TY - JOUR
T1 - Immortalized liver endothelial cells
T2 - A cell culture model for studies of motility and angiogenesis
AU - Huebert, Robert C.
AU - Jagavelu, Kumaravelu
AU - Liebl, Ann F.
AU - Huang, Bing Q.
AU - Splinter, Patrick L.
AU - Larusso, Nicholas F.
AU - Urrutia, Raul A.
AU - Shah, Vijay H.
N1 - Funding Information:
We acknowledge Helen Hendrickson for technical support, Angela Mathison for assistance with pathway analysis, and Theresa Johnson for secretarial support. This work was supported by grants DK59615-06 (Shah), HL086990 (Shah), DK24031 (LaRusso), P30DK084567 (LaRusso), and the Loan Repayment Program (Huebert) from the National Institutes of Health; by the Hartz Foundation; and the Mayo Foundation.
PY - 2010/12
Y1 - 2010/12
N2 - Hepatic sinusoidal endothelial cells (HSECs) are a unique subpopulation of fenestrated endothelial cells lining the hepatic sinusoids and comprising the majority of endothelial cells within the liver. HSECs not only have important roles in blood clearance, vascular tone, and immunity, but also undergo pathological changes, contributing to fibrosis, angiogenesis, and portal hypertension. There are few cell culture models for in vitro studies of motility and angiogenesis as primary cells are time-consuming to isolate, are limited in number, and often lack features of pathological vasculature. The aim of this study was to generate an immortalized cell line derived from HSECs that mimic pathological vasculature and allows detailed molecular interventions to be pursued. HSECs were isolated from mouse liver using CD31-based immunomagnetic separation, immortalized with SV40 large T-antigen, and subcloned on the basis of their ability to endocytose the acetylated low-density lipoprotein (AcLDL). The resulting cell line, transformed sinusoidal endothelial cells (TSECs), maintains an endothelial phenotype as well as some HSEC-specific features. This is evidenced by typical microscopic features of endothelia, including formation of lamellipodia and filopodia, and a cobblestone morphology of cell monolayers. Electron microscopy showed maintenance of a limited number of fenestrae organized in sieve plates. TSECs express numerous endothelia-specific markers, including CD31 and von Willebrand's factor (vWF), as detected by PCR array, immunoblotting, and immunofluorescence (IF). Functionally, TSECs maintain a number of key endothelial features, including migration in response to angiogenic factors, formation of vascular tubes, endocytosis of AcLDL, and remodeling of extracellular matrix. Their phenotype most closely resembles the pathological neovasculature associated with chronic liver disease, in which cells become proliferative, defenestrated, and angiogenic. Importantly, the cells can be transduced efficiently with viral vectors. TSECs should provide a reproducible cell culture model for high-throughput in vitro studies pertaining to a broad range of liver endothelial cell functions, but likely broader endothelial cell biology as well.
AB - Hepatic sinusoidal endothelial cells (HSECs) are a unique subpopulation of fenestrated endothelial cells lining the hepatic sinusoids and comprising the majority of endothelial cells within the liver. HSECs not only have important roles in blood clearance, vascular tone, and immunity, but also undergo pathological changes, contributing to fibrosis, angiogenesis, and portal hypertension. There are few cell culture models for in vitro studies of motility and angiogenesis as primary cells are time-consuming to isolate, are limited in number, and often lack features of pathological vasculature. The aim of this study was to generate an immortalized cell line derived from HSECs that mimic pathological vasculature and allows detailed molecular interventions to be pursued. HSECs were isolated from mouse liver using CD31-based immunomagnetic separation, immortalized with SV40 large T-antigen, and subcloned on the basis of their ability to endocytose the acetylated low-density lipoprotein (AcLDL). The resulting cell line, transformed sinusoidal endothelial cells (TSECs), maintains an endothelial phenotype as well as some HSEC-specific features. This is evidenced by typical microscopic features of endothelia, including formation of lamellipodia and filopodia, and a cobblestone morphology of cell monolayers. Electron microscopy showed maintenance of a limited number of fenestrae organized in sieve plates. TSECs express numerous endothelia-specific markers, including CD31 and von Willebrand's factor (vWF), as detected by PCR array, immunoblotting, and immunofluorescence (IF). Functionally, TSECs maintain a number of key endothelial features, including migration in response to angiogenic factors, formation of vascular tubes, endocytosis of AcLDL, and remodeling of extracellular matrix. Their phenotype most closely resembles the pathological neovasculature associated with chronic liver disease, in which cells become proliferative, defenestrated, and angiogenic. Importantly, the cells can be transduced efficiently with viral vectors. TSECs should provide a reproducible cell culture model for high-throughput in vitro studies pertaining to a broad range of liver endothelial cell functions, but likely broader endothelial cell biology as well.
KW - angiogenesis
KW - culture
KW - endothelial
KW - liver
KW - motility
KW - sinusoidal
UR - http://www.scopus.com/inward/record.url?scp=78649680847&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=78649680847&partnerID=8YFLogxK
U2 - 10.1038/labinvest.2010.132
DO - 10.1038/labinvest.2010.132
M3 - Article
C2 - 20644520
AN - SCOPUS:78649680847
SN - 0023-6837
VL - 90
SP - 1770
EP - 1781
JO - Laboratory Investigation
JF - Laboratory Investigation
IS - 12
ER -