Analyzing the Role of TNF and#945; and Autophagy in Regulation of TGF and#946; Induced Epithelial to Mesenchymal Transition in Cancer Cells

Abstract

Cancer typically develops in a chronic inflammatory setting causal to the release of a plethora of cytokines. In this regard, the progression of liver cancer, hepatocellular carcinoma (HCC) is almost always associated with persistent inflammation. Components of the inflammatory tumor microenvironment (TME) thus plays a critical role in HCC pathogenesis. As a consequence of the intricate crosstalk, HCC is often associated with complex carcinogenesis rendering hindrances to chemotherapy. Considering the importance of this crosstalk between secretary products from the tumor milieu with HCC cells, in this study, we selected two cytokines prevalent in HCC TME; TGF-and#946; and TNF-and#945; and investigated their effect on HCC progression, especially their implications on epithelial to mesenchymal transition (EMT) and on cellular homeostatic process- autophagy. EMT is a pre-requisite for cancer cells to disseminate; and autophagy, based on current evidences is considered a dual edged sword which can have both cancer promoting or inhibitory effects. We observed SMAD2 signalling dependent significant elevation of EMT markers upon TGF-and#946; (transforming growth factor- and#946;) exposure to HCC cells. Interestingly, simultaneous exposure to another cytokine, TNF-and#945; significantly reduced TGF-and#946; induced EMT by activating the expression of inhibitory SMAD7 and elevating intracellular ROS (reactive oxygen species) levels leading to cell death. TNF-and#945; mediated antagonism of TGF-and#946; induced effects was further validated in another cell type- human osteosarcoma cells as well. Importantly, TGF-and#946; exposed cells undergoing EMT, showed induction of autophagy, which when inhibited pharmacologically using CQDP (chloroquine di-phosphate) or genetically (siATG5), drastically enhanced ROS and suppressed EMT. In contrary, quenching of ROS by NAC (N-acetyl cysteine), reduced autophagy and resulted in a significant elevation of EMT. TNF-and#945; (tumor necrosis factor-and#945;) was found to inhibit EMT by elevating ROS levels and reducing autophagy. We hence prove that regulation of R