Activation of the Unfolded Protein Response (UPR) Is Associated with Cholangiocellular Injury, Fibrosis and Carcinogenesis in an Experimental Model of Fibropolycystic Liver Disease

Chaobo Chen,Carmen Berasain,August Vidal,Roger J Davis,Alberto Villanueva,Christian Trautwein,Amaia Lujambio,Agustín Tortajada,Kang Zheng,Rafael Bañares,Leticia Colyn,Tony Bruns,María Arechederra,Yulia A Nevzorova,Hui Ye,Javier Vaquero,Francisco Javier Cubero,Maite G Fernández‐Barrena ,Mohamed Ramadan Mohamed,Hanghang Wu,Raúl Torres-Ruíz ,Johanna Reißing ,Matı́as A Avila ,Eduardo Martı́nez-Naves ,Sandra Rodríguez‐Perales ,Leonard J Nelson,M Ujue Latasa ,María J Iraburu ,María Isabel Peligros

doi:10.3390/cancers14010078

Abstract

Simple SummaryPolycystic liver disease (PLD) is a group of rare disorders that result from structural changes in the biliary tree development in the liver. In the present work, we studied alterations in molecular mechanisms and signaling pathways that might be responsible for these pathologies. We found that activation of the unfolded protein response, a process that occurs in response to an accumulation of unfolded or misfolded proteins in the lumen of the endoplasmic reticulum, as well as the scarring of the liver tissue, contribute to the pathogenesis of PLD and the development of cancer. As a preclinical animal model we have used mutant mice of a specific signaling pathway, the c-Jun N-terminal kinase 1/2 (Jnk1/2). These mice resemble a perfect model for the study of PLD and early cancer development.Fibropolycystic liver disease is characterized by hyperproliferation of the biliary epithelium and the formation of multiple dilated cysts, a process associated with unfolded protein response (UPR). In the present study, we aimed to understand the mechanisms of cyst formation and UPR activation in hepatocytic c-Jun N-terminal kinase 1/2 (Jnk1/2) knockout mice. Floxed JNK1/2 (Jnkf/f) and Jnk∆hepa animals were sacrificed at different time points during progression of liver disease. Histological examination of specimens evidenced the presence of collagen fiber deposition, increased α-smooth muscle actin (αSMA), infiltration of CD45, CD11b and F4/80 cells and proinflammatory cytokines (Tnf, Tgfβ1) and liver injury (e.g., ALT, apoptosis and Ki67-positive cells) in Jnk∆hepa compared with Jnkf/f livers from 32 weeks of age. This was associated with activation of effectors of the UPR, including BiP/GRP78, CHOP and spliced XBP1. Tunicamycin (TM) challenge strongly induced ER stress and fibrosis in Jnk∆hepa animals compared with Jnkf/f littermates. Finally, thioacetamide (TAA) administration to Jnk∆hepa mice induced UPR activation, peribiliary fibrosis, liver injury and markers of biliary proliferation and cholangiocarcinoma (CCA). Orthoallografts of DEN/CCl4-treated Jnk∆hepa liver tissue triggered malignant CCA. Altogether, these results suggest that activation of the UPR in conjunction with fibrogenesis might trigger hepatic cystogenesis and early stages of CCA.

Highlights

JNKs are a family of evolutionarily conserved mitogen-activated protein kinases (MAPKs) activated by tumor necrosis factor (TNF) that play an important role in converting extracellular stimuli into a wide range of cellular responses, including inflammatory response, stress response, cell death, cell differentiation and cell proliferation [1,2]
Alpha-smooth muscle actin protein overexpression was evident at 32 weeks of age in Jnk∆hepa mice (Figure 1C)
Our findings reveal that Jnk∆hepa mice spontaneously develop hepatic histological and molecular traits indicative of liver injury, fibrosis and a remarkable cystogenesis accompanied by early activation of the Unfolded Protein Response (UPR)

Summary

Introduction

JNKs are a family of evolutionarily conserved mitogen-activated protein kinases (MAPKs) activated by tumor necrosis factor (TNF) that play an important role in converting extracellular stimuli into a wide range of cellular responses, including inflammatory response, stress response, cell death, cell differentiation and cell proliferation [1,2]. In order to characterize the combined functions of the JNK genes, using cell-type-specific deletion models is essential. By implementing this strategy, we were the first to report that aged hepatocyte-specific Jnk1/2 (Jnk∆hepa) knockout mice present bile duct hyperplasia. The deletion of Jnk1/2 in an experimental model of chronic liver disease was strongly associated with cell death, cholestasis and cholangiocyte proliferation [1]. A few months later, Manieri and colleagues [6] reported that changes in bile acid metabolism in Jnk∆hepa mice may contribute to cholangiocyte proliferation and hepatoblast maturation, causing bile duct hyperplasia and cholangiocyte injury, which leads to cholangiocarcinoma (CCA) development at late stages. A very recent paper [7] confirmed these findings and identified a molecular link between JNK and RIPK1 as a mechanistic trigger of murine polycystic liver disease (PLD)

Objectives

Methods

Results

Conclusion