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Sinusoidal endothelial cells from guinea pig liver synthesize and secrete cellular fibronectin in vitro
Author(s) -
Rieder Hartmut,
Ramadori Giuliano,
Dienes HansPeter,
Zum Büschenfelde KarlHermann Meyer
Publication year - 1987
Publication title -
hepatology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.488
H-Index - 361
eISSN - 1527-3350
pISSN - 0270-9139
DOI - 10.1002/hep.1840070511
Subject(s) - fibronectin , endothelial stem cell , biology , microbiology and biotechnology , hepatocyte , cell culture , secretion , sodium dodecyl sulfate , in vitro , cell , biochemistry , genetics
Endothelial liver cells were obtained from guinea pig by enzymatic digestion and centrifugal elutriation. Cells were cultured on gelatin and fibronectin pretreated culture vessels. Endothelial cells were characterized by phase‐contrast microscopy, electron microscopy and the presence of Factor VIII‐related antigen. Fibronectin secretion was determined in cell‐free supernatants by a sensitive and specific ELISA and localized on fixed cultured cells by immunofluorescence. [ 35 S ]Methionine endogeneously labeled fibronectin was immunoprecipitated from supernatants and cellular lysates and displayed on sodium dodecyl sulfate polyacrylamide slab gel electrophoresis. After attachment to the culture vessel, one day after plating, endothelial cells start to produce fibronectin as measured by ELISA and demonstrated by immunoprecipitation and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. Secretion of fibronectin increases as cells proliferate to form a confluent monolayer. By immunofluorescence, fibronectin is visualized inside permeabilized cells and as a fibrillar network on the cell surface. Underneath the cell bodies, fibronectin‐positive material is present as short strands. From supernatants and cellular lysates, fibronectin is immunoprecipitated with an apparent M r of about 235,000 obviously larger than plasma fibronectin with an M r of 220,000, which behaves electrophoretically like fibronectin isolated from early hepatocyte cultures. As endothelial cells incorporate [ 3 H]fucose in fibronectin, whereas hepatocytes do not, we conclude that endothelial cells in contrast to hepatocytes produce cellular fibronectin. Endothelial cells, therefore, are probably the cellular source of the fibronectin present in the space of Disse. The significance of this finding with respect to fibrotic liver disease is discussed.