Objectives:
Our aim is to identify biological markers that allow diagnosis at a molecular disease stage, facilitating patient stratification and early intervention. Here, we report the development of an in vitro model of human articular cartilage that enables identification of disease indicators in a well-defined context (without co-morbidity).
Methods:
Chondrocyte progenitor cells were isolated from human articular cartilage and used to generate cartilage-like tissue in vitro using a permeable transwell system. The expression of genes characterizing chondrocyte differentiation stages was analyzed using PCR/QPCR. The tissue was characterized for cartilage–specific ECM components using immunohistochemistry and quantitative biochemical methods. Biomechanical properties were assessed using compression testing.
Results:
A protocol for differentiation of human chondrocyte progenitor cells into cartilage-like tissue was established, creating tissue discs of ~6mm diameter and 1mm height. Full-depth differentiation and matrix synthesis occurred over a 28-day period. The tissue had biomechanical properties comparable to articular cartilage. Immunohistochemical analyses showed stratification of the tissue, with collagen I expression restricted to superficial layer and abundance of collagen II and varying glycosaminoglycans in the cartilage matrix proper. Exposure of mature constructs to cyclical loading or to inflammatory cytokines induced mechanoresponse genes and MMP-13/ADAMTS-4 mediated cartilage breakdown, respectively.
Conclusion:
De novo formation of artificial human cartilage tissue in vitro from chondrocyte progenitor cells provides a new tool for analyzing cartilage responses to external stimuli (loading, inflammation, drugs). This model offers continuous tissue supply, a high level of reproducibility and allows for within and between patient analyses.