Coculture of equine mesenchymal stem cells and mature equine articular chondrocytes results in improved chondrogenic differentiation of the stem cells
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Bone marrow derived mesenchymal stem cells (MSCs) can be used to repair articularcartilage defects, these cells should be properly stimulated so that they could differentiatemorphologically and hold cellular synthetic features closer to maturely differentiatedchondrocytes. It is well known that tissue specific environment plays an important role incell fate determination. Once improved isolation, proliferation and differentiationprotocols have been developed, the likelihood of spontaneous differentiation of MSCs intodivergent lineages will be reduced, thus increasing their value for cartilage repair. Thepurpose of this study was to improve chondrogenic differentiation of equine MSCs usingcoculture with mature equine articular chondrocytes (ACs), along with the determinationof the effect of adding transforming growth factor (TGF) β1 in the pellet culture system.Following confirmation of multilineage (adipogenic, osteogenic and chondrogenic)differentiation, isolated MSCs, ACs and coculture of both cell types were transferred intopellet culture system in a DMEM-based medium supplemented with or without TGFβ1.Chondrogenic differentiation was evaluated histologically and the relative mRNAexpressions of collagen type 1 α1 (COL1A1), collagen type 2 α1(COL2A1), aggrecan (ACAN)and SRY-box 9 (SOX9) were estimated by quantitative RT-PCR. Cocultured cells showeddiffuse distribution of extracellular matrix (ECM), whereas in chondrocyte pellets it wasmore localized to central regions. Expression of COL2A1, ACAN and SOX9 genes were higherin cocultured pellets when compared to MSCs and ACs-composed pellets. Addition of TGFβ1in chondrogenic differentiating medium did not consistently amplify expression of the abovementioned genes. Differentiation of equine MSCs was enhanced by coculturing in associationwith mature ACs, improving expression of cartilage-specific genes and producing a morehomogeneous production of ECM within the newly formed cocultured cartilage. The use ofthe coculture system could possibly enhance the capacity of MSC-derived chondrocytes tobuild up stable articular cartilage-like constructs, which could play an important role inarticular cartilage repair and regeneration.