Abstract C29: Free fatty acid induced epithelial to mesenchymal transition (EMT) in hepatocellular carcinoma cells

Background: The incidence and mortality rates of hepatocellular carcinoma (HCC) have steadily increased in the United States over the past few decades. From an epidemiological perspective, it has been observed that HCC related mortality rates are significantly higher in obese patients indicating a possible correlation between obesity and metastatic progression of HCC. At the physiological level, obesity manifests an elevation in free fatty acid levels (FFA) levels which is thought to be detrimental for liver cells. While most previous studies have focused on cytotoxic effects of elevated FFA, very few studies have investigated their potential role in progression of liver cancer. In a previous study, we found reduced expression of desmoplakin (DSP) in FFA treated HepG2 cells, suggesting a loss of cell adhesion – one of the features of cells undergoing metastatic progression via EMT. Hence, we hypothesized that elevated FFA levels possess the ability to induce EMT in HCC cells. Methodology and Results: We studied the in vitro effects of FFA on HepG2 and Hep3B cells – two human HCC cell lines of different etiologies. We performed all experiments using FFA conjugated to BSA (bovine serum albumin), and used BSA as treatment control. We first evaluated the overall impact of FFA treatment on cell adhesion. We performed a dispase-based dissociation assay and found significant loss of cell adhesion in FFA treated cells. Next, using scratch wound-healing assay and modified Boyden9s chamber assay, we found that FFA significantly enhanced the migration and invasiveness of HCC cells. We further confirmed the occurrence of EMT by measuring the change in expression levels of various markers (Keratin 8/18, Vimentin, E-cadherin) in FFA treated cells using Western blots. Additionally, we screened the change in expression levels of several transcription factors that have been described as inducers of EMT using qRT-PCR. Our studies indicated enhanced expression of a number of EMT-related transcription factors including SNAIL, TWIST, ZEB1 and FOXC2, suggesting a potential transcriptional regulatory mechanism by which elevated FFA could induce EMT. Furthermore, we evaluated the possibility of a direct role of DSP loss in inducing EMT. Using siRNA mediated DSP knock-down we found enhanced expression of EMT markers in both HepG2 and Hep3B cells. Conclusions and future perspectives: Our studies provide evidence that elevated FFA levels may be involved in metastatic progression of HCC cells through transcriptional activation of EMT pathways. Future in vivo studies based on mouse xenograft model will provide additional support to confirm the effects of elevated FFA and DSP loss on enhancing aggressiveness and metastatic potential of HCC. These results may impact HCC therapeutic intervention and drug development, as well as future studies on obesity as a risk factor for cancer tumorigenesis and progression. Citation Format: Aritro Nath, Christina Chan. Free fatty acid induced epithelial to mesenchymal transition (EMT) in hepatocellular carcinoma cells. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C29.