title: Chondrocyte redifferentiation in 3D: the effect of adhesion site density and substrate elasticity.
abstract: To obtain sufficient cell numbers for cartilage tissue engineering with autologous chondrocytes, cells are typically expanded in monolayer culture. As a result, they lose their chondrogenic phenotype in a process called dedifferentiation, which can be reversed upon transfer into a 3D environment. We hypothesize that the properties of this 3D environment, namely adhesion site density and substrate elasticity, would influence this redifferentiation process. To test this hypothesis, chondrocytes were expanded in monolayer and their phenotypical transition was monitored. Agarose hydrogels manipulated to give different RGD adhesion site densities and mechanical properties were produced, cells were incorporated into the gels to induce redifferentiation, and constructs were analyzed to determine cell number and extracellular matrix production after 2 weeks of 3D culture. The availability of adhesion sites within the gels inhibited cellular redifferentiation. Glycosaminoglycan production per cell was diminished by RGD in a dose-dependent manner and cells incorporated into gels with the highest RGD density, remained positive for collagen type I and produced the least collagen type II. Substrate stiffness, in contrast, did not influence cellular redifferentiation, but softer gels contained higher cell numbers and ECM amounts after 2 weeks of culture. Our results indicate that adhesion site density but not stiffness influences the redifferentiation process of chondrocytes in 3D. This knowledge might be used to optimize the redifferentiation process of chondrocytes and thus the formation of cartilage-like tissue.
authors.affiliation: Institute for Biomechanics, ETH Zurich, Zurich, Switzerland.
authors.forename: Elena
authors.initials: E
authors.lastname: Schuh
authors.forename: Sandra
authors.initials: S
authors.lastname: Hofmann
authors.forename: Kathryn
authors.initials: K
authors.lastname: Stok
authors.forename: Holger
authors.initials: H
authors.lastname: Notbohm
authors.forename: Ralph
authors.initials: R
authors.lastname: Müller
authors.forename: Nicole
authors.initials: N
authors.lastname: Rotter
mesh_terms: D000818:Animals; D002417:Cattle; D002448:Cell Adhesion / Q000187:drug effects; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods*; D054337:Cell Dedifferentiation / Q000187:drug effects; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D024042:Collagen Type I / Q000378:metabolism; D024043:Collagen Type II / Q000378:metabolism; D004548:Elasticity / Q000187:drug effects*; D020100:Hydrogels / Q000737:chemistry* / Q000494:pharmacology*; D007150:Immunohistochemistry; D009842:Oligopeptides / Q000494:pharmacology; D012685:Sepharose / Q000737:chemistry; D012710:Serum Albumin, Bovine / Q000378:metabolism; D034421:Sus scrofa
chemical_list: D024042:Collagen Type I; D024043:Collagen Type II; D020100:Hydrogels; D009842:Oligopeptides; D012710:Serum Albumin, Bovine; C047981:arginyl-glycyl-aspartic acid; D012685:Sepharose