P0302 : Constitutive gp130 activation accelerates transformation of proliferating human hepatocytes via increased levels of oxidative stress

Abstract

Background and Aims: Transforming growth factor (TGF)-b is a ubiquitously expressed cytokine with fundamental roles in various aspects of cell physiology. In carcinogenesis, TGF-b signaling plays a dual role. While it suppresses the proliferation of epithelial cells and adenoma cells at early stages by inducing growth arrest and apoptosis, it triggers epithelial to mesenchymal transition (EMT) and gain of metastatic abilities at later stages of carcinoma development. The molecular mechanisms underlying this ’TGF-bswitch’ are only beginning to be unravelled. To mimic the pathophysiological situation as closely as possible, we exposed the cells to TGFb long-term. Through this approach we aimed to identify those cooperating factors and signaling pathways that cause HCC cells to interpret the TGF-b signal in a tumor progressive way. Methods: In vitro comparison of migratory behaviour of various HCC cell lines treated long-term (>10 days) with TGF-b. Analysis of regulatory networks and target genes underlying the TGF-b treatment. Results: HCC cell lines that have undergone EMT secrete TGF-b and show elevated levels of Smad2/3 phosphorylation indicating an autocrine regulatory feedback loop. Inhibition of TGF-b by LY2109 abrogates autocrine stimulation and diminishes the migratory potential of mesenchymal HCC cells. Silencing of either TGF-bR1 or Smad4 indicated the importance of canonical TGF-b/Smad signaling HCC cell migration. Short-term treatment of cells with TGF-b could not improve migratory abilities. Interestingly, long-term TGF-b treatment revealed crucial differences between mesenchymal HCC cell lines. While HLF cells showed an increase in migration when treated with TGF-b for more than 10 days, SNU449 displayed a dramatic reduction in migration. However, both cell lines displayed no modulation in Smad phosphorylation, indicating a change in the utilization of TGF-b signaling in long-term treated SNU449 cells. Conclusions: EMT-transformed HCC cells establish an autocrine TGF-b loop which stimulates migration. However, TGF-b cannot add up to the autocrine loop but causes a different, even opposing reaction, over time. Interpretation of long-term TGF-b signaling, which mimics the patient’s situation more closely, depends on duration and intensity and is controlled by co-acting factors and signaling pathways. P0301 Withdrawn