Abstract 2309: Hepatitis C virus core protein in combination with interleukin-6 induces epithelial-mesenchymal transition through upregulation of Snail and E2A in human hepatocyte-derived cells

Abstract

Abstract Chronic inflammation gives a great impact on the development of Hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC). Indeed, the interleukin-6 (IL-6)/STAT3 axis, an inflammatory signaling pathway, is known to promote the oncogenic role of the HCV core protein (Core) (Yoshida et al., J Exp Med 2002). In the Core-induced hepatocarcinogenesis in mice, it has recently been suggested that epithelial-mesenchymal transition (EMT) was involved in phenotypic alteration of hepatocytes under TGF-β exposure (Battaglia et al., PLos ONE 2009); however, contribution of either the EMT regulators, including Snail, SLUG, and E2A, or IL-6 to the Core-induced EMT has not yet been investigated. Thus, the AIM of this study was to assess whether Core upregulated the EMT regulator expressions in an IL-6-dependent manner in human hepatocyte-derived cells. Methods: Two human cell lines OUMS-29 and HAK-1A were used in this study. OUMS-29 is a fetus-derived immortalized hepatocyte cell line, which preserves cell polarity and hapatocyte-specific protein production. HAK-1A resembles well-differentiated HCC cells, which has been established from an HCV-related cirrhotic patient (Yano et al., Hepatology 1993). The Core plasmids (myc-tagged and FLAG-tagged) were prepared, and the HCV NS5A plasmid was kindly provided from Dr. Sakamoto. The cells were transfected with the plasmids and were incubated with or without IL-6 (PeproTech Inc., NJ). Protein expression levels in whole cell lysates and nuclear extracts were examined by Western blot analysis. Morphological change at single cell level in the Core-induced EMT was analyzed by immunocytochemistry for E-cadherin and tight junction-associated molecules using a confocal laser scanning microscope (FLUOVIEW FV300; Olympus, Tokyo, Japan). Results: In the Western blot analysis, the protein expression levels of Snail and E2A were increased in both the whole cell lysates and the nuclear extracts from the Core-transfected cells, but not in those samples from the NS5A-transfected cells. The increase in the EMT regulators was augmented in an IL-6-dependent manner. The Core-transfected cells showed a clear decrease in the expression level of occludin after exposure to IL-6. Immunocytochemistry showed the similar decrease in occludin in the Core-transfected cells treated with IL-6. Conclusion: We found that EMT was induced by Core, but not by another HCV-derived protein NS5A. It was suggested that the Core-induced EMT involving Snail and E2A was regulated by the IL-6-mediated inflammatory signaling pathway, which has been shown to promote hapatocarcinogenesis (Naugler et al, Science 2007). Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2309.