Identification of IGF binding proteins in bovine milk and the demonstration of IGFBP-3 synthesis and release by bovine mammary epithelial cells

Insulin-like growth factor system components are synthesized and secreted b y mammary epithelial cells and multiple IGF binding proteins (IGFBP) are fo und in milk of various species. This study was conducted to identify the IG FBP in bovine milk, to compare them with those found in blood, and to ident ify the cell(s) responsible for mammary IGFBP synthesis. Bovine blood, milk , and cell culture-conditioned media were analyzed and characterized with W estern ligand blot procedures for specific IGFBP. Electrophoresis and [I-12 5]IGF-II ligand blot analyses of the samples indicated that, unlike serum a nd mammary primary cell culture-conditioned media, milk required removal of casein in order to accurately disclose all IGFBP. Immunoprecipitation stud ies identified IGFBP-2, -3, -4, and -5 in blood, milk, and primary cell cul ture conditioned media. The IGFBP were present at higher concentrations in serum than in milk, and milk concentrations were greater than that shown in conditioned media from primary cultures of bovine mammary cells. Northern analysis detected IGFBP-3 messenger RNA in extracts from fresh tissue and c ells in culture, and in situ hybridization studies with fresh tissue utiliz ing probes for IGFBP-3 and alpha S-1-casein showed that the mRNA for IGFBP- 3 is predominant in the secretory epithelial cells, when compared to other tissue cell types.