Primary cultures of rat hepatocytes as a model system of canalicular development, biliary secretion, and intrahepatic cholestasis. I. Distribution of filipin-cholesterol complexes during de novo formation of bile canaliculi.
Hepatocytes in primary monolayer culture reconstitute structural intact bile canaliculi sealed by tight junctions. Using filipin as a cytochemical marker for cholesterol-like membrane components in conjunction with the techniques of freeze fracture and thin sectioning, we have studied the distribution of cholesterol during the development of the biliary pole of cultured hepatocytes. It was found that the development of bile canaliculi is characterized in its very early stage by huge accumulations of filipin-cholesterol complexes located at distinct domains of the contiguous membrane. They were surrounded by junction formation zones almost devoid of these complexes, in which the alignment of intramembranous particles takes place. Maturation of the bile canaliculi was accompanied by dispersion of cholesterol within the canalicular membrane and its removal by segregation of cholesterol-rich membrane whorls and vesicles into the lumen. Finally, the luminal membranes, and particularly the areas studied with microvilli, contained only very few filipin-cholesterol complexes. In some cases, these seemed to be still arranged in small clusters. These alterations suggest a crucial role of cholesterol-rich membrane domains during initiation of a biliary polarity. On the other hand, cholesterol-poor (thus probably more fluid) areas might be required for the assembly of tight junctions, and appear to constitute the secretory active apical membrane present in the mature bile canaliculus.
D000818:Animals; D001648:Bile Canaliculi / Q000032:analysis / Q000648:ultrastructure*; D001653:Bile Ducts, Intrahepatic / Q000648:ultrastructure*; D002462:Cell Membrane / Q000032:analysis / Q000648:ultrastructure; D002478:Cells, Cultured; D002784:Cholesterol / Q000032:analysis*; D005372:Filipin; D007365:Intercellular Junctions / Q000032:analysis / Q000648:ultrastructure; D008099:Liver / Q000648:ultrastructure*; D008297:Male; D008854:Microscopy, Electron; D008871:Microvilli / Q000032:analysis; D009024:Morphogenesis; D051381:Rats; D011919:Rats, Inbred Strains
D005372:Filipin; D002784:Cholesterol