Abstract
The ability to pre-clinically evaluate new cartilage substitution therapies in viable physiological biotribological models, such as the femoral-tibial joint would be advantageous. Methods for osteochondral (OC) plug culture have been developed and the aim of this study was to extend these methods to organ culture of whole femoral condylar and tibial osteochondral tissues.
Porcine femoral condyles and tibial plateau were aseptically dissected. The majority of cancellous bone was removed leaving intact cartilage and a layer of cortical bone. OC plugs were from porcine knee condyles. “Whole joint” tissues and OC plugs were cultured in defined medium and the viability of the cartilage at day 0, 8 or 14 days of culture assessed by XTT assay and LIVE/DEAD staining. Histological analysis (H&E; alcian blue staining) was used to determine cell number and visualise glycosominoglycans (GAGs). GAG levels were quantified in the cartilage using the dimethylene blue assay.
XTT conversion by OC plug cartilage reduced significantly between day 0 and day 8 with no further change between day 8 and 14. GAG levels did not change. “Whole joint” tissue behaved similarly with reduced XTT conversion between days 0 and 8 (femoral only) and days 0 and 14 (femoral and tibial). LIVE/DEAD staining showed the majority of cells remained alive in the mid and deep cartilage zones. There was a band of mainly dead cells in the surface zone, from day 0. There was no change in the GAG levels over the 14 day culture period.
In conclusion, large cuts of femoral and tibial osteochondral tissues were maintained in organ culture for extended periods. Surface zone chondrocytes rapidly lost membrane integrity ex-vivo whereas mid- and deep zone chondrocytes remained viable. It is hypothesised that physiological loading in a novel physically interactive bioreactor will improve the viability and will be the focus of future studies.
