Abstract 24

Gene expression of passaged human articular chondrocytes

Zhen Lin¹, Jiake Xu¹, Craig Willers¹, Jonathan Fitzgerald², David Wood¹, and Ming Hao Zheng¹

¹Department of Orthopaedic Surgery, QEII Medical Centre, University of Western Australia, Perth, WA, ²Biological Engineering Division, Massachusetts Institute of Technology, Cambridge, Massachusetts, U.S.

Abstract:
Autologous chondrocyte implantation (ACI) is considered to be one of the best cellular engineering approaches for the treatment of articular cartilage injury. ACI is able to repair cartilage defects with hyaline-like histology and improved function over conventional surgical treatments. ACI employs in vitro cultured autologous chondrocytes to repair articular cartilage injury, but the biosynthetic profile of cultured chondrocytes has been shown to be altered during in vitro monolayer cultivation. Thus, this study investigated the expression profile of several chondrocyte associated gene clusters in serially passaged human articular chondrocytes by quantitative real-time PCR. The gene clusters include extracellular matrix proteins (aggrecan, type I collagen, type II collagen, type X collagen, fibromodulin, fibronectin, and link protein), matrix proteinases (MMP-1, MMP-3, MMP-9, MMP-13, ADAMTS-4 and ADAMTS-5), proteinase inhibitors (TIMP-1, TIMP-2 and TIMP-3), cytokines (IL-1â, TGFâ, TNFá, and IGF-1), transcription factors (Sox-9, c-fos and c-jun), and intercellular signaling (COX-2, MAPk1, and NOS2). Results obtained by clustering analysis (Euclidean distance) showed that with increasing passage number, the gene expression of the matrix proteins aggrecan, type II collagen, and fibromodulin decreased, while fibronectin and link protein increased. Matrix proteinases, MMP3, 9, 13 and ADAMTS-4, 5, decreased expression especially to passage 6, whilst the proteinase inhibitors, TIMP1, 2, 3, remained constant. Cytokine IL-1 also showed increased expression with serial chondrocyte culture. No significant alternation in TNF-á, TGF-â, IGF-1, or transcriptional factors, Sox-9, cfos, or c-jun expression were observed. These results suggest that the chondrocytic gene expression profile is altered at various degrees with increasing passage number, though the gene expression levels of transcriptional factors which contribute to hyaline cartilage regeneration remain unchanged. This data may prove important for the future development of a more specific and efficacious cultivation technique for human articular chondrocyte-based therapies.

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