Summary Objective This study investigated a novel approach to induce chondrogenic differentiation of human mesenchymal stem cells (hMSC). We hypothesized that a structured three-dimensional co-culture using hMSC and chondrocytes would provide chondroinductive cues to hMSC without inducing hypertrophy. Method In an effort to promote optimal chondrogenic differentiation of hMSC, we created bilaminar cell pellets (BCPs), which consist of a spherical population of hMSC encased within a layer of juvenile chondrocytes (JC). In addition to histologic analyses, we examined proteoglycan content and expression of chondrogenic and hypertrophic genes in BCPs, JC pellets, and hMSC pellets grown in the presence or absence of TGFβ following 21 days of culture in either growth or chondrogenic media. Results In either growth or chondrogenic media, we observed that BCPs and JC pellets produced more proteoglycan than hMSC pellets treated with TGFβ. BCPs and JC pellets also exhibited higher expression of the chondrogenic genes Sox9, aggrecan, and collagen 2A1, and lower expression of the hypertrophic genes matrix metalloproteinase-13, Runx2, collagen 1A1, and collagen 10A1 than hMSC pellets. Histologic analyses suggest that juvenile chondrocytes promote chondrogenic differentiation of cells in BCPs without hypertrophy. Furthermore, when cultured in hypoxic and inflammatory conditions intended to mimic the injured joint microenvironment, BCPs produced significantly more proteoglycan than either JC pellets or hMSC pellets. Conclusion The BCP co-culture promotes a chondrogenic phenotype without hypertrophy and, relative to pellet cultures of hMSCs or JCs alone, is more resistant to the adverse conditions anticipated at the site of articular cartilage repair.

The percentage of TUNEL positive chondrocytes following intraarticular fracture is much higher than that reported for chronic degenerative conditions such as osteoarthritis and rheumatoid arthritis. These data provide strong evidence that chondrocyte apoptosis is a consequence of intraarticular fracture and suggest that chondrocyte apoptosis may play a particularly significant role in the subsequent development of post-traumatic arthritis.

Intra-articular administration of the caspase inhibitor Z-VAD-fmk effectively blocks chondrocyte PCD following experimental osteochondral injury in this model. Inhibition of chondrocyte PCD rescues chondrocytes that would otherwise die, limiting subsequent cartilage loss. To our knowledge, this study is the first to demonstrate that short-term inhibition of chondrocyte PCD leads to long-term preservation of cartilage in vivo.

Conclusion: this result suggests that BMP signaling inhibits RC muscle atrophy but promotes fatty infiltration.