Articular cartilage lines the surfaces of joints and functions to absorb shock and distribute load. To date, few strategies exist for restoring damaged articular surfaces; therefore, cartilage tissue engineering (TE) has emerged as a means to generate functional replacement tissues. To optimize growth and maturation of TE constructs, various methodologies have been employed, including 3D culture, coupled with mechanical stimulation [1] and growth factor supplementation [2]. Recently, studies have shown that temporal application of TGF-β3 enhances the compressive properties and GAG content of chondrocyte-laden hydrogels to near-native levels after the removal of this morphogen [3, 4]. However, as chondrocytes may prove impractical for clinical use, mesenchymal stem cells (MSCs) have been increasingly utilized in cartilage TE. While MSCs are able to undergo chondrogenesis and deposit cartilaginous extracellular matrix (ECM), they have not demonstrated functional parity with chondrocytes [5]. One recent study showed MSCs were able to maintain a chondrocytic phenotype after brief exposure of TGF-β3, though mechanical and biochemical properties were not assessed [6]. In this study, we evaluated these properties in chondrocyte- and MSC-laden hydrogels with transient exposure of TGF-β3 in a chemically defined medium. In addition, we explored the effects of varying seeding density in MSC-laden constructs on functional properties. We hypothesized that transient application of TGF-β3 would improve functional properties of MSC-laden constructs in a seeding density-dependent manner, and that these changes would be marked by differences in cartilaginous gene expression, particularly of enzymes involved in proteoglycan synthesis.

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