Different approaches have been used to study milk related traits in farm animals, reaching from statistical dissection of phenotypic variation to the search of candidate genes with major phenotypic effects. The aim of this study was to develop a new research tool devoted to in vitro studies of physiological pathways responsible for mamary gland development, lactation remodeling and immune response, supported by a user friendly map based bioinformatics tool for integration of different types of data. We established goat mammary gland derived primary epithelial cell line with predominantly epithelial morphology, responsive to lactogenic hormones and exhibiting regeneration potential in hetrologous mouse system. The response of primary epithelial cells to pathogenic bacterium Mycoplasma agalactiae was studied using RNA sequencing approach and 1553 differentially expressed genes were detected 24 h post infection. The majority of differentially expressed genes belonged to cell cycle regulating genes, pro-inflammatory cytokines, chemokines and genes involved in lipid metabolism. Bioinformatic analysis of 359 putative target sites for mammary gland expressed miRNAs revealed polymorphic miRNA target sites for bta-miR-199b, -199a-5p, and -361 in the IL1B gene and for -miR-126 in the CYP11B1 gene. Graphical integration of different types of data to DairyVis platform allowed identification of genomic regions with higher number of potential functional elements that deserve further experimental analysis. The newly developed MEC line and integration of bioinformatics tools into DairyVis database present a promising methodological support for further research in the field of lactation biology.
COBISS.SI-ID: 2932360
Up to date there has been no research done on mammary gland stem cells in goat, but there have been some studies done on epithelial fractions isolated from cow's mammary glands. Stem cells reside in basal cell population of mammary epithelium, which been proven for human and mice but not for mammary glands of other species. Although there is a vast of antibodies reacting with mice and human epitopes, isolation of pure stem cell population is still impossible. This is because definitive stem cell markers have not been defined so far. In our work we used in vitro and in vivo approaches to show that stem cells exist in caprine mammary gland. In an assay of clonogenic proliferation of progenitors we plated low numbers of epithelial cells mixed with mitomycin C treated fibroblasts on collagen coated dishes. The colonies that arose 6-8 days after plating were morphologically and immunofluorescently characterized. According to expression of specific lineage markers, namely cytokeratins 14 and 18, we determined the frequencies of progenitors. Percentage of progenitors was highly dependent on the sample tested, medium used, and age of the animal from which the sample was recovered. In an in vitro transplantation experiments we showed that the stem cells are capable of regeneration of mammary epithelial structures in collagen gels, after their transplantation under the renal capsule of NOD/SCID mice. The xenogels were removed after 4-6 weeks and within them structures that resemble the epithelium in mammary tissue arose, where cytokeratin 14-positive myoepithelial cells line the cytokeratin 18-positive luminal cells, which open into a lumen. This is a proof that there are cells in goat mammary gland that are capable of proliferation and regeneration.