What Are the Challenges and Complications of Sterilizing Autografts with Liquid Nitrogen for Malignant Bone Tumors? A Preliminary Report

Abstract

Background Reconstruction of defects after resection of malignant bone tumors with liquid nitrogen-sterilized recycled autografts is an alternative to bone allografts and endoprostheses in resource-constrained environments. Most studies reporting favorable outcomes with liquid nitrogen-sterilized autografts for bone reconstruction are geographically restricted to a few countries, and the technical challenges of routinely using liquid nitrogen intraoperatively, especially when using the pedicle freezing technique, has not been documented. Questions/purposes (1) What are the technical challenges of liquid nitrogen sterilization of bone tumors for inexperienced surgeons? (2) What are the complications associated with the procedure? Methods Between May 2017 and October 2019, 88 patients underwent limb salvage procedures for malignant bone tumors of the extremities at our institution. An endoprosthesis was used for reconstruction of the defect following resection in 45% (40 of 88) of these patients, mostly in adults (median age 21 years; range 9 to 68). In the remaining 55% (48 of 88) of patients undergoing biological reconstruction, liquid nitrogen-sterilized autograft was used in 90% (43 of 48), extracorporeal irradiation-sterilized autograft was used in 4% (2 of 48) and allograft was used in 6% (3 of 48). Of the 43 patients receiving liquid nitrogen-sterilized autograft, 5% (2 of 43) were excluded due to loss to follow-up and the remaining 95% (41 of 43) were included for the analysis. Liquid nitrogen-sterilized autograft was the preferred method of reconstruction at our institution during the study period, unless the patient had an indication for prosthesis reconstruction; extracorporeal irradiation-sterilized autograft was used due to resource constraints with liquid nitrogen and allograft was used when patients insisted. All surgical procedures were performed by the same team of trained orthopaedic oncology surgeons. The medical records of the included 41 patients were retrieved using an institutional database search in this retrospective study, and all were used to ascertain technical challenges associated with the operations as well as early (within 3 weeks of the index procedure) and late complications (those occurring 3 weeks or more after surgery). The technical challenges were defined as follows: the quantity of liquid nitrogen to be used; arranging, storing and handling of liquid nitrogen in the operating room, type and size of the container to be used for sterilization, the positioning of the container during pedicle freezing, level of fibular osteotomy for pedicle freezing of tibia, soft tissue protection, limb rotation during pedicle freezing, managing tourniquet time, and any other intraoperative factors with the use of liquid nitrogen for sterilizing the autograft. As our experience with the technique gradually grew, the answers to the above-mentioned factors were determined. Considering the removal of autograft as the endpoint of interest, survival of the autograft was determined by Kaplan-Meier analysis. The median (range) patient age was 14 years (2 to 49), and 54% (22 of 41) were males. Osteosarcoma was the most common diagnosis (68%, [28 of 41]) followed by Ewing’s sarcoma (20%, [8 of 41]). On presentation, 27% of patients (11 of 41) had radiological evidence of pulmonary metastasis. Tumors were seen frequently around the knee (39% [16 of 41] proximal tibia and 22% [9 of 41] distal femur). Before resection 85% (35 of 41) underwent neoadjuvant chemotherapy. Sixty-six percent (27 of 41) underwent pedicle-freezing and the remaining 34% (14 of 41) underwent free-freezing of the tumor segment of the bone. The median (range) duration of surgery was 280 minutes (210 to 510). The patients were followed up for a median (range) duration of 21 months (5 to 30); two patients were lost to follow-up. Results With gradual experience using liquid nitrogen-sterilization over time at our institution, we determined that the following factors helped us in performing liquid nitrogen-sterilization more efficiently. For every operation 15 L to 20 L of unsterilized liquid nitrogen was arranged, 1 or 2 days before the procedure, and stored in industrial-grade cryocylinders in the operating complex. During the procedure, the operating surgeons wore additional plastic aprons under the surgical gowns, surgical goggles, and rubber boots. The staff managing the liquid nitrogen in the operating room wore thermal protective gloves. For most of the pedicle freezing procedures, we used a cylindrical stainless-steel container that was 30 cm in height and 15 cm in diameter, with a narrow opening. The container was kept on a separate moveable cart that was placed next to the operating table at a slightly lower level, and it was wrapped in multiple cotton rolls, plastic sheets, surgical sheets, and a crepe bandage. For pedicle freezing of the tibia, we performed the fibular osteotomy at least 5 cm away from the planned surgical margin, roughly around the axis of rotation of the limb. The soft tissue at the base of the delivered bone segment was dissected for at least 5 cm beyond the planned surgical margin of bone, and was protected with multiple layers of cotton rolls, plastic drapes, a single roll of Esmarch and crepe bandage. The tumor segment was externally rotated during pedicle freezing for all anatomic sites (proximal tibia, distal tibia, proximal humerus, and proximal femur). The tourniquet was inflated just before pedicle freezing to prevent tumor dissemination and not before the initial incision in all pedicle freezing procedures. Thirty-nine percent of patients (16 of 41) experienced complications associated with the procedures, and 15% (6 of 41) underwent revision surgery. Early complications (occurring within 3 weeks of the index procedure) were skin necrosis in four of 16 patients, intraoperative fracture in one of 16, superficial infection in one of 16, and neurapraxia in one of 16 patients. Late complications (occurring 3 weeks or more after surgery) were resorption of the recycled bone in four of 16 patients, nonunion of the osteotomy site in two of 12, delayed union of the osteotomy site in one of 16, collapse of the recycled bone in one of 16, and local recurrence in 1 of 16 patients. Kaplan-Meier survivorship free from removal of autograft at 2 years after surgery was 92% (95% confidence interval 89 to 96). Conclusion Liquid nitrogen-sterilization is an alternative technique that requires some training and experience for the surgeon to become proficient in treating primary malignant bone tumors. Because it is widely available, it may be an option worth exploring in resource-constrained environments, where allografts and endoprostheses cannot be procured. The methods we developed to address the technical challenges will require more study and experience, but we believe these observations will aid others who may wish to use and evaluate liquid nitrogen sterilization of extremity bone sarcomas. Level of Evidence Level IV, therapeutic study.

Do Liquid Nitrogen–treated Tumor-bearing Nerve Grafts Have the Capacity to Regenerate, and Do They Pose a Risk of Local Recurrence? A Study in Rats

“…We reported that liquid nitrogen kills normal and tumor tissues, and developed reconstruction methods using liquid nitrogen-treated tumor-bearing bone after wide excision of malignant bone tumors [23,29,51,53,59,60]. Furthermore, the efficacy of liquid nitrogen treatment for malignant bone tumors has been reported worldwide [19,24,36,37,47,56,58]. However, we only performed hematoxylin-eosin staining to investigate local recurrence in the midportion of the nerve.…”

“…We have developed a reconstruction method using liquid nitrogen-treated tumor-bearing bone after wide excision of malignant bone tumors [23,29,51,53,59,60]. This technique is now used worldwide [19,24,36,37,47,56,58]. We applied this freezing technique to the nerves and thought that using liquid nitrogen-treated tumor-bearing nerves for peripheral nerve reconstruction after wide excisions may provide good functional outcomes.…”

Does A Single Osteotomy Technique for Frozen Autograft (Pedicled Freezing) in Patients With Malignant Bone Tumors of the Long Bones Achieve Union and Local Tumor Control?

“…This technique also obviates the need for a bone bank or special equipment [10,16]. Its osteoinductive and osteoconductive capabilities have been shown to be superior to those of hyperthermia [4,7,16,20]. Further, Xu et al [20] studied the regeneration process in a pedicle frozen bone model in rabbits and the patterns of angiogenesis and revitalization.…”

Engineering Multifunctional Hydrogel With Osteogenic Capacity for Critical-Size Segmental Bone Defect Repair

“…Currently, conventional treatments like bone autografts and allografts and bone cement reconstruction have been used to repair bone defects with primary stability and long-term regeneration; however, these strategies also show some drawbacks ( Bai et al, 2020b ; Butscheidt et al, 2021 ; Chen et al, 2021 ). Among them, bone autografts are considered the gold standard for treatment of bone defects owing to the excellent osteoconduction, osteoinduction, and osteogenesis in bone regeneration ( Hindiskere et al, 2020 ). However, some disadvantages, such as donor-site morbidity, high cost, and long rehabilitation time, limit the application of bone autografts ( Boriani et al, 2017 ; Fahmy-Garcia et al, 2018 ; Lim et al, 2019 ; Han et al, 2020 ).…”