TY - JOUR
T1 - Isolation and morphologic characterization of bile duct epithelial cells from normal rat liver
AU - Ishii, Motoyasu
AU - Vroman, Benjamin
AU - LaRusso, Nicholas F.
PY - 1989/11
Y1 - 1989/11
N2 - To study directly the functions of the cells that line the bile ducts inside the liver, we developed a new technique for isolating intrahepatic bile duct epithelial cells (IBDECs) from normal rat liver. Parenchymal and nonparenchymal cells were separated from whole liver by enzymatic digestion and mechanical disruption; subpopulations of individual nonparenchymal cells then were isolated by serial counterflow elutriation, isopycnic centrifugation, and immunoaffinity separation with a specific monoclonal antibody against an antigen on the plasma membrane of IBDECs. Using this approach, we isolated 1.2 ± 0.2 × 106 (mean ± SE) viable (>95% trypan blue exclusion) cells, >95% of which were identified as IBDECs by morphologic appearance and specific cytochemical markers. The IBDECs averaged 7.4 ± 0.16 μm in diameter and retained their in situ appearance, including morphologic polarity. They appeared as single cells or as cell doublets attached by tight junctions that excluded ruthenium red. Microvilli were abundant and were restricted to the apical (i.e., luminal) domain of the plasma membrane. Coated pits were observed on both apical and basolateral cell surfaces. Internally, IBDECs contained a well-developed system of organelles, including mitochondria, Golgi, and discrete types of vesicles, such as coated vesicles, multivesicular bodies, and lysosomes. These results indicate that a highly purified suspension of viable, morphologically intact, and polar IBDECs can be prepared from normal rat liver using a novel approach that separates liver cells on the basis of size, density, and specific membrane components. The availability of such a model will allow experimental studies to be performed directly on IBDECs, an approach that has not previously been possible.
AB - To study directly the functions of the cells that line the bile ducts inside the liver, we developed a new technique for isolating intrahepatic bile duct epithelial cells (IBDECs) from normal rat liver. Parenchymal and nonparenchymal cells were separated from whole liver by enzymatic digestion and mechanical disruption; subpopulations of individual nonparenchymal cells then were isolated by serial counterflow elutriation, isopycnic centrifugation, and immunoaffinity separation with a specific monoclonal antibody against an antigen on the plasma membrane of IBDECs. Using this approach, we isolated 1.2 ± 0.2 × 106 (mean ± SE) viable (>95% trypan blue exclusion) cells, >95% of which were identified as IBDECs by morphologic appearance and specific cytochemical markers. The IBDECs averaged 7.4 ± 0.16 μm in diameter and retained their in situ appearance, including morphologic polarity. They appeared as single cells or as cell doublets attached by tight junctions that excluded ruthenium red. Microvilli were abundant and were restricted to the apical (i.e., luminal) domain of the plasma membrane. Coated pits were observed on both apical and basolateral cell surfaces. Internally, IBDECs contained a well-developed system of organelles, including mitochondria, Golgi, and discrete types of vesicles, such as coated vesicles, multivesicular bodies, and lysosomes. These results indicate that a highly purified suspension of viable, morphologically intact, and polar IBDECs can be prepared from normal rat liver using a novel approach that separates liver cells on the basis of size, density, and specific membrane components. The availability of such a model will allow experimental studies to be performed directly on IBDECs, an approach that has not previously been possible.
UR - http://www.scopus.com/inward/record.url?scp=0024441370&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0024441370&partnerID=8YFLogxK
U2 - 10.1016/0016-5085(89)91695-8
DO - 10.1016/0016-5085(89)91695-8
M3 - Article
C2 - 2792660
AN - SCOPUS:0024441370
SN - 0016-5085
VL - 97
SP - 1236
EP - 1247
JO - Gastroenterology
JF - Gastroenterology
IS - 5
ER -