Evaluation of the Complex Transcriptional Topography of Mesenchymal Stem Cell Chondrogenesis for Cartilage Tissue Engineering

Mesenchymal stem cells (MSCs) are a promising cell source for cartilage tissue engineering given their chondrogenic potential. This potential has yet to be fully realized, as the mechanical properties of MSC-based constructs are lower than those of chondrocyte-based constructs cultured identically. The aim of this study was to better understand the transcriptional underpinnings of this functional limitation. Matched chondrocytes and MSCs from three donors were cultured in agarose in a defined medium containing transforming growth factor β3 (TGF-β3). We evaluated the compressive mechanical properties and matrix deposition of maturing constructs over 56 days. Transcriptional differences between the two cell types were assessed on day 0 and 28 via microarray analysis and real-time polymerase chain reaction; differential deposition of matrix molecules was assessed by immunohistochemistry. Although the mechanical and biochemical properties of cell-seeded constructs improved with culture duration, MSC values plateaued at day 28, and remained lower than chondrocyte values. Using microarray analysis, 324 genes were identified as mis-expressed during chondrogenesis. Differential expression of 18 genes was validated, and differential deposition of proteoglycan 4 and TGF-beta-induced 68 kDa protein (TGFBI) was confirmed. Temporal expression profiles of these 18 genes showed that some genes were never expressed (chondromodulin), some were expressed at lower levels (proteoglycan 4), and some were expressed only at later time points (TGFBI) in MSCs compared to chondrocytes. These findings further define the complex transcriptional topography of MSC chondrogenesis, and provide new benchmarks for optimizing the growth of MSC-based engineered cartilage

Surface Zone Articular Chondrocytes Modulate the Bulk and Surface Mechanical Properties of the Tissue-Engineered Cartilage

The central hypothesis of functional tissue engineering is that an engineered construct can serve as a viable replacement tissue in vivo by replicating the structure and function of native tissue. In the case of articular cartilage, this requires the reproduction of the bulk mechanical and surface lubrication properties of native hyaline cartilage. Cartilage tissue engineering has primarily focused on achieving the bulk mechanical properties of native cartilage such as the compressive aggregate modulus and tensile strength. A scaffold-free self-assembling process has been developed that produces engineered cartilage with compressive properties approaching native tissue levels. Thus, the next step in this process is to begin addressing the friction coefficient and wear properties of these engineered constructs. The superficial zone protein (SZP), also known as lubricin or PRG4, is a boundary mode lubricant that is synthesized by surface zone (SZ) articular chondrocytes. Under conditions of high loading and low sliding speeds, SZP reduces friction and wear at the articular surface. The objective of this investigation was to determine whether increasing the proportion of SZ chondrocytes in cartilage constructs, in the absence of external stimuli such as growth factors and mechanical loading, would enhance the secretion of SZP and improve their frictional properties. In this study, cartilage constructs were engineered through a self-assembling process with varying ratios of SZ and middle zone (MZ) chondrocytes (SZ:MZ): 0:100, 25:75, 50:50, 75:25, and 100:0. Constructs containing different ratios of SZ and MZ chondrocytes did not significantly differ in the glycosaminoglycan composition or compressive aggregate modulus. In contrast, tensile properties and collagen content were enhanced in nearly all constructs containing greater amounts of SZ chondrocytes. Increasing the proportion of SZ chondrocytes had the hypothesized effect of improving the synthesis and secretion of SZP. However, increasing the SZ chondrocyte fraction did not significantly reduce the friction coefficient. These results demonstrate that additional factors, such as SZP-binding macromolecules, surface roughness, and adhesion, need to be examined to modulate the lubrication properties of engineered cartilage