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Background/Aims : RCAS1, which is recognized by the specific antibody 22-1-1, was first identified as a tumor-associated antigen in gynecological carcinomas. RCAS1 is the ligand of a putative receptor present on lymphocytes, the expression of which is enhanced by lymphocyte activation. RCAS1 inhibits the growth of receptor-bearing cells and induces apoptotic death. Here we examined RCAS1 expression in biliary diseases. Methods : RCAS1 expression was immunohistochemically examined on tissue samples. Apoptotic death was analyzed by DNA fragmentation detection method. RCAS1 production by cell lines was investigated by flow cytometry, enzyme-linked immunosorbent assay, and reverse transcription–polymerase chain reaction. Results : All cholangiocarcinoma cell lines examined clearly expressed RCAS1 at both the protein and RNA level. Immunohistochemically, RCAS1 was expressed in a high percentage of biliary adenocarcinomas (85.9% of intrahepatic cholangiocarcinomas, 96.4% of extrahepatic cholangiocarcinomas and gallbladder carcinomas). Apoptotic tumor-infiltrating lymphocytes could be found in these specimens. RCAS1 expression was frequently detected also in biliary epithelial cells in cases of immune-mediated cholangitis (74.2% in primary biliary cirrhosis, 66.6% in graft-versus-host disease), although the staining pattern for RCAS1 was different compared with cancer cells. Conclusions : RCAS1 is highly expressed not only in cancer cells but also in non-tumor bile duct cells subjected to immune attack.

Backgroundl Aims: In the fibrotic response to liver injury, hepatic stellate cells are activated, leading to the myofibroblastic cell shape, with actin cytoskeletal reorganization and increased extracellular matrix production. The reorganization of actin cytoskeleton suggests that the small GTP binding protein Rho might modulate the process of this myofibroblastic change. The aim of this study was to investigate the role of Rho in the phenotypic changes of hepatic stellate cells. Methods: The phenotypic changes were investigated by the overexpression of Rho regulator, Rho GDI or dominant negative mutant of Rho in mouse hepatic stellate cell line, GRX cells. In activated rat hepatic stellate cells, the effects of microinjection of Botulinus toxin C3, which is the specific inhibitor for Rho, were analyzed. Furthermore, the effect of C3 on the type I collagen accumulation in hepatic stellate cells was investigated. Results: Overexpression of Rho GDI or the dominant negative mutant of Rho caused the shrinkage cell shape and suppressed stress fiber formation. Microinjection of toxin C3 caused a markedly distorted cell shape and the disappearance of stress fibers in rat stellate cells. In addition, C3 strongly suppressed collagen accumulation in activated stellate cells. Conclusions: These results suggest that Rho regulates the actin cytoskeletal reorganization, and may be implicated in the collagen accumulation in activated stellate cells. These findings provide evidence for the role of Rho in the myofibroblastic phenotype in hepatic stellate cells.

Background/Aims : The tumor-associated antigen, RCAS1, has been reported to be expressed in various types of cancer, including cholangiocarcinoma. We measured serum RCAS1 levels in patients with intrahepatic cholangiocellular carcinoma (CCC) and other hepatobiliary diseases, and examined the clinical significance of serum RCAS1 as a tumor marker. Methods : Sera collected from the patients and healthy volunteers were used for ELISA for RCAS1. The values of RCAS1 for CCC patients were compared to those of other tumor marker proteins. Results : Serum RCAS1 levels exceeded the normal limit in a high percentage (73.9%) of CCC patients. The positivity rate was higher than those of CA19-9 and CEA. No correlation was found between the RCAS1 and CA19-9 concentrations. Serum RCAS1 was positive in many cases that were negative for CA19-9. Surgical resection of CCC reduced the RCAS1 level to within the normal range. On the other hand, serum RCAS1 levels were elevated in very few cases of benign hepatobiliary disease. Conclusions : As a tumor marker in CCC, RCAS1 is, at least, of complementary value to CA19-9 and CEA. Measuring serum RCAS1 contributes to the diagnostic accuracy, and is useful for estimating tumor progression or therapeutic effect.

Background/Aims: p160ROCK, a serine/threonine protein kinase, is a direct RhoA target mediating RhoA-induced assembly of focal adhesions and stress fibers. Recently, Rho signaling pathways were reported to play an important role in the activation of rat hepatic stellate cells (HSC). The aim of this study was to investigate the mechanism of action of a p160ROCK-specific inhibitor, Y-27632, on cultured rat HSC. Methods: HSC were isolated from normal rat livers and cultured on fibronectin-coated dishes. The cell morphology and actin cytoskeleton were studied with phase contrast and fluorescence microscopy, respectively. Immunoblot analysis was used to examine phosphorylation of focal adhesion kinase and extracellular signal-regulated kinase, and the expression of cell cycle-associated proteins. HSC proliferation was measured by quantitating the percentage of cells that exhibited nuclear incorporation of 5-bromodeoxyuridine. Type I collagen gene expression and accumulation in HSC culture media were evaluated by Northern blot and enzyme-linked immunosorbent assay, respectively. Results: Y-27632 consistently blocked cell spreading and suppressed RhoA-induced formation of stress fibers in HSC. In addition, Y-27632 inhibited phosphorylation of focal adhesion kinase and extracellular signal-regulated kinase. Cells treated with Y-27632 failed to proliferate, in contrast to untreated spread cells. This shape-dependent block in cell proliferation correlated with a failure to increase cyclin D1 protein level and to down-regulate the cell cycle inhibitor p27. Y-27632 decreased type I collagen gene expression and accumulation in HSC culture media. Conclusions: Our findings indicate that p160ROCK-mediated actin stress fiber assembly is involved in the pathophysiology of hepatic fibrogenesis and suggest that inhibitors of the RhoA-ROCK pathway might be useful therapeutically in liver fibrogenesis.