Research Article: Lymphotoxin Signaling Is Initiated by the Viral Polymerase in HCV-linked Tumorigenesis

Date Published: March 21, 2013

Publisher: Public Library of Science

Author(s): Yannick Simonin, Serena Vegna, Leila Akkari, Damien Grégoire, Etienne Antoine, Jacques Piette, Nicolas Floc’h, Patrice Lassus, Guann-Yi Yu, Arielle R. Rosenberg, Michael Karin, David Durantel, Urszula Hibner, Stanley M. Lemon.


Exposure to hepatitis C virus (HCV) typically results in chronic infection that leads to progressive liver disease ranging from mild inflammation to severe fibrosis and cirrhosis as well as primary liver cancer. HCV triggers innate immune signaling within the infected hepatocyte, a first step in mounting of the adaptive response against HCV infection. Persistent inflammation is strongly associated with liver tumorigenesis. The goal of our work was to investigate the initiation of the inflammatory processes triggered by HCV viral proteins in their host cell and their possible link with HCV-related liver cancer. We report a dramatic upregulation of the lymphotoxin signaling pathway and more specifically of lymphotoxin-β in tumors of the FL-N/35 HCV-transgenic mice. Lymphotoxin expression is accompanied by activation of NF-κB, neosynthesis of chemokines and intra-tumoral recruitment of mononuclear cells. Spectacularly, IKKβ inactivation in FL-N/35 mice drastically reduces tumor incidence. Activation of lymphotoxin-β pathway can be reproduced in several cellular models, including the full length replicon and HCV-infected primary human hepatocytes. We have identified NS5B, the HCV RNA dependent RNA polymerase, as the viral protein responsible for this phenotype and shown that pharmacological inhibition of its activity alleviates activation of the pro-inflammatory pathway. These results open new perspectives in understanding the inflammatory mechanisms linked to HCV infection and tumorigenesis.

Partial Text

Persistent HCV infection affects about 170 million people worldwide [1] and is one of the most common causes of chronic liver disease [2]. Infected individuals typically suffer from chronic liver inflammation that can last several decades and lead to progressive fibrotic liver that can culminate in hepatic cirrhosis and hepatocellular carcinoma (HCC) (for review see [3]).

Persistent HCV infection is a major cause of chronic liver disease. In particular chronic inflammation, resulting from continuous immune response against infected hepatocytes, is associated with necro-inflammatory changes, liver fibrosis and cirrhosis and HCC development (for review see [45]). The molecular mechanisms involved in initiation and in fuelling of this process, sometimes over very long periods, are still incompletely understood (for review see [7]). In this report we show an upregulation of a pro-inflammatory cytokine, LTβ, and its downstream targets, NF-κB and CXCL10, in HCV-related tumors and in several cellular models based on expression of HCV proteins. The most spectacular alteration of this inflammatory signaling pathway was a very strong upregulation of LTβ expression in nine out of ten liver tumors of transgenic mice with liver-targeted expression of HCV proteins. The one exception (animal 440 in Fig. 2) had high levels of LTβ transcripts and protein both in the tumoral and peri-tumoral liver samples, suggestive of an ongoing inflammation unrelated to HCV. Augmented LTβ expression was also observed in several hepatocyte cell lines harboring the totality or a subset of HCV proteins or solely NS5B, the RNA dependent RNA polymerase. However, it was not detectable in non-tumoral regions of FL-N/35 transgenic livers despite the presence of detectable viral RNA transcripts. In this context it is noteworthy that while efficient cytokine induction by NS5B requires high levels of the enzyme [26], the expression of HCV proteins is typically over 10–100 fold higher in cellular models compared to the transgenic mouse livers analyzed here [46], probably accounting for lack of LT expression in the livers of the FL-N/35 animals. Interestingly, the level of viral RNA in mouse tumors is comparable to that found in peritumoral liver (data not shown). Although we cannot exclude possible variations of NS5B protein expression between the non-tumoral and the tumoral tissues, as well as within individual cells, our data suggest that LT activation might not initiate tumorigenesis, but rather contributes to tumor progression in this animal model. Indeed, strong LTβ expression in 100% of tumors together with complete abrogation of HCV-linked tumorigenesis in animals invalidated for canonical NF-κB signaling, which acts both as an upstream activator and a downstream effector of LT pathway, prompt us to speculate that an autoregulatory loop involving LT and NF-κB might exist in HCV-linked HCC.




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