Conjugated linoleic acids (CLAs) are a group of dietary fatty acids that are widely marketed as weight loss supplements. The isomer responsible for this effect is the trans-10, cis-12 CLA isomer (10e12z). 10e12z CLA increases lean body mass by reducing both lipid storage and lipid synthesis as well as adipocyte differentiation. These studies were designed to address whether 10e12z CLA directly or indirectly altered adipose function, and whether adipose dysfunction induced by 10e12z CLA impacted the development of the mammary epithelium. Adipocyte differentiation is regulated by the ligand activated receptor Peroxisome Proliferator-Activated Receptor (PPAR) γ. 10e12z CLA did not alter PPARγ transcriptional activation of an in vitro reporter assay, nor did it affect lipid storage induced by the thiazolidinedione rosiglitazone, a potent PPARγ ligand, suggesting that CLA did not interact directly with PPARγ. However, gene expression analysis by microarray suggested that 10e12z CLA increased expression of inflammatory signaling cascades and extracellular matrix remodeling proteins consistent with NF-κB activation by Toll-Like Receptors (TLRs). TLR4 is activated by fatty acids, but 10e12z CLA did not activate TLR4 in reporter assays. However, consistent with NF-κB activation, COX-2 was induced by 10e12z CLA in mouse adipocytes both in vitro and in vivo. COX-2 was not directly responsible for 10e12z CLA-induced alterations in lipid storage and differentiation, but it did induce epidermal growth factor (EGF) receptor (EGFR) ligands in a prostaglandin F_2α (PGF_2α)-dependent manner. Inflammation, extracellular matrix (ECM) remodeling and activation of EGF receptors are consistent with activation of proliferative signaling that occurs in breast cancer. Therefore, the influence of 10e12z CLA-induced adipocyte-derived factors on the mammary epithelium was examined. Wild type mice fed 10e12z CLA exhibited increased ECM remodeling and decreased mammary adipose content. Proliferation in the mammary gland was also increased two-fold by these treatments as measured by bromodeoxyuridine incorporation into DNA. 10e12z treated adipocytes in vitro recapitulated both the increased proliferation and invasion seen in vitro. Gene expression profiles of mammary glands from mice fed 10e12z CLA matched profiles of adipocytes, but not mammary epithelial cells, or macrophages grown in vitro. These results demonstrated that 10e12z CLA altered the expression of adipokines in the mammary fat pad, and that these factors govern the response of other tissues to 10e12z CLA. Although this isomer is reported to possess anti-carcinogenic activities the results of these investigations demonstrate that 10e12z CLA exerts maked effects on adipose-epithelial interactions and therefore, caution is advised with regard to consumption of this fatty acid as a dietary supplement.