Correlation between free radical content and mechanical properties of ribose-protected irradiation sterilized cortical bone allografts

Abstract

Event Abstract Back to Event Correlation between free radical content and mechanical properties of ribose-protected irradiation sterilized cortical bone allografts Tarik Attia1, 2*, Gagan Minhas2, Xing Ze Lu2*, Marc Grynpas1, 2* and Thomas Willett1, 2, 3* 1 University of Toronto, Institute of Biomaterials and Biomedical Engineering, Canada 2 Mount Sinai Hospital, Lunenfeld Research Institute, Canada 3 University of Toronto, Department of Surgery, Canada Introduction: Human bone transplants are used in the orthopaedic reconstruction of skeletal defects; however, bone allografts have insufficient mechanical properties[1]. In order to ensure the safety of the patient, bone transplants are sterilized to reduce the risk of pathogens. γ-irradiation is the gold standard, as it is an effective method to destroy pathogens. This method, however, causes damage to the bone collagen [2],[3]. Our lab has developed a novel method for improving static mechanical properties, tested, in 3-point-bending by using ribose to modify collagen network connectivity of bone prior/during the irradiation[4]. The purpose of this study is to understand the mechanism by which the ribose treatment protects the mechanical properties of bone from irradiation. We measured the free radical content of the bone and examined its correlation to the mechanical properties of different ribose concentration treatments. Method: Cortical bone beams were cut from human femurs with final dimensions of 2x4x50-60mm. The beams were separated into 5 different treatment groups: normal (N; not irradiated), irradiated (I), and ribose pre-treated + irradiation with a concentration of 0.9M, 1.2M and 2.0M. Subsequently, after irradiation each specimen was placed into an Electron Paramagnetic Resonance (EPR) cavity so that the entire cavity was filled. EPR spectra were collected running at -80oC to ensure stable free radical content. After EPR, 3-point bending tests to fracture were conducted. Results: 30kGy of γ-irradiation created large free radical contents in both the organic and mineral phases of the conventionally irradiated bone. Non-irradiated controls contained barely detectable amounts. The 0.9M ribose treatment resulted in 43%, 1.2M in 40% and the 2.0M in 44% less free radical content immediately after irradiation in comparison to the I-group (p<0.0001)(Fig.1A). No statistically difference between the R-groups were detected. Conventional irradiation greatly decreased the work-to-fracture of the bone by 45% (p<0.001). Ribose-treatment protected the work-to-fracture of the irradiated bone by 30% for 0,9M, 22% for 1.2M and 39% for 2.0M with no statistically difference between the R-groups (Fig.1B). Discussion: The formation of macromolecular free radicals in the organic phase of bone is a known result of scission sites in the collagen and defects in the mineral generated by γ–irradiation, which contributes to the loss of toughness. This study demonstrates that our ribose pre-treatment reduces the amount of free radicals formed and prevents some loss of toughness, independent of the ribose concentration. Significance: This study provides further evidence towards understanding the mechanism by which ribose pre-treatment protects the mechanical properties of irradiation sterilized cortical bone. Mount Sinai Allograft Technologies; Timothy Burrow at University of Toronto, Department of Chemistry; Funding: Canadian Institutes of Health Research (CIHR)