Role of the hepatitis C virus/Amphiregulin axis in hepatocellular carcinoma-associated angiogenesis

The role of Hepatitis C Virus/Amphiregulin axis in HCC associated Angiogenesis MOHAMMED BENKHEIL, SAM NOPPEN, JAN PAESHUYSE, JOHAN NEYTS, SANDRA LIEKENS REGA INSTITUTE FOR MEDICAL RESEARCH, KU LEUVEN, MINDERBROEDERSSTRAAT 10, B-3000 LEUVEN, BELGIUM Hepatocellular carcinoma (HCC) associated with Hepatitis C Virus (HCV) shows a higher grade of vascularization compared to HCC with other underlying etiologies, suggesting that HCV may play a role in the establishment of a more angiogenic tumor microenvironment (REF). Angiogenesis is a major determinant of tumor malignancy since the newly formed vessels allow tumor progression and metastatic outgrowth. Thus, it is important to gain more insight into the potential role of HCV in this process. Amphiregulin (AREG), an angiogenic factor with prominent expression in the liver has been shown to be upregulated in HCV-infected hepatoma cells (Pei et al, 2011). Here, we further investigated the potential role of AREG in HCV-induced angiogenesis. Using microarray, we found a 16-fold upregulation of AREG in HCV-infected hepatoma Huh7 5.1 cells. This finding was confirmed by RTqPCR and by Western-blot and ELISA on total cellular lysates and conditioned media, respectively. To identify the viral proteins responsible for AREG upregulation, Huh7 5.1 cells were transiently transfected with plasmids encoding individual HCV proteins. This analysis revealed an increased expression of cellular AREG by HCV non-structural proteins. Next, we investigated whether recombinant human AREG (rhAREG) mediates the expression of a selection of angiogenic factors including the vascular endothelial growth factors family members VEGF-A, VEGF-B, VEGF-C and VEGF-D. The latter is also known as c-Fos induced growth factor (FIGF). Treatment of starved Huh7 5.1 with rhAREG resulted in an increased expression of c-Fos and FIGF. An increased expression of FIGF was also found in HCV-infected Huh7 5.1 cells. Our data suggest a significant upregulation of AREG by HCV and in particular by HCV non-structural proteins. Upregulation of FIGF in Huh7 5.1 cells, either after infection with HCV or after treatment with rhAREG, suggests the existence of an HCV/AREG/FIGF angiogenesis axis. Pei R, Chen H, Lu L, Zhu W, Beckebaum S, Cicinnati V, Lu M, Chen X. Hepatitis C virus infection induces the expression of amphiregulin, a factor related to the activation of cellular survival pathways and required for efficient viral assembly. Journal of General Virology. 2011 Oct; 92(Pt 10):2237-48.status: publishe