Effects of SCCO2, Gamma Irradiation, and Sodium Dodecyl Sulfate Treatments on the Initial Properties of Tendon Allografts.

Sterile and decellularized allograft tendons are viable biomaterials used in reconstructive surgeries for dense connective tissue injuries. Established allograft processing techniques including gamma irradiation and sodium dodecyl sulfate (SDS) can affect tissue integrity. Supercritical carbon dioxide (SCCO2) represents a novel alternative that has the potential to decellularize and sterilize tendons with minimized exposure to denaturants, shortened treatment time, lack of toxic residues, and superior tissue penetration, and thus efficacy. This study attempted to develop a single-step hybrid decellularization and sterilization protocol for tendons that involved SCCO2 treatment with various chemical additives. The processed tendons were evaluated with mechanical testing, histology, scanning electron microscopy (SEM), and Fourier-transform infrared (FTIR) spectroscopy. Uniaxial mechanical testing showed that tendons treated with SCCO2 and additive NovaKillTM Gen2 and 0.1% SDS had significantly higher (p < 0.05) ultimate tensile stress (UTS) and Young's modulus compared to gamma-irradiated and standard-SDS-treated tendons. This was corroborated by the ultrastructural intactness of SCCO2-treated tendons as examined by SEM and FTIR spectroscopy, which was not preserved in gamma-irradiated and standard SDS-treated tendons. However, complete decellularization was not achieved by the experimented SCCO2-SDS protocols used in this study. The present study therefore serves as a concrete starting point for development of an SCCO2-based combined sterilization and decellularization protocol for allograft tendons, where additive choice is to be optimized.

Ribose pre-treatment can protect the fatigue life of γ-irradiation sterilized bone.

Structural bone allografts are often sterilized with γ-irradiation to decrease infection risk, which unfortunately degrades the bone collagen connectivity, making the bone weak and brittle. In previous studies, we successfully protected the quasi-static mechanical properties of human cortical bone by pre-treating with ribose, prior to irradiation. This study focused on the quasi-static and fatigue tensile properties of ribose treated irradiated sterilized bone allografts. Seventy-five samples were cut from the mid-shaft diaphysis of human femurs into standardized dog-bone shape geometries for quasi-static and fatigue tensile testing. Specimens were prepared in sets of three adjacent specimens. Each set was made of a normal (N), irradiated (I) and ribose pre-treated + irradiation (R) group. The R group was incubated in a 1.2 M ribose solution before γ-irradiation. The quasi-static tensile and decalcified tests were conducted to failure under displacement control. The fatigue samples were tested under cyclic loading (10 Hz, peak stress of 45MP, minimum-to-maximum stress ratio of 0.1) until failure or reaching 10 million cycles. Ribose pre-treatment significantly improved significantly the mechanical properties of irradiation sterilized human bone in the quasi-static tensile and decalcified tests. The fatigue life of the irradiated group was impaired by 99% in comparison to the normal control. Surprisingly, the R-group has significantly superior properties over the I-group and N-group (p < 0.01, p < 0.05) (> 100%). This study shows that incubating human cortical bone in a ribose solution prior to irradiation can indeed improve the fatigue life of irradiation-sterilized cortical bone allografts.

Autograft or Allograft? Irradiated or Not? A Contrast Between Autograft and Allograft in Anterior Cruciate Ligament Reconstruction: A Meta-analysis.

PURPOSE: To compare the clinical outcomes and adverse events associated with irradiated and nonirradiated allografts in anterior cruciate ligament (ACL) reconstruction. METHODS: PubMed, Web of Science, and EMBASE were searched for randomized controlled trials from January 1990 to March 2018 to compare autograft with allograft in ACL reconstruction. Both objective and subjective outcomes of the function and adverse events were meta-analyzed. Two comparisons were summarized: (1) autograft and nonirradiated allograft and (2) autograft and irradiated allograft. The bias risk was based on the Cochrane Handbook for Systematic Reviews of Interventions. The overall risk ratio or weighted mean difference was calculated using a fixed- or random-effects model. Heterogeneity between studies was evaluated by the Q and the I2 statistics. RESULTS: Eleven trials were included in this review for meta-analysis. A total of 1,172 patients were involved (465 autograft and 461 nonirradiated allograft; 141 autograft and 138 irradiated allograft patients). The average follow-up varied from 2 to >10 years. The mean patient age varied from 22 to 32.8 years. The total failure rate was 2.5%. Our analyses demonstrated better clinical outcomes in autograft than irradiated allograft, which could be observed clearly through the International Knee Documentation Committee score (3.84; 95% confidence interval [CI], 1.93-5.76; P < .0001; I2 = 0%), Lysholm score (2.94; 95% CI, 0.66-5.22; P = .01; I2 = 0%), and Tegner score (0.14; 95% CI, -0.08 to 0.36; P = .22; I2 = 0%) with fewer adverse events 0.20 (95% CI, 0.11-0.39; P < .00001; I2 = 0%). There were no significant differences in autograft and nonirradiated allograft groups (P = .47, P = .27, P = .24, and P = .24, respectively). CONCLUSIONS: Autograft offered greater advantages in functional outcomes and adverse events than irradiated allograft in ACL reconstruction; however, there were no significant differences between autograft and nonirradiated allograft in ACL reconstruction. LEVEL OF EVIDENCE: Level II, meta-analysis of Level I and Level II studies.

Effects of supercritical fluid CO2 and 25 kGy gamma irradiation on the initial mechanical properties and histological appearance of tendon allograft.

Tendon allografts, when autograft options are limited or when obtaining an autograft is not aligned with the patients' best interest, play an important role in tendon and ligament reconstruction. To minimize the risk of infectious disease transmission tissue banks perform screening tests and the allografts cleaned are sterilized. The current study examines and compares the initial mechanical properties and histological appearance of supercritical CO2 (SCCO2)-treated and gamma-irradiated porcine extensor tendons. Thirty intact porcine forelimb extensor tendons randomized equally into three groups: control group, gamma-irradiation group, and SCCO2-treated group. Once treated, histological assessment and histomorphologic measurements were made on the histological sections obtained from each tendon while stiffness and ultimate failure loads were evaluated from tensile testing. Histological evaluation of gamma-irradiated tendons showed significant disruption to the hierarchical morphology of the fascicle bundles, which was not evident in SCCO2-treated specimens. Histomorphologic measurements showed a significant increase for measured dead space (void) between tendon fibrils of the gamma-irradiated group comparing to both control and SCCO2 treated groups (p < 0.01). There was a significant reduction in the ultimate failure load for tendons treated by gamma-irradiation compared to the control group (p < 0.05). No statistically significant difference was detected between control and SCCO2-treated tendons in the ultimate failure load. Stiffness values were not significantly different between three-study groups. This study suggests that while gamma-irradiation has a deleterious effect on mechanical properties of tendon tissue, SCCO2 does not alter the biomechanical properties and the histological structure of porcine extensor tendons.

The effect of ribose pre-treatment of cortical bone on γ-irradiation sterilization effectiveness.

Reconstruction of large skeletal defects is a significant and challenging issue. Tissue banks often use γ-irradiation (15-35 kGy) to sterilize bone allografts, which, however, drastically impairs the post-yield mechanical properties. In previous studies, we reported the development of a method that protects human bone collagen connectivity through ribose crosslinking while still undergoing γ-irradiation. Given these promising results, the next step was to determine if the presence of ribose within the bone tissue would interfere with the effectiveness of the γ-irradiation sterilization against bacteria. This study had two stages. The aim of the first stage was to assess the protective effect of ribose in solution using a Bacillus pumilus spore strip model. The aim of the second stage was to assess the protective effect of ribose (R) on spores within a human cortical bone model in comparison to conventionally irradiated bone (I). Treatment of B. pumilus spore strips with ribose in solution led to temperature-dependent effects on spore viability versus spore strips treated with PBS alone. Ribose solution at 60 °C led to a notable two logs decrease in spore count relative to PBS at 60 °C. In the human bone model, the number of spores in the I and R groups were greatly decreased in comparison to the non-irradiated N group. No spore colonies were detected in the R group (n = 4) whereas two of the four plates of group I formed colonies. This study provides evidence that the method of pre-treating bone with ribose crosslinking prior to irradiation sterilization, while improving irradiation sterilized bone allograft quality, also may improve the effectiveness of the sterilization process.

Does a different dose of gamma irradiation have the same effect on five different types of tendon allografts? - a biomechanical study.

INTRODUCTION: The goals of our study were to evaluate the biomechanical differences between five tendons and the changes in biomechanical properties caused by irradiation. METHODS: Achilles, quadriceps, semitendinosus + gracilis (STG), tibialis anterior (TA) and the peroneus longus (PL) were harvested from 30 donors. Group A contained 50 tendons without gamma irradiation. The groups were irradiated with a dose of 21 kGy (group B 50 tendons) and with a dose of 42 kGy (group C 50 tendons). The grafts were soaked in a radio-protectant solution and frozen at -80 °C. Cyclic loading tests were performed followed by load to failure tests. Young modulus of elasticity, maximum force, strain at tensile strength and strain at rupture were calculated. RESULTS: The Achilles tendons had significantly lower Young modulus than the TA (p = 0.0036) in group A. The Achilles showed significantly lower than PL (p = 0.000042) and TA (p = 0.00142) in group B and C. The quadriceps and the ST (p = 0.0037) provided poorer values than the TA (p = 0.0432) in group C. We found no difference in maximum loads among the tendons in group A. The maximum load of the Achilles and quadriceps showed better results than the PL (p = 0.0016), (p = 0.0018) and the STG (p = 0.0066), (p = 0.0019) in group C. The TA had similar results like the Achilles and quadriceps. DISCUSSION AND CONCLUSIONS: The vulnerability of gamma irradiation of TA was less than Achilles and quadriceps tendons.

Development of a novel method for the strengthening and toughening of irradiation-sterilized bone allografts.

Reconstruction of large skeletal defects is a significant and challenging issue. Bone allografts are often used for such reconstructions. However, sterilizing bone allografts by using γ-irradiation, damages collagen and causes the bone to become weak, brittle and less fatigue resistant. In a previous study, we successfully protected the mechanical properties of human cortical bone by conducting a pre-treatment with ribose, a natural and biocompatible agent. This study focuses on examining possible mechanisms by which ribose might protect the bone. We examined the mechanical properties, crosslinking, connectivity and free radical scavenging potentials of the ribose treatment. Human cortical bone beams were treated with varying concentration of ribose (0.06-1.2 M) and γ-irradiation before testing them in 3-point bending. The connectivity and amounts of crosslinking were determined with Hydrothermal-Isometric-Tension testing and High-Performance-Liquid-Chromatography, respectively. The free radical content was measured using Electron Paramagnetic Resonance. Ribose pre-treatment improved the mechanical properties of irradiation sterilized human bone in a pre-treatment concentration-dependent manner. The 1.2 M pre-treatment provided >100% of ultimate strength of normal controls and protected 76% of the work-to-fracture (toughness) lost in the irradiated controls. Similarly, the ribose pre-treatment improved the thermo-mechanical properties of irradiation-sterilized human bone collagen in a concentration-dependent manner. Greater free radical content and pentosidine content were modified in the ribose treated bone. This study shows that the mechanical properties of irradiation-sterilized cortical bone allografts can be protected by incubating the bone in a ribose solution prior to irradiation.

Effect of a novel sterilization method on biomechanical properties of soft tissue allografts.

PURPOSE: Evaluate allograft tissue commonly used in soft tissue reconstruction to determine whether stiffness and strength were significantly altered after grafts were treated with different sterilization methods. Unprocessed, irradiated, and grafts treated with supercritical CO2 were compared. METHODS: Thirty-eight anterior or posterior tibialis tendons were obtained from a tissue bank (Allograft Innovations, Gainesville, FL). Group I was unprocessed, group II was sterilized with gamma irradiation (20-28 kGy), and group III was sterilized with supercritical CO2. The grafts were pretensioned to 89 N for 300 s. Specimens were then loaded from 50 to 300 N at 0.5 Hz for 250 cycles before being loaded to failure at 50 mm/min. Dependent variables were compared between sterilization groups with one-way ANOVA (P < 0.05) and equivalence trial. RESULTS: There was no significant difference in load to failure or failure stress among groups I, II, and III. Group III resulted in 27-36 % lower stiffness than group I and II. This difference was significant at 1, 10, 50, 100, and 250 cycles. There was no significant difference in stiffness between group I and group II. CONCLUSION: The two sterilization methods tested in this study do not affect allograft strength. The supercritical CO2 sterilization method resulted in significantly lower stiffness than unprocessed and irradiated allografts. However, the stiffness and strength of all groups tested were greater than that of published values of the native intact anterior cruciate ligament (ACL). This study provides previously unpublished mechanical test data on a new sterilization technique that will assist surgeons to decide which allograft to use in ACL reconstruction surgery. LEVEL OF EVIDENCE: III.

Porphyrin-adsorbed Allograft Bone: A Photoactive, Antibiofilm Surface.

BACKGROUND: Allograft bone is commonly used to augment bone stock. Unfortunately, allograft is prone to bacterial contamination and current antimicrobial therapies are inadequate. Photoactivated porphyrins combat bacterial growth by production of reactive oxygen species (ROS); however, to our knowledge, they have not been tested in the setting of allograft bone. QUESTIONS/PURPOSES: We asked: (1) Does 5,10,15,20-tetrakis-(4-aminophenyl)-porphyrin (TAPP) stably adsorb to morselized, mineralized allograft? (2) Does Staphylococcus aureus acquire TAPP from TAPP-allograft? (3) Is TAPP-allograft antibacterial to S. aureus? (4) Is ROS production critical for antimicrobial activity? (5) Does illuminated TAPP-allograft dislodge biofilm? (6) Could other photoactive dyes (TAPP, TMPyP, TSP, THP, and methylene blue) confer antimicrobial properties to allograft? METHODS: TAPP adsorption to allograft (TAPP-allograft), its localization in S. aureus, and TAPP-allograft long-term stability were determined spectrophotometrically. Antimicrobial activity was measured while activated with light or in the dark during incubation with S. aureus or after allograft biofilm formation. Glutathione was added to illuminated TAPP-allograft to quench ROS and antimicrobial activity was determined. Light-dependent antimicrobial activity of other photoactive dyes (TMPyP, TSP, THP, and methylene blue) adsorbed to allograft was also tested. RESULTS: We found (1) porphyrins strongly adhere to bone allograft; and (2) the bacteria are not able to sequester TAPP from the TAPP-allograft; (3) when illuminated, TAPP-allograft is resistant to bacterial adherence; (4) the effects of TAPP are inhibited by the radical scavenger glutathione, indicating ROS-dependent antimicrobial activity; (5) illumination of TAPP-allograft disrupts biofilms; and, (6) other photoactive dyes impede biofilm formation on allograft bone in the presence of light. CONCLUSIONS: Porphyrins stably associate with allograft and are inactive until illuminated. Illuminated TAPP-allograft markedly reduces bacterial colonization, which is restored in the presence of radical scavengers. Finally, illuminated TAPP-allograft disrupts biofilms. CLINICAL RELEVANCE: The findings of this in vitro study suggest that loading bone allograft with biocompatible porphyrins before surgery might allow increased sterility of the allograft during implantation. Future testing in an animal model will determine if these in vitro activities can be used to prevent allograft-based infection in an establishing osteomyelitis.

A Prospective Randomized Study of Anterior Cruciate Ligament Reconstruction With Autograft, γ-Irradiated Allograft, and Hybrid Graft.

PURPOSE: To compare the results of patients who underwent anterior cruciate ligament (ACL) reconstruction with autograft, γ-irradiated allograft, or hybrid graft in a prospective randomized study with a minimum clinical follow-up period of 5 years. METHODS: In this prospective, randomized, comparative study, 102 patients with ACL tears underwent ACL reconstruction with autograft (gracilis and semitendinosus tendons), γ-irradiated allograft (tibialis anterior tendons), or hybrid graft (γ-irradiated tibialis anterior tendon allograft and semitendinosus tendon autograft). Laboratory testing of the erythrocyte sedimentation rate and C-reaction protein level were performed; clinical results were evaluated with the KT-1000 arthrometer (MEDmetric, San Diego, CA), Lachman test, Lysholm score, Tegner activity score, and International Knee Documentation Committee evaluation (both objective and subjective). RESULTS: There were 32 patients in the autograft group, 31 in the hybrid graft group, and 32 in the γ-irradiated allograft group at last follow-up. No differences were found among the 3 groups regarding patient demographic data and the duration from injury to operation (P > .05). The C-reaction protein and erythrocyte sedimentation rate values were statistically higher in the γ-irradiated allograft group than in the other 2 groups on the third, seventh, and fourteenth days (P < .05). No significant differences were found between the autograft and hybrid graft groups (P > .05). The KT-1000 examination showed more anterior laxity in the γ-irradiated allograft group than in the other 2 groups (P < .05). No significant differences in the Lachman test and pivot-shift test findings were found among the 3 groups (P > .05). The Lysholm score, Tegner activity score, and International Knee Documentation Committee evaluation (subjective and objective) showed no differences among the 3 groups (P > .05). CONCLUSIONS: Patients undergoing primary ACL reconstruction with hybrid graft or autograft had satisfactory and similar objective and subjective clinical results. On KT-1000 measurement of anteroposterior knee laxity, both the autograft and hybrid graft groups showed statistically significant differences compared with the γ-irradiated allograft group. LEVEL OF EVIDENCE: Level II, prospective comparative study.

Meniscal allograft sterilisation: effect on biomechanical and histological properties.

Sterilisation of allografts are a crucial step in ensuring safety and viability. Current sterilisation standards such as 25 kGy gamma irradiation (γ) can have adverse effects on the ultrastructure and biomechanical properties of allograft tissue. Supercritical CO2 (SCCO2) technology, represents an improved sterilisation process that potentially preserves tissue properties. This study aimed to test the effect of SCCO2 sterilisation on the biomechanical and histological properties of the meniscus and compare this to the current standard of γ. Thirty-two 18-month old ovine menisci were randomly assigned into three groups for sterilisation (SCCO2, γ and control). After treatment, biomechanical indentation testing (stiffness and stress relaxation) or histological analysis [percentage of void, cells and extracellular matrix (ECM) per slide] was undertaken. Both SCCO2 and gamma groups displayed an increase in stiffness and stress relaxation as compared to control, however, this difference was lesser in samples treated with SCCO2. No significant histological quantitative differences were detected between SCCO2 and control specimens. Gamma-treated samples demonstrated a significant increase in void and decrease in ECM. Interestingly, both treatment groups demonstrated a decreasing mean void and increasing ECM percentage when analysed from outer to inner zones. No significant differences were detected in all-endpoints when analysed by section. SCCO2 sterilisation represents a potential feasible alternative to existing sterilization techniques such as γ.

Tendon allograft sterilized by peracetic acid/ethanol combined with gamma irradiation.

BACKGROUND: Research and clinical applications have demonstrated that the effects of tendon allografts are comparable to those of autografts when reconstructing injured tendons or ligaments, but allograft safety remains problematic. Sterilisation could eliminate or decrease the possibility of disease transmission, but current methods seldom achieve satisfactory sterilisation without affecting the mechanical properties of the tendon. HYPOTHESIS: Peracetic acid-ethanol in combination with low-dose gamma irradiation (PE-R) would inactivate potential deleterious microorganisms without affecting mechanical and biocompatible properties of tendon allograft. STUDY DESIGN: Controlled laboratory design. METHODS: HIV, PPV, PRV and BVDV inactivation was evaluated. After verifying viral inactivation, the treated tendon allografts were characterised by optical microscopy, scanning electron microscopy and tensile testing, and the cytocompatibility was assessed with an MTT assay and by subcutaneous implantation. RESULTS: Effective and efficient inactivation of HIV, PPV, PRV and BVDV was observed. Histological structure and ultrastructure were unchanged in the treated tendon allograft, which also exhibited comparable biomechanical properties and good biocompatibility. CONCLUSION: The preliminary results confirmed our hypothesis and demonstrated that the PE-R tendon allograft has significant potential as an alternative to ligament/tendon reconstruction. CLINICAL RELEVANCE: Tendon allografts have been extensively used in ligament reconstruction and tendon repair. However, current sterilisation methods have various shortcomings, so PE-R has been proposed. This study suggests that PE-R tendon allograft has great potential as an alternative for ligament/tendon reconstruction. WHAT IS KNOWN ABOUT THIS SUBJECT: Sterilisation has been a great concern for tendon allografts. However, most sterilisation methods cannot inactivate viruses and bacteria without impairing the mechanical properties of the tendon allograft. WHAT THIS STUDY ADDS TO EXISTING KNOWLEDGE: Peracetic acid/ethanol with gamma irradiation can effectively inactivate viruses and bacteria. Meanwhile, tendon allografts sterilised by this method maintain their physiological tendon structure, biomechanical integrity and good compatibility.

The effect of sterilization and storage on the viscoelastic properties of human tendon allografts - Continued: Storage for 0 to 4 months.

The role of donor-derived tendons, also known as allografts, in anterior cruciate ligament replacement surgeries is steadily increasing. Before surgery, temporary storage and, in most cases, sterilization are essential. It is, thus, crucial to determine how these procedures alter the grafts' biomechanical properties. The purpose of this research was to analyze the effect of different sterilization methods (native, frozen, frozen + 21 kGy gamma irradiation, frozen + 21 kGy electron beam irradiation) and storage durations (0 to 4 months) on the deformation and creep of two tendon types (tibialis anterior, peroneus longus). 80 tibialis anterior and 83 peroneus longus tendons from 51 human cadavers were included. The samples were removed, placed in a radio-cryoprotectant solution, then slowly cooled, sterilized and stored at -80 °C. All groups were subject to 60 s static creep test with 250 N load. Deformation during the loading phase, creep during static loading, and the ratio of these two were evaluated. Deformation at the end of the loading phase and creep consistently exhibited significantly smaller values in the tibialis anterior compared to the peroneus longus type, as well as in electron beam-sterilized grafts as opposed to gamma beam-sterilized ones. Prolonged storage periods (within 0 to 4 months) resulted in a notable increase in these values, particularly in deformation. Based on the experimental data, the tibialis anterior tendon type and sterilization by gamma beam irradiation are better choices for anterior cruciate ligament reconstruction than the peroneus longus and sterilization by electron beam. Increased storage time affects negatively the evaluated mechanical properties.

Anterior cruciate ligament reconstruction with autografts compared with non-irradiated, non-chemically treated allografts.

PURPOSE: Allograft anterior cruciate ligament (ACL) reconstruction obviates donor site morbidity and may accelerate postoperative recovery. However, allograft use can lead to increased rates of surgical failure, particularly when chemical processing or irradiation is used. Few studies have rigorously evaluated the comparative outcomes of autografts and fresh-frozen allograft tissue for ACL reconstruction. METHODS: We performed a PubMed search to identify and systematically evaluate outcomes of autograft and non-chemically treated non-irradiated allograft tissue in ACL reconstruction between 1980 and 2012. We included studies with Level of evidence of I to III, determinate graft treatment, a minimum of 25 patients per treatment arm, a minimum 2-year follow-up, and selected subjective and objective outcome measures. RESULTS: After the exclusion of 585 citations, we isolated 11 studies for further review. All patients showed improvement in clinical outcome measures and knee stability end points from injury to definitive surgical management. No statistically significant differences were detected between autografts and non-chemically processed non-irradiated allografts in Lysholm scores, International Knee Documentation Committee (IKDC) scores, Lachman examinations, pivot-shift testing, KT-1000 measurements, or failure rates. CONCLUSIONS: Further large-scale, well-designed studies are required to better evaluate the comparative outcomes after fresh-frozen allograft ACL reconstruction. The current study suggests that the results after autograft ACL reconstruction are comparable to those using non-chemically processed nonirradiated allograft tissue. LEVEL OF EVIDENCE: Systematic review of Level I to III studies.

Sterilization of tendon allografts: a method to improve strength and stability after exposure to 50 kGy gamma radiation.

Terminal sterilization of tendon allografts with high dose gamma irradiation has deleterious effects on tendon mechanical properties and stability after implantation. Our goal is to minimize these effects with radio protective methods. We previously showed that radio protection via combined crosslinking and free radical scavenging maintained initial mechanical properties of tendon allografts after irradiation at 50 kGy. This study further evaluates the tissue response and simulated mechanical degradation of tendons processed with radio protective treatment, which involves crosslinking in 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide followed by soaking in an ascorbate/riboflavin-5-phosphate solution. Control untreated and treated tendons were irradiated at 50 kGy and implanted in New Zealand White rabbit knees within the joint capsule for four and 8 weeks. Tendons were also exposed to cyclic loading to 20 N at one cycle per 12 s in a collagenase solution for 150 cycles, followed by tension to failure. Control irradiated tendons displayed increased degradation in vivo, and failed prematurely during cyclic processing at an average of 25 cycles. In contrast, radio protected irradiated tendons displayed greater stability following implantation over 8 weeks, and possessed strength at 59 % of native tendons and modulus equivalent to that of native tendons after cyclic loading in collagenase. These results suggest that radio protective treatment improves the strength and the stability of tendon allografts.

The use of deep frozen and irradiated bone allografts in the reconstruction of tibial plateau fractures.

To investigate the clinical behavior of deep frozen and irradiated bone allografts in the treatment of depressed tibial plateau fractures. Twenty-two patients with a tibial plateau fracture were treated with cancellous bone allografts. The bone allograft preparation process included fresh-freezing at -70 °C for 4 weeks and gamma-irradiation at 25 kGy. All of the patients were followed for 1-2 years. The clinical effects were assessed using the Rasmussen score for tibial head fractures and X-rays. Postoperatively, the average excellent and fair Rasmussen scores were 88.9%. Only one patient developed an infection, with no integration between allograft and recipient bone observed. All of the other bone allografts were incorporated successfully, and no osteoporosis or sclerosis was observed. The frozen and gamma-irradiated bone allograft is a good alternative in the treatment of tibial plateau fractures, which we have shown can integrate with the surrounding host bone.

The effect of sterilization on mechanical properties of soft tissue allografts.

One major concern regarding soft tissue allograft use in surgical procedures is the risk of disease transmission. Current techniques of tissue sterilization, such as irradiation have been shown to adversely affect the mechanical properties of soft tissues. Grafts processed using Biocleanse processing (a proprietary technique developed by Regeneration Technologies to sterilize human tissues) will have better biomechanical characteristics than tissues that have been irradiated. Fifteen pairs of cadaveric Achilles tendon allografts were obtained and separated into three groups of 10 each. Three treatment groups were: Biocleanse, Irradiated, and Control (untreated). Each specimen was tested to determine the biomechanical properties of the tissue. Specimens were cyclically preloaded and then loaded to failure in tension. During testing, load, displacement, and optical strain data were captured. Following testing, the cross sectional area of the tendons was determined. Tendons in the control group were found to have a higher extrinsic stiffness (slope of the load-deformation curve, p = .005), have a higher ultimate stress (force/cross sectional area, p = .006) and higher ultimate failure load (p = .003) than irradiated grafts. Biocleanse grafts were also found to be stiffer than irradiated grafts (p = .014) yet were not found to be statistically different from either irradiated or non-irradiated grafts in terms of load to failure. Biocleanse processing seems to be a viable alternative to irradiation for Achilles tendon allografts sterilization in terms of their biomechanical properties.

Evaluation of safety and efficacy of radiation-sterilized bone allografts in reconstructive oral surgery.

Bone grafting allows reconstruction of the atrophied or destroyed alveolar process. In orthopaedics and traumatology allogeneic grafting has been used to restore defects of osseous tissue for over 60 years. In order to improve safety of the graft recipient, sterilized allogeneic grafts have been use. The aim of the study was to assess the direct and long-term outcomes following augmentation of atrophied alveolar processes with the use of radiation-sterilized allogeneic bone grafts. Sixty-eight patients were surgically treated between 2004 and 2011: 29 underwent open sinus floor elevation, post-extraction alveoli augmentation was performed in 16 subjects and 23 underwent reconstruction of the atrophied alveolar process. Augmentation of bone defects used bone granulate in 63 patients and bone blocks stabilized with titanium screws in 5 patients. PRF membranes collected from the patient's blood were also used in all the procedures. In each of the cases optimal dimensions of the alveolar process were obtained allowing embedment of BIOMET 3I dental implant/-s. In all the patients the defects were successfully restored with implant-supported prostheses. Radiation-sterilized allogeneic bone grafts proved to be safe and effective for the patients and manageable for the surgeon constituting a good alternative to autogeneic material.

Radioprotection provides functional mechanics but delays healing of irradiated tendon allografts after ACL reconstruction in sheep.

Successful protection of tissue properties against ionizing radiation effects could allow its use for terminal sterilization of musculoskeletal allografts. In this study we functionally evaluate Achilles tendon allografts processed with a previously developed radioprotective treatment based on (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide) crosslinking and free radical scavenging using ascorbate and riboflavin, for ovine anterior cruciate ligament reconstruction. Arthroscopic anterior cruciate ligament (ACL) reconstruction was performed using double looped allografts, while comparing radioprotected irradiated and fresh frozen allografts after 12 and 24 weeks post-implantation, and to control irradiated grafts after 12 weeks. Radioprotection was successful at preserving early subfailure mechanical properties comparable to fresh frozen allografts. Twelve week graft stiffness and anterior-tibial (A-T) translation for radioprotected and fresh frozen allografts were comparable at 30 % of native stiffness, and 4.6 and 5 times native A-T translation, respectively. Fresh frozen allograft possessed the greatest 24 week peak load at 840 N and stiffness at 177 N/mm. Histological evidence suggested a delay in tendon to bone healing for radioprotected allografts, which was reflected in mechanical properties. There was no evidence that radioprotective treatment inhibited intra-articular graft healing. This specific radioprotective method cannot be recommended for ACL reconstruction allografts, and data suggest that future efforts to improve allograft sterilization procedures should focus on modifying or eliminating the pre-crosslinking procedure.

Radioprotection and cross-linking of allograft bone in the presence of vitamin E.

Bone allografts are the preferred method for bone augmentation in over 500,000 orthopedic surgical procedures in the US. Sterilization by ionizing radiation is the most effective method of minimizing the bioburden of bone allografts; however, radiation causes chain scission of collagen, resulting in the reduction of the allografts' mechanical strength. In this study, we doped bone allografts with vitamin E as radioprotectant using a novel two-step process to protect the collagen architecture against radiation damage and to preserve the mechanical strength of the construct. In addition, combining the radioprotectant with a cross-linking agent further minimized collagen degradation and further preserved the mechanical strength of the allografts. Both vitamin E and combined vitamin E/genipin-treated allograft were less cytotoxic to both osteoblasts and osteoclasts when compared to irradiated-only allografts. Host bone-allograft unionization was faster in a rat calvaria defect model with vitamin E-treated and combined vitamin E and genipin-treated allograft when compare to irradiated-only allografts. This method can enable the efficient and uniform radioprotective treatment of bone allograft of desired shapes for sterilization with improved mechanical strength and biointegration.