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Abstract
Introduction: The rational use of allograft in bone grafting applications is hindered by current processing and sterilization techniques which impair the mechanical and biological properties of the graft, resulting in a failure to extract the optimal performance out of the donor tissue. Supercritical fluid (SCF) technology is a novel technique, which is capable of achieving terminal sterilization to regulatory standards, while facilitating delipidation of antigenic factors producing a ‘clean’ graft. Its use is
hindered by a lack of formative information regarding its effect on the mechanical and biological properties of allograft bone. Methods: The effect of SCF treatment on the mechanical properties of cortical bone was characterized by in vitro mechanical testing in quasi-static compression in three orientations, as well as 3pt- and 4pt- bending, torsion and dynamic 3pt-bending. Both elastic and plastic mechanical properties were evaluated and compared to untreated controls and gamma irradiation at a low and ‘standard’ dose. The osteoconductivity and osteoinductivity of SCF treated allograft bone was tested in vivo using a bilateral metaphyseal critical-sized defect model in rabbits and an athymic rat ectopic muscle pocket model respectively. The SCF treated grafts were compared to gamma irradiation and analyzed using radiography, computed tomography, and both qualitative and
quantitative histology and immunohistochemistry for both studies. Results: SCF treatment preserved the mechanical properties of bone under all loading modalities tested. This highlights its benign effect on the structure and interaction of bones constituents under both quasi-static and clinically relevant fatigue loading scenarios. Gamma irradiation had a deleterious dose-dependent effect under all loading modalities, which was most significant in fatigue. These findings indicate that clinicians should be
cautious when using gamma irradiated grafts in load-bearing applications. The in vivo studies showed that both gamma irradiation and SCF treatment maintained the osteoconductive traits of allograft bone, with both grafts able to facilitate healing in a metaphyseal critical-sized defect. DBM treated using the same methods preserved its osteoinductivity, producing new bone ectopically in the established screening bioassay. These results substantiate the assertion from the mechanical investigation that
the structure of the bone was preserved following SCF treatment; allowing the normal attachment and proliferation of cells on both mineralized allograft and in DBM. The preservation of osteoinductivity in DBM indicates that this treatment is also gentle to endogenous growth factors. The extraction properties inherent in this treatment also proved to be an ideal final cleaning step for preparing the graft, enabling the removal of cellular debris and processing artifact, and consequently limiting the severity of the immune response compared to the corresponding gamma irradiated graft. Gamma irradiation resulted in an increase graft resorption in both in vivo investigations. Conclusion: SCF technology has considerable expediency for the processing and sterilization of allograft bone, producing safe and efficacious grafts for use in a range of clinical applications. Relevance: Gamma irradiation impairs the mechanical properties of bone and increases resorption which may inhibit its clinical efficacy. SCF sterilization preserves the mechanical and biological properties of allograft bone, allowing the optimal extraction of the inherent properties in donor tissue, and providing clinicians with a viable alternative to autograft.