Background/Purpose During Osteoarthritis (OA), chondrocytes loose their initial anabolic properties, undergoing morphological and biochemical modifications similar to those encountered during chondrocyte dedifferentiation. However, the molecular mechanisms underlying this process are poorly known. In order to understand the morphological changes, we interested in the expression of two molecules involved in actin cytoskeletal organization, namely Zyxin-related protein 1 (ZRP-1) and in regulation of cell interaction with the extracellular matrix (ECM), meaning the integrin complex α_vβ₃. Concerning the biochemical changes,  we followed the expression of leptin, an OA-related adipokine, during chondrocytes dedifferentiation. We next assessed pathways potentially involved in its production, namely TGFβ and Wnt/β-catenin, since they are both involved in chondrocytes phenotype maintenance and OA development. 

Methods Human articular cartilage was obtained from macroscopically preserved areas within OA hip after prosthetic surgery. Isolated chondrocytes were cultured in monolayer during 14 days sothat dedifferentiation happen, shifting from round (day1) to fibroblastic-like (day 14) shape. Classical dedifferentiation and hypertrophic markers were assed by qRT PCR, ELISA or western blotting. ZRP-1 was detected by western blotting and α_vβ₃complex was vizualized by immunohistochemistry using LM609 antibody. Components of TGFβ and Wnt/β-catenin were detected by western blotting and involvement of these pathways in leptin production was investigated by stable lentiviral silencing of Smad1 and β-catenin.

Results Chondrocytes monolayer culture induced typical Sox9 and COLIIA1 gene expression lost, as well as COLIA1, Runx-2 and MMP-13 gain. We found that ZRP-1 and α_vβ₃ complex expression came with dedifferentiation. Similarly, leptin was absent in primary chondrocytes whereas present in dedifferentiated chondrocytes, and significantly more under prednisolone. ALK1/ALK5 ratio was shifted during dedifferentiation, from high ALK5 and p-Smad2 expression in primary chondrocytes to high endoglin, ALK1 and p-Smad1/5 expression in dedifferentiated counterpart. Moreover, inactive GSK3β and correspondant active β-catenin were found in dedifferentiated chondrocytes, whereas absent from primary cells. Smad1 and β-catenin stable lentiviral silencing led to a significative decrease in leptin production by dedifferentiated chondrocytes. 

Conclusion ZRP-1, α_vβ₃ integrin complex, leptin expression and its associated signaling might represent novel markers for chondrocyte loss of primary phenotype.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/monolayer-culture-induced-the-expression-of-zyxin-related-protein-1-zrp-1-%ce%b1v%ce%b23-integrin-complex-and-leptin-in-human-articular-chondrocytes/