title: Effects of cryopreservation on the depth-dependent elastic modulus in articular cartilage and implications for osteochondral grafting.
abstract: 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.
authors.forename: David
authors.initials: D
authors.lastname: Kahn
authors.forename: Clifford
authors.initials: C
authors.lastname: Les
authors.forename: Yang
authors.initials: Y
authors.lastname: Xia
mesh_terms: D000818:Animals; D001696:Biomechanical Phenomena; D016025:Bone Transplantation; D002358:Cartilage, Articular / Q000166:cytology* / Q000378:metabolism; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism; D015925:Cryopreservation; D004285:Dogs; D055119:Elastic Modulus; D006025:Glycosaminoglycans / Q000378:metabolism; D014022:Tissue Survival
chemical_list: D006025:Glycosaminoglycans