Chondrogenesis by bone marrow-derived mesenchymal stem cells grown in chondrocyte-conditioned medium for auricular reconstruction

Bone marrow-derived mesenchymal stem cells (BMSCs) can be obtained by minimally invasive means and would be a favourable source for cell-based cartilage regeneration. However, controlling the differentiation of the BMSCs towards the desired chondrogenic pathway has been a challenge hampering their application. The major aim of the present study was to determine if conditioned medium collected from cultured auricular chondrocytes could promote chondrogenic differentiation of BMSCs. Auricular chondrocytes were isolated and grown in BMSC standard culture medium (SM) that was collected and used as chondrocyte-conditioned medium (CCM). The BMSCs were expanded in either CCM or SM for three passages. Cells were seeded onto fibrous collagen scaffolds and precultured for 2 weeks with or without transforming growth factor-beta 3 (TGF-β3). After preculture, constructs were implanted subcutaneously in nude mice for 6 and 12 weeks and evaluated with real-time polymerase chain reaction, histology, immunohistochemistry and biochemistry. Real-time polymerase chain reaction results showed upregulation of COL2A1 in the constructs cultured in CCM compared with those in SM. After 12 weeks in vivo, abundant neocartilage formation was observed in the implants that had been cultured in CCM, with or without TGF-β3. In contrast, very little cartilage matrix formation was observed within the SM groups, regardless of the presence of TGF-β3. Osteogenesis was only observed in the SM group with TGF-β3. In conclusion, CCM even had a stronger influence on chondrogenesis than the supplementation of the standard culture medium with TGF-β3, without signs of endochondral ossification. Efficient chondrogenic differentiation of BMSCs could provide a promising alternative cell population for auricular regeneration.