Culturing Osteochondral Explants Under Rotary Shaking or After Removing Bone Marrow Elements Increases Explant Cellular Viability

Abstract

Background: Reduced viability in the deepest zones of osteochondral allografts (OCAs) can weaken the subchondral interface, potentially increasing the risk of failure. This reduction may result from nutritional imbalances due to uneven media distribution or interference from bone marrow elements. Purpose: To investigate whether culturing OCAs using a rotary shaker or removing the bone marrow elements would increase graft cellular viability. Study Design: Controlled laboratory study. Methods: Bovine osteochondral explants were stored for 28 days at 4°C under 3 different conditions (n = 6 explants per group): static (control group), rotary shaker at 150 rpm (shaker group), and static after removal of bone marrow elements using a Waterpik device (Waterpik group). Chondrocyte viability was assessed using live/dead staining across the entire tissue and in each zone (superficial, middle, deep). Subchondral bone viability was assessed using TUNEL (terminal deoxynucleotidal transferase–mediated biotin–deoxyuridine triphosphate nick-end labeling) staining to detect apoptotic cells. Results: Both shaker (64.2%; P = .010) and Waterpik (65.6%; P = .005) conditions showed significantly higher chondrocyte viability compared with control (49.8%). When samples were analyzed by zone, the shaker and Waterpik groups displayed higher cellular viability at the middle zone (shaker = 60.6%, P < .001; Waterpik = 56.1%, P < .001) and deep zone (shaker = 63.1%, P = .018; Waterpik = 61.5%, P = .025) than the control group (25.6% at middle zone; 32.8% at deep zone). Additionally, shaker (56.7%; P = .018) and Waterpik (51.4%; P = .007) groups demonstrated a lower percentage of apoptotic cells in subchondral bone compared with control (88.0%). No significant differences were observed between the shaker and Waterpik groups in any of the analyses. Conclusion: Both rotary shaking and removal of bone marrow elements during storage of osteochondral explants led to higher chondrocyte viability at the middle and deep zones of the graft compared with the static storage condition. Enhancing nutrition delivery to the graft could improve its quality, potentially improving outcomes of OCA transplantation. Clinical Relevance: The use of a rotary shaker or the removal of bone marrow elements may significantly improve the culture conditions, increasing graft viability and integrity after OCA storage.