Alvaro Santos-Laso1, Laura Izquierdo-Sanchez1,2, Pedro M Rodrigues1, Bing Q Huang3, Mikel Azkargorta2,4, Ainhoa Lapitz1, Patricia Munoz-Garrido1, Ander Arbelaiz1, Francisco J Caballero-Camino1,5, Maite G Fernández-Barrena2,6, Raul Jimenez-Agüero1, Josepmaria Argemi6, Tomas Aragon6, Felix Elortza2,4, Marco Marzioni7, Joost P H Drenth8, Nicholas F LaRusso3, Luis Bujanda1,2, Maria J Perugorria1,2,9, Jesus M Banales1,2,9. 1. Department of Liver and Gastrointestinal Diseases, Biodonostia Research Institute - Donostia University Hospital, University of the Basque Country (UPV/EHU), San Sebastian, Spain. 2. National Institute for the Study of Liver and Gastrointestinal Diseases (CIBERehd, Instituto de Salud Carlos III), Madrid, Spain. 3. Division of Gastroenterology and Hepatology, Mayo Clinic, Rochester, MN, USA. 4. Proteomics Platform, CIC bioGUNE, ProteoRed-ISCIII, Bizkaia Science and Technology Park, Derio, Spain. 5. Department of Organic Chemistry I, Center for Innovation in Advanced Chemistry (ORFEO-CINQA), University of the Basque Country (UPV/EHU), San Sebastian, Spain. 6. Division of Hepatology, CIMA-University of Navarra, Pamplona, Spain. 7. Department of Gastroenterology and Hepatology, Università Politecnica delle Marche, Ancona, Italy. 8. Department of Gastroenterology and Hepatology, Radboud University Medical Center, Nijmegen, The Netherlands. 9. IKERBASQUE, Basque Foundation for Science, Bilbao, Spain.
Abstract
BACKGROUND & AIMS: Polycystic liver diseases (PLDs) are genetic disorders characterized by progressive development of multiple biliary cysts. Recently, novel PLD-causative genes, encoding for endoplasmic reticulum (ER)-resident proteins involved in protein biogenesis and transport, were identified. We hypothesized that aberrant proteostasis contributes to PLD pathogenesis, representing a potential therapeutic target. METHODS: ER stress was analysed at transcriptional (qPCR), proteomic (mass spectrometry), morphological (transmission electron microscopy, TEM) and functional (proteasome activity) levels in different PLD models. The effect of ER stress inhibitors [4-phenylbutyric acid (4-PBA)] and/or activators [tunicamycin (TM)] was tested in polycystic (PCK) rats and cystic cholangiocytes in vitro. RESULTS: The expression levels of unfolded protein response (UPR) components were upregulated in liver tissue from PLD patients and PCK rats, as well as in primary cultures of human and rat cystic cholangiocytes, compared to normal controls. Cystic cholangiocytes showed altered proteomic profiles, mainly related to proteostasis (ie synthesis, folding, trafficking and degradation of proteins), marked enlargement of the ER lumen (by TEM) and hyperactivation of the proteasome. Notably, chronic treatment of PCK rats with 4-PBA decreased liver weight, as well as both liver and cystic volumes, of animals under baseline conditions or after TM administration compared to controls. In vitro, 4-PBA downregulated the expression (mRNA) of UPR effectors, normalized proteomic profiles related to protein synthesis, folding, trafficking and degradation and reduced the proteasome hyperactivity in cystic cholangiocytes, reducing their hyperproliferation and apoptosis. CONCLUSIONS: Restoration of proteostasis in cystic cholangiocytes with 4-PBA halts hepatic cystogenesis, emerging as a novel therapeutic strategy.
BACKGROUND & AIMS:Polycystic liver diseases (PLDs) are genetic disorders characterized by progressive development of multiple biliary cysts. Recently, novel PLD-causative genes, encoding for endoplasmic reticulum (ER)-resident proteins involved in protein biogenesis and transport, were identified. We hypothesized that aberrant proteostasis contributes to PLD pathogenesis, representing a potential therapeutic target. METHODS: ER stress was analysed at transcriptional (qPCR), proteomic (mass spectrometry), morphological (transmission electron microscopy, TEM) and functional (proteasome activity) levels in different PLD models. The effect of ER stress inhibitors [4-phenylbutyric acid (4-PBA)] and/or activators [tunicamycin (TM)] was tested in polycystic (PCK) rats and cystic cholangiocytes in vitro. RESULTS: The expression levels of unfolded protein response (UPR) components were upregulated in liver tissue from PLD patients and PCKrats, as well as in primary cultures of human and rat cystic cholangiocytes, compared to normal controls. Cystic cholangiocytes showed altered proteomic profiles, mainly related to proteostasis (ie synthesis, folding, trafficking and degradation of proteins), marked enlargement of the ER lumen (by TEM) and hyperactivation of the proteasome. Notably, chronic treatment of PCKrats with 4-PBAdecreased liver weight, as well as both liver and cystic volumes, of animals under baseline conditions or after TM administration compared to controls. In vitro, 4-PBA downregulated the expression (mRNA) of UPR effectors, normalized proteomic profiles related to protein synthesis, folding, trafficking and degradation and reduced the proteasome hyperactivity in cystic cholangiocytes, reducing their hyperproliferation and apoptosis. CONCLUSIONS: Restoration of proteostasis in cystic cholangiocytes with 4-PBA halts hepatic cystogenesis, emerging as a novel therapeutic strategy.
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