title: Maintenance of hepatic sinusoidal endothelial cell phenotype in vitro using organ-specific extracellular matrix scaffolds.
abstract: Sinusoidal endothelial cells (SECs) are notoriously difficult to culture in vitro. SECs represent a highly specialized endothelial cell (EC) population, and traditional methods of SEC isolation from the liver initiate a process of SEC dedifferentiation. Acellular extracellular matrix (ECM) scaffolds were investigated in a physiologically relevant in vitro culture model for their ability to maintain SEC phenotype. The cell culture model used SECs only or a coculture of SECs with hepatocytes on ECM substrates derived from the liver (L-ECM), bladder (UBM-ECM), or small intestine submucosa (SIS-ECM). The effect of the ECM substrate upon SEC dedifferentiation was evaluated using scanning electron microscopy (SEM) and confocal microscopy. When SECs alone were cultured on uncoated glass slides, collagen I, UBM-ECM, or SIS-ECM, SECs showed signs of dedifferentiation after 1 day. In contrast, SECs alone cultured on L-ECM maintained their differentiated phenotype for at least 3 days, indicated by the presence of many fenestrations on SEC surface, expression of anti-rat hepatic sinusoidal endothelial cells mouse IgG MoAb (SE-1), and lack of expression of CD31. When SECs were cocultured with hepatocytes on any of the ECM scaffolds, the SECs maintained a near-normal fenestrated phenotype for at least 1 day. However, SEM revealed that the shape, size, frequency, and organization of the fenestrations varied greatly depending on ECM source. At all time points, SECs cocultured with hepatocytes on L-ECM maintained the greatest degree of differentiation. The present study demonstrated that the acellular ECM scaffold derived from the liver maintained SEC differentiation in culture longer than any of the tested substrate materials. The replacement of complex tissues and 3-dimensional organs may require specialized scaffolds to support multiple, functional cell phenotypes.
authors.affiliation: Department of Bioengineering, McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15219, USA.
authors.forename: Tiffany L
authors.initials: TL
authors.lastname: Sellaro
authors.forename: Anjani K
authors.initials: AK
authors.lastname: Ravindra
authors.forename: Donna Beer
authors.initials: DB
authors.lastname: Stolz
authors.forename: Stephen F
authors.initials: SF
authors.lastname: Badylak
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001672:Biocompatible Materials; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology*; D005109:Extracellular Matrix; D008099:Liver / Q000166:cytology*; D010641:Phenotype; D051381:Rats; D017207:Rats, Sprague-Dawley; D023822:Tissue Engineering / Q000295:instrumentation*
chemical_list: D001672:Biocompatible Materials