Amphiregulin contributes to the transformed phenotype of human hepatocellular carcinoma cells.

Hepatocellular carcinoma is a major cause of cancer-related deaths. Current treatments are not effective, and the identification of relevant pathways and novel therapeutic targets are much needed. Increasing evidences point to the activation of the epidermal growth factor receptor (EGFR) as an important mechanism in the development of hepatocarcinoma. We previously described that amphiregulin (AR), a ligand of the EGFR, is not expressed in healthy liver but is up-regulated during chronic liver injury, the background on which most liver tumors develop. Now, we have studied the expression and role of AR in human hepatocarcinoma. AR expression and function was studied in human liver tumors and cell lines. AR is expressed in human hepatocellular carcinoma tissues and cell lines and behaves as a mitogenic and antiapoptotic growth factor for hepatocarcinoma cells. We provide several lines of evidence, including AR silencing by small interfering RNAs and inhibition of amphiregulin by neutralizing antibodies, showing the existence of an AR-mediated autocrine loop that contributes to the transformed phenotype. Indeed, interference with endogenous AR production resulted in reduced constitutive EGFR signaling, inhibition of cell proliferation, anchorage-independent growth, and enhanced apoptosis. Moreover, knockdown of AR potentiated transforming growth factor-beta and doxorubicin-induced apoptosis. Conversely, overexpression of AR in SK-Hep1 cells enhanced their proliferation rate, anchorage-independent growth, drug resistance, and in vivo tumorigenic potential. These observations suggest that AR is involved in the acquisition of neoplastic traits in the liver and thus constitutes a novel therapeutic target in human hepatocarcinoma.

Novel role for amphiregulin in protection from liver injury.

Clinically, the Fas and Fas ligand system plays a central role in the development of hepatocyte apoptosis, a process contributing to a broad spectrum of liver diseases. Therefore, the development of therapies aimed at the inhibition of hepatocyte apoptosis is a major issue. Activation of the epidermal growth factor receptor has been shown to convey survival signals to the hepatocyte. To learn about the endogenous response of epidermal growth factor receptor ligands during Fas-mediated liver injury we investigated the expression of epidermal growth factor, transforming growth factor alpha, heparin-binding epidermal growth factor-like growth factor, betacellulin, epiregulin, and amphiregulin in the liver of mice challenged with Fas-agonist antibody. Amphiregulin expression, barely detectable in healthy liver, was significantly up-regulated. Amphiregulin administration abrogated Fas-mediated liver injury in mice and showed direct anti-apoptotic effects in primary hepatocytes. Amphiregulin activated the Akt and signal transducer and activator of transcription-3 survival pathways, and up-regulated Bcl-xL expression. Amphiregulin knock-out mice showed signs of chronic liver damage in the absence of any noxious treatment, and died faster than wild type mice in response to lethal doses of Fas-agonist antibody. In contrast, these mice were more resistant against sublethal liver damage, supporting the hypothesis that chronic liver injury can precondition hepatocytes inducing resistance to subsequent cell death. These results show that amphiregulin is a protective factor induced in response to liver damage and that it may be therapeutic in liver diseases.

Amphiregulin: an early trigger of liver regeneration in mice.

BACKGROUND AND AIMS: Liver regeneration is a unique response directed to restore liver mass after resection or injury. The survival and proliferative signals triggered during this process are conveyed by a complex network of cytokines and growth factors acting in an orderly manner. Activation of the epidermal growth factor receptor is thought to play an important role in liver regeneration. Amphiregulin is a member of the epidermal growth factor family whose expression is not detectable in healthy liver. We have investigated the expression of amphiregulin in liver injury and its role during liver regeneration after partial hepatectomy. METHODS: Amphiregulin gene expression was examined in healthy and cirrhotic human and rat liver, in rodent liver regeneration after partial hepatectomy, and in primary hepatocytes. The proliferative effects and intracellular signaling of amphiregulin were studied in isolated hepatocytes. The in vivo role of amphiregulin in liver regeneration after partial hepatectomy was analyzed in amphiregulin-null mice. RESULTS: Amphiregulin gene expression is detected in chronically injured human and rat liver and is rapidly induced after partial hepatectomy in rodents. Amphiregulin expression is induced in isolated hepatocytes by interleukin 1beta and prostaglandin E(2), but not by hepatocyte growth factor, interleukin 6, or tumor necrosis factor alpha. We show that amphiregulin behaves as a primary mitogen for isolated hepatocytes, acting through the epidermal growth factor receptor. Finally, amphiregulin-null mice display impaired proliferative responses after partial liver resection. CONCLUSIONS: Our findings indicate that amphiregulin is an early-response growth factor that may contribute to the initial phases of liver regeneration.