Effects of processing and gamma radiation on mechanical properties and organic composition of frozen, freeze-dried and demineralised human cortical bone allograft.

Bone processing and radiation were reported to influence mechanical properties of cortical bones due in part to structural changes and denaturation of collagen composition. This comparative study was to determine effects of bone processing on mechanical properties and organic composition, and to what extent the radiation damaging after each processing. Human femur cortical bones were processed by freezing, freeze-drying and demineralisation and then gamma irradiated at 5, 15, 20, 25 and 50 kGy. In the compression test, freeze drying significantly decreased the Young's Modulus by 15%, while demineralisation reduced further by 90% (P < 0.05) when compared to the freezing. Only demineralisation significantly reduced ultimate strength of bone by 93% (P < 0.05). In the bending test, both freeze drying and demineralisation significantly reduced the ultimate strength and the work to failure. Radiation at 25 kGy showed no effect on compression for ultimate strength in each processing group. However, high dose of 50 kGy significantly reduced bending ultimate strength by 47% in demineralisation group. Alterations in collagen in bones irradiated at 25 and 50 kGy showed by the highest peak of the amide I collagen in the Fourier Transfer Infra-Red spectra indicating more collagen was exposed after calcium was removed in the demineralised bone, however radiation showed no effect on the collagen crosslink. The study confirmed that demineralisation further reduced the ability to resist deformation in response to an applied force in freeze-dried bones due to calcium reduction and collagen composition. Sterilisation dose of 25 kGy has no effect on mechanical properties and collagen composition of the processed human cortical bone.

Deep-Freezing Temperatures During Irradiation Preserves the Compressive Strength of Human Cortical Bone Allografts: A Cadaver Study.

BACKGROUND: Gamma irradiation, which minimizes the risk of infectious disease transmission when human bone allograft is used, has been found to negatively affect its biomechanical properties. However, in those studies, the deep-freezing temperature during irradiation was not necessarily maintained during transportation and sterilization, which may have affected the findings. Prior reports have also suggested that controlled deep freezing may mitigate the detrimental effects of irradiation on the mechanical properties of bone allograft. QUESTION/PURPOSE: Does a controlled deep-freezing temperature during irradiation help preserve the compressive mechanical properties of human femoral cortical bone allografts? METHODS: Cortical bone cube samples, each measuring 64 mm3, were cut from the mid-diaphyseal midshaft of five fresh-frozen cadaver femurs (four male donors, mean [range] age at procurement 42 years [42 to 43]) and were allocated via block randomization into one of three experimental groups (with equal numbers of samples from each donor allocated into each group). Each experimental group consisted of 20 bone cube samples. Samples irradiated in dry ice were subjected to irradiation doses ranging from 26.7 kGy to 27.1 kGy (mean 26.9 kGy) at a deep-freezing temperature below -40°C (the recommended long-term storage temperature for allografts). Samples irradiated in gel ice underwent irradiation doses ranging from 26.2 kGy and 26.4 kGy (mean 26.3 kGy) in a freezing temperature range between -40°C and 0°C. Acting as controls, samples in a third group were not subjected to gamma irradiation. The mechanical properties (0.2% offset yield stress, ultimate compression stress, toughness, and the Young modulus) of samples from each group were subsequently evaluated via axial compression loading to failure along the long axis of the bone. The investigators were blinded to sample group during compression testing. RESULTS: The mean ultimate compression stress (84 ± 27 MPa versus 119 ± 31 MPa, mean difference 35 [95% CI 9 to 60]; p = 0.005) and toughness (3622 ± 1720 kJ/m3 versus 5854 ± 2900 kJ/m3, mean difference 2232 [95% CI 70 to 4394]; p = 0.009) of samples irradiated at a higher temperature range (-40°C to 0°C) were lower than in those irradiated at deep-freezing temperatures (below -40°C). The mean 0.2% offset yield stress (73 ± 28 MPa versus 109 ± 38 MPa, mean difference 36 [95% CI 11 to 60]; p = 0.002) and ultimate compression stress (84 ± 27 MPa versus 128 ± 40 MPa, mean difference 44 [95% CI 17 to 69]; p < 0.001) of samples irradiated at a higher temperature range (-40°C to 0°C) were lower than the nonirradiated control group samples. The mean 0.2% offset yield stress (73 ± 28 MPa versus 101 ± 28 MPa, mean difference 28 [95% CI 3 to 52]; p = 0.02; effect size = 1.0 [95% CI 0.8 to 1.2]) of samples irradiated at higher temperature range (-40°C to 0°C) were no different with the numbers available to those irradiated at deep-freezing temperature. The mean toughness (3622 ± 1720 kJ/m3 versus 6231 ± 3410 kJ/m3, mean difference 2609 [95% CI 447 to 4771]; p = 0.02; effect size = 1.0 [95% CI 0.8 to 1.2]) of samples irradiated at higher temperature range (-40°C to 0°C) were no different with the numbers available to the non-irradiated control group samples. The mean 0.2% offset yield stress, ultimate compression stress, and toughness of samples irradiated in deep-freezing temperatures (below -40°C) were not different with the numbers available to the non-irradiated control group samples. The Young modulus was not different with the numbers available among the three groups. CONCLUSION: In this study, maintenance of a deep-freezing temperature below -40°C, using dry ice as a cooling agent, consistently mitigated the adverse effects of irradiation on the monotonic-compression mechanical properties of human cortical bone tissue. Preserving the mechanical properties of a cortical allograft, when irradiated in a deep-freezing temperature, may have resulted from attenuation of the deleterious, indirect effects of gamma radiation on its collagen architecture in a frozen state. Immobilization of water molecules in this state prevents radiolysis and the subsequent generation of free radicals. This hypothesis was supported by an apparent loss of the protective effect when a range of higher freezing temperatures was used during irradiation. CLINICAL RELEVANCE: Deep-freezing temperatures below -40°C during gamma irradiation may be a promising approach to better retain the native mechanical properties of cortical bone allografts. A further study of the effect of deep-freezing during gamma radiation sterilization on sterility and other important biomechanical properties of cortical bone (such as, tensile strength, fracture toughness, and fatigue) is needed to confirm these findings.

Does bone marrow aspirate help enhance the integration of gamma irradiated allograft bone?

Bone allografts donated by other individuals offer a viable alternative to autograft. Risks of disease transmission are overcome by sterilizing the bone; unfortunately sterilization methods generally affect bone functional properties including osteogenic potential and biomechanical integrity. This study aimed to determine any enhancement effect when gamma sterilised allografts was impregnated with autologous bone marrow in improving the rate and quality of integration in metaphyseal-tibial defects of rabbits. Almost all subjects showed 50% of the defect being covered by new bones by the third week and smaller residual defect size in the treated group at the fifth week. Hounsfield units at the defect site showed increasing healing in all samples, with the treated group having an apparent advantage although insignificant (p > 0.05). In the histopathological score evaluating healing over cortical and cancellous bone at the fracture site showed only slight variations between the groups (p > 0.05). Therefore no enhanced healing by the autologous bone marrow was observed when added to the bone allografts in treating the unicortical defects.

Improving knowledge of doctors and paramedics through effective bone procurement workshop: A cognitive approach.

BACKGROUND: Procurement of bone allograft must be performed by trained personnel. Improper handling and lack of knowledge during bone procurement will lead to contamination hence jeopardizing quality of the procured bones and expose bone recipients to risks of infection in post-operative phase. Bone procurement workshop is the fundamental training programme to enhance skill among personnel who has been or will be involved in bone procurement. This study evaluated the effectiveness of the workshop contents including teaching materials by assessing the knowledge on bone procurement among the participants before and after the workshop. METHODS: Bone procurement workshop was held for 2 days for doctors and paramedics. The knowledge on bone procurement was evaluated in pre- and post-assessments by answering self administration questionnaire before and after the workshop, respectively. RESULTS: A total of 50 participants comprised of doctors and paramedics attended the workshop however only 15 (55.6%) doctors and 12 (44.4%) paramedics completed the assessments. Overall, the mean total score for the post-assessment (61.4%) was significantly higher (p < 0.05) than that of the pre-assessment score (32.2%). The mean values of correct responses for the post-assessment was significantly higher (p < 0.05) than that of the pre-assessment in all five topics given during the workshop. The correct responses for the pre- and the post- assessments in the respective group of the doctors and paramedics were also statistically significant (p < 0.05). In the pre-assessment, the doctors had the highest score in Surgical Approach & Reconstruction (50%) while the paramedics had the highest score in Donor Screening & Selection Criteria (33.3%). In the post-assessment, the doctors had the highest score in Donor Screening & Selection Criteria (70%) while the paramedics in Packaging & Transportation (65.8%). CONCLUSIONS: The assessment managed to show that the workshop contents and teaching materials were effective in improving the cognitive knowledge of the personnel who would get involved in bone procurement under the National Donation Programme.

Performance of cooling materials and their composites in maintaining freezing temperature during irradiation and transportation of bone allografts.

PURPOSE: Bone allografts supplied by University Malaya Medical Centre Bone Bank are sterilized by gamma radiation at 25 kGy in dry ice (DI) to minimize radiation effects. Use of cheaper and easily available cooling materials, gel ice (GI) and ice pack (IP), was explored. Composites of DI and GI were also studied for the use in routine transportations and radiation process. METHODS: (a) Five dummy bones were packed with DI, GI, or IP in a polystyrene box. The bone temperatures were monitored while the boxes were placed at room temperature over 96 h. Durations for each cooling material maintaining freezing temperatures below -40°C, -20°C, and 0°C were obtained from the bone temperature over time profiles. (b) Composites of DI (20, 15, 10, 5, and 0 kg) and GI were used to pack five dummy bones in a polystyrene box. The durations maintaining varying levels of freezing temperature were compared. RESULTS: DI (20 kg) maintained temperature below -40°C for 76.4 h as compared to 6.3 h in GI (20 bags) and 4.0 h in IP (15 packs). Composites of 15DI (15 kg DI and 9 GI bags) and 10DI (10 kg DI and 17 GI bags) maintained the temperature below -40°C for 61 and 35.5 h, respectively. CONCLUSION: Composites of DI and GI can be used to maintain bones in deep frozen state during irradiation, thus avoiding radiation effects on biomechanical properties. Sterile frozen bone allograft with preserved functional properties is required in clinical applications.

Assessing bone banking activities at University of Malaya medical centre.

The main advantage of establishing in-house bone banks is its ability to readily provide allograft bones for local surgeries. Bone procurement activities of our university bone bank during the 10 years of operation were reviewed. Socio-demographic data of donors, types of bone procured, cases of rejected bones and types of allograft bones transplanted are presented. From 179 potential donors, 73 % were accepted with 213 procured bones. Femoral head was the common bone transplanted (45 %), as it was also the most common procured (82 %). Bones were rejected mainly due to non-technical reasons (83 %) rather than positive results of microbiological (13 %) and serological (4 %) tests. Comprehensive data could not be obtained for further analysis due to difficulties in retrieving information. Therefore, quality assurance system was improved to establish more systematic documentations, as the basis of good banking practice with process control hence allowing traceability.

A retrospective study on annual evaluation of radiation processing for frozen bone allografts complying to quality system requirements.

Bone allografts have been used widely to fill up essential void in orthopaedic surgeries. The benefit of using allografts to replace and reconstruct musculoskeletal injuries, fractures or disease has obtained overwhelming acceptance from orthopaedic surgeons worldwide. However, bacterial infection and disease transmission through bone allograft transplantation have always been a significant issue. Sterilization by radiation is an effective method to eliminate unwanted microorganisms thus assist in preventing life threatening allograft associated infections. Femoral heads procured from living donors and long bones (femur and tibia) procured from cadaveric donors were sterilized at 25 kGy in compliance with international standard ISO 11137. According to quality requirements, all records of bone banking were evaluated annually. This retrospective study was carried out on annual evaluation of radiation records from 1998 until 2012. The minimum doses absorbed by the bones were ranging from 25.3 to 38.2 kGy while the absorbed maximum doses were from 25.4 to 42.3 kGy. All the bones supplied by our UMMC Bone Bank were sterile at the required minimum dose of 25 kGy. Our analysis on dose variation showed that the dose uniformity ratios in 37 irradiated boxes of 31 radiation batches were in the range of 1.003-1.251, which indicated the doses were well distributed.