title: Designing a fibrotic microenvironment to investigate changes in human liver sinusoidal endothelial cell function.
abstract: The deposition of extracellular matrix (ECM) proteins by hepatic cells during fibrosis leads to the stiffening of the organ and perturbed cellular functions. Changes in the elasticity of liver tissue are manifested by altered phenotype in hepatic cells. We have investigated changes in human liver sinusoidal endothelial cells (hLSECs) that occur as the elastic modulus of their matrix transitions from healthy (6kPa) to fibrotic (36kPa) conditions. We have also investigated the role played by Kupffer cells in the dedifferentiation of hLSECs. We report the complete loss of fenestrae and the expression of CD31 at the surface as a result of increasing elastic moduli. LSECs exhibited a greater number of actin stress fibers and vinculin focal adhesion on the stiffer substrate, as well. A novel finding is that these identical trends can be obtained on soft (6kPa) substrates by introducing an inflamed microenvironment through the addition of Kupffer cells. hLSEC monocultures on 6kPa gels exhibited fenestrae that were 140.7±52.6nm in diameter as well as a lack of surface CD31 expression. Co-culturing hLSECs with rat Kupffer cells (rKCs) on 6kPa substrates, resulted in the complete loss of fenestrae, an increase in CD31 expression and in a well-organized cytoskeleton. These results demonstrate that the increasing stiffness of liver matrices does not solely result in changes in hLSEC phenotype. Even on soft substrates, culturing hLSECs in an inflamed microenvironment can result in their dedifferentiation. Our findings demonstrate the interplay between matrix elasticity and inflammation in the progression of hepatic fibrosis.
authors.affiliation: Department of Chemical Engineering, Virginia Tech, Blacksburg, VA 24061, United States. Electronic address: ford89@vt.edu.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Ford
authors.affiliation: School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA 24061, United States. Electronic address: jaing@vt.edu.
authors.forename: Gaurav
authors.initials: G
authors.lastname: Jain
authors.affiliation: Department of Chemical Engineering, Virginia Tech, Blacksburg, VA 24061, United States; School of Biomedical Engineering and Sciences, Virginia Tech, Blacksburg, VA 24061, United States; ICTAS Center for Systems Biology of Engineered Tissues, Virginia Tech, Blacksburg, VA 24061, United States. Electronic address: padmar@vt.edu.
authors.forename: Padmavathy
authors.initials: P
authors.lastname: Rajagopalan
mesh_terms: D000818:Animals; D054337:Cell Dedifferentiation; D002478:Cells, Cultured; D060833:Cellular Microenvironment; D018920:Coculture Techniques; D042783:Endothelial Cells / Q000378:metabolism* / Q000473:pathology; D005109:Extracellular Matrix / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007728:Kupffer Cells / Q000378:metabolism / Q000473:pathology; D008099:Liver / Q000378:metabolism* / Q000473:pathology; D008103:Liver Cirrhosis / Q000378:metabolism* / Q000473:pathology; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000096:biosynthesis; D051381:Rats
chemical_list: D019408:Platelet Endothelial Cell Adhesion Molecule-1
keywords: Fenestrae; Fibrosis; Kupffer cells; Liver sinusoidal endothelial cells; Matrix stiffness