Cryopreservation of intact articular cartilage (AC) may provide a source of tissues that will enhance long-term results of osteochondral transplantation for large, unconfined joint defects. This study investigated the effects of increasing dimethyl sulfoxide (Me2SO) concentrations on chondrocyte recovery in intact porcine AC after a rapid-cooling cryopreservation technique. Osteochondral dowels 10 mm in diameter were harvested, immersed in various concentrations of Me2SO (1 M, 3 M, 5 M, 6 M, and 7 M), and cryopreserved using a rapid-cooling technique by plunging into liquid nitrogen after initial cooling in an ice-water bath. Fresh controls, toxicity controls, and an experimental group were compared. Slices 70 μm thick (divided into three layers - superficial, middle, and deep) were evaluated after membrane integrity dye staining by fluorescence microscopy. High concentrations of Me2SO (6 M and 7 M) demonstrated significant protection from cryoinjury, likely due to vitrification, but this was compromised by significant cellular toxicity. Low concentrations of Me2SO (1 M and 3 M) exhibited significantly less cellular toxicity but did not protect against cryoinjury. The superficial layer demonstrated significantly increased cellular toxicity compared with the deeper layers at all Me2SO concentrations. Me2SO at 6 M demonstrated the highest recovery of chondrocytes after the cryopreservation procedure, with 42% recovery compared with fresh controls (59% recovery compared with toxicity controls), with the middle layer of the sample demonstrating the optimal balance between cellular toxicity and cryoprotection with an intact cell recovery of 56% (compared with fresh controls). This study demonstrated that concentrations of Me2SO sufficient to result in vitrification have significant potential to successfully cryopreserve intact AC. Further investigation is required to limit potential causes cell loss, including cryoprotectant toxicity, devitrification, and heat transfer.
ASJC Scopus subject areas
- Biomedical Engineering
- General Biochemistry,Genetics and Molecular Biology
- Cell Biology