oct1 and Sp1 transcription factors. The pattern of DNA-protein complexes of Sp1 transcription factor significantly differed between virgin heifers and lactating cows. Transition from pregnancy to lactation was associated with changes in the DNAprotein binding patterns of NFI and Oct1 transcription factors.
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