ENTACTIN GENE-EXPRESSION IN NORMAL AND FIBROTIC RAT-LIVER AND IN RAT-LIVER CELLS

BACKGROUND: Entactin, a constituent of basement membranes, is a sulfat ed glycoprotein with binding sites for laminin, collagen type IV, fibr onectin, and cell surfaces. As it is known that excess matrix depositi on and sinusoidal basement membrane formation is a central characteris tic of liver fibrogenesis, we investigated whether the entactin gene i s expressed in normal and in damaged rat liver and which cell types ar e able to express this gene. In addition, we were interested in the ce llular origin and time course of laminin synthesis, a matrix protein c losely associated with entactin. EXPERIMENTAL DESIGN: Entactin gene ex pression was analyzed in normal, acutely and chronically damaged rat l ivers (CCl4-model) by immunohistochemistry and in situ detection of sp ecific transcripts. Rat fat-storing cells (FSC) (Ito), hepatocytes, Ku pffer cells, liver endothelial cells, arterial smooth muscle cells (SM C), and skin fibroblasts (SF) were isolated according to standard tech niques. Entactin gene expression in cultured cells was examined by sod ium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of imm unoprecipitates, Northern blot analysis, and immunocytochemistry. RESU LTS: In normal liver, entactin was detected in the vessel walls as con tinuous deposits and in a spot-like fashion along the sinusoids. Entac tin was detectable among the cells of the inflammatory infiltrates of acutely damaged liver and in connective tissue septa, in the walls of newly formed vessels and bile ducts of fibrotic liver. Laminin distrib ution in the vessels was similar, but it was additionally present in t he space of Disse of damaged liver. By in situ hybridization, few enta ctin-positive cells were found in normal liver sections. Strongly posi tive cells were scattered over the injured parenchyma of acutely and c hronically damaged livers. Northern blot analysis of total RNA extract ed from normal and damaged liver tissue showed a distinct increase of entactin specific transcripts during development of fibrosis. Hybridiz ation of total RNA from cultured FSC, hepatocytes, Kupffer cells, endo thelial cells, SMC, and SF revealed entactin specific mRNA in FSC, SMC , SF, and endothelial cells; laminin mRNA was found in FSC and SF. Syn thesis and secretion of both proteins were found in FSC, SMC and SF. E ntactin and laminin gene expression increased in parallel to FSC durin g time in culture. CONCLUSIONS: Among the main liver cells, FSC show t he highest entactin gene expression and might therefore play the domin ant role in the synthesis of this protein in normal and fibrotic liver . However, endothelial cells and liver myofibroblasts could also contr ibute to entactin production.