Abstracts 1072 to 1346
Authors: Delphine Quelard, Elise Lavergne, Ismail Hendaoui, Harri Elamaa, Estelle Robert, Bruno Turlin, Karim Boudjema, Jean-Pierre Campion, Taina Pihlajaniemi, Bruno Clement, Orlando Musso, Matthew Lawless, Suzanne Norris, Alpha-1 Adrenergic, Receptor Stat3, I Hepatocytes, Chang Han, William Bowen, George Michalopoulos, Tong Wu, Dawidson Gomes, Michele Rodrigues, Marcus Gomez, M Leite, Anton Bennett, Michael Nathanson, Michele Ro- Drigues, Wnt Signaling, Wolfram Goessling, Trista North, Leonard Zon, Leo- Nard Zon, Elisabeth Barfod, Ann Moore, Michael Roe, Steven Lidofsky
Journal: Hepatology (2006)

Collagen XVIII (C18) is a basement membrane component and a plasma protein secreted by hepatocytes.Three variants [NC303, NC493 and NC728] differ in their N-terminal noncollagenous domains (NC).NC303 is the basement membrane form; NC493 and NC728 are alternatively spliced forms driven by a liverspecific promoter.NC728 contains a 235-aa module of unknown function homologous to the secreted frizzled-related protein (SFRP) inhibitors of the Wnt-b-catenin pathway, hence named frizzled.Here, we investigated the biological function of the frizzled module of C18 (FZC18).We analyzed 108 human liver tissue samples by QRT-PCR and immunohistochemistry (12 normal; 35 hepatocellular carcinomas [HCC] and 25 matching non tumor samples; 9 cirrhoses not complicated with HCC; 9 cholestases; 9 benign tumors and 9 colorectal cancer [CRC] metastases).Compared with normal livers, steady-state mRNA levels of NC728 were 6 folds higher both in cirrhoses and in benign tumors and 3 folds higher in cholestases.In metastatic CRC, the expression was similar to that of normal liver.In HCC, NC728 was 3 folds higher than in normal livers.HCCs with pure trabecular pattern (nϭ8) showed 5.8-fold higher expression than other HCCs (nϭ27).Immunoperoxidase showed that NC728 was located in the pericellular matrix of hepatocytes both in cirrhoses and in HCCs, particularly at sites of cell-cell and cell-extracellular matrix interactions.These findings were confirmed by transfection of either FZC18 or full-length NC728 cDNAs in mouse S315 hepatoma cells followed by confocal microscopy analysis with anti-tag antibodies.Then, we tested whether FZC18 is involved in b-catenin signaling by using HepG2 (HCC) and HCT116 (CRC) cell lines harboring activating b-catenin mutations.Assays using a b-catenin responsive reporter showed a dose-dependent inhibition of b-catenin signaling by FZC18 and SFRPs 1; 3 and 5.These effects were associated with a decrease in b-catenin protein stability, cyclin D1 and c-myc protein expression in HCT116 cells and cyclin D1 promoter activity in HepG2 cells.Similarly to SFRPs 1; 3 and 5, transfection of FZC18 cDNA decreased colony formation by 60-90% and induced tumor cell death in HepG2 and HCT116 cells.Moreover, in HEK 293T and in Huh-7 hepatoma cells (wild-type b-catenin), the induction of Wnt signaling by Wnt-1 or Wnt-3a was suppressed by FZC18.Taken together, these data indicate that the frizzled module of C18 is an extracellular inhibitor of Wnt-b-catenin signaling that may quench Wnts in the tumor microenvironment and thus could be involved in liver tissue patterning.