Cryopreservation of articular cartilage is often used in storage of experimental samples and osteochondral grafts, but the depth-dependence and concentration of glycosaminoglycan (GAG) are significantly altered when cryogenically stored without a cryoprotectant, which will reduce cartilage stiffness and affect osteochondral graft function and long-term viability. This study investigates our ability to detect changes due to cryopreservation in the depth-dependent elastic modulus of osteochondral samples. Using a direct-visualization method requiring minimal histological alterations, unconfined stepwise stress relaxation tests were performed on four fresh (never frozen) and three cryopreserved (−20 °C) canine humeral head osteochondral slices 125 ± 5 μm thick. Applied force was measured and tissue images were taken at the end of each relaxation phase using a 4× objective. Intratissue displacements were calculated by tracking chondrocytes through consecutive images for various intratissue depths. The depth-dependent elastic modulus was compared between fresh and cryopreserved tissue for same-depth ranges using analysis of variance (ANOVA) with Tukey post-test with a 95% confidence interval. Cryopreservation was found to significantly alter the force–displacement profile and reduce the depth-dependent modulus of articular cartilage. Excessive collagen fiber folding occurred at 40–60% relative depth, producing a “black line” in cryopreserved tissue. Force–displacement curves exhibited elongated toe-region in cryopreserved tissue while fresh tissue had nonmeasurable toe-region. Statistical analysis showed significant reduction in the elastic modulus and GAG concentration throughout the tissue between same-depth ranges. This method of cryopreservation significantly reduces the depth-dependent modulus of canine humeral osteochondral samples.
Effects of Cryopreservation on the Depth-Dependent Elastic Modulus in Articular Cartilage and Implications for Osteochondral Grafting
Manuscript received August 22, 2014; final manuscript received November 10, 2014; accepted manuscript posted November 20, 2014; published online March 6, 2015. Assoc. Editor: Ram Devireddy.
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Kahn, D., Les, C., and Xia, Y. (May 1, 2015). "Effects of Cryopreservation on the Depth-Dependent Elastic Modulus in Articular Cartilage and Implications for Osteochondral Grafting." ASME. J Biomech Eng. May 2015; 137(5): 054502. https://doi.org/10.1115/1.4029182
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