Di-N-Octylphthalate Acts as a Proliferative Agent in Murine Cell Hepatocytes by Regulating the Levels of TGF-β and pro-Apoptotic Proteins

Di-n-octylphthalate (DNOP) is a phthalate used in the manufacturing of a wide variety of polyvinyl chloride (PVC)-containing medical and consumer products. A study on chronic exposure to DNOP in rodents showed the development of pre-neoplastic hepatic lesions following exposure to a tumor initiator. The objective of this study was to identify the mechanisms by which DNOP leads to pre-neoplastic hepatic lesions. Healthy mouse hepatocyte AML-12 and FL83B cells were treated with DNOP (10, 100 and 1,000 ppm). The rate of cell proliferation was increased in treated cells with DNOP in a concentration-dependent manner as early as 24 h. Using quantitative PCR, DNOP caused an increase in expression of transforming growth factor-β (tgf-β) in both cell lines, and primary culture mouse hepatocytes. The novel selective TGF-β receptor type I/II (TβRI/II) inhibitor LY2109761 impaired the effect of DNOP on cell proliferation. The presence of pro-apoptotic proteins decreased in the presence of 1,000 ppm DNOP. In in-vitro and ex-vivo models, DNOP did not change the methylation status of any tumor suppressor gene studied. Our observation indicates that DNOP, through an increase in the expression of tgf-β and a decrease in the presence of pro-apoptotic proteins, acts as a proliferative agent in normal mouse hepatocytes. We also studied the morphological and functional changes of the mouse liver upon a short-term treatment of DNOP for a month. Although DNOP had no effect on body weight or liver weight, it caused a decrease in albumin and increase in vimentin indicating an epithelial-to-mesenchymal transition state in mouse hepatocytes. Mice exposed to DNOP displayed cholestasis, which was reflected in an increase in hepatic bile acids and glutathione levels.