Abstract
Background
In the ongoing development of bioresorbable implants, there has been a particular focus on magnesium (Mg)-based alloys. Several Mg alloys have shown promising properties, including a lean, bioresorbable magnesium-zinc-calcium (Mg-Zn-Ca) alloy designated as ZX00. To our knowledge, this is the first clinically tested Mg-based alloy free from rare-earth elements or other elements. Its use in medial malleolar fractures has allowed for bone healing without requiring surgical removal. It is thus of interest to assess the resorption behavior of this novel bioresorbable implant.
Questions/purposes
(1) What is the behavior of implanted Mg-alloy (ZX00) screws in terms of resorption (implant volume, implant surface, and gas volume) and bone response (histologic evaluation) in a sheep model after 13 months and 25 months? (2) What are the radiographic changes and clinical outcomes, including patient-reported outcome measures, at a mean of 2.5 years after Mg-alloy (ZX00) screw fixation in patients with medial malleolar fractures?
Methods
A sheep model was used to assess 18 Mg-alloy (ZX00) different-length screws (29 mm, 24 mm, and 16 mm) implanted in the tibiae and compared with six titanium-alloy screws. Micro-CT was performed at 13 and 25 months to quantify the implant volume, implant surface, and gas volume at the implant sites, as well as histology at both timepoints. Between July 2018 and October 2019, we treated 20 patients with ZX00 screws for medial malleolar fractures in a first-in-humans study. We considered isolated, bimalleolar, or trimalleolar fractures potentially eligible. Thus, 20 patients were eligible for follow-up. However, 5% (one patient) of patients were excluded from the analysis because of an unplanned surgery for a pre-existing osteochondral lesion of the talus performed 17 months after ZX00 implantation. Additionally, another 5% (one patient) of patients were lost before reaching the minimum study follow-up period. Our required minimum follow-up period was 18 months to ensure sufficient time to observe the outcomes of interest. At this timepoint, 10% (two patients) of patients were either missing or lost to follow-up. The follow-up time was a mean of 2.5 ± 0.6 years and a median of 2.4 years (range 18 to 43 months).
Results
In this sheep model, after 13 months, the 29-mm screws (initial volume: 198 ± 1 mm3) degraded by 41% (116 ± 6 mm3, mean difference 82 [95% CI 71 to 92]; p < 0.001), and after 25 months by 65% (69 ± 7 mm3, mean difference 130 [95% CI 117 to 142]; p < 0.001). After 13 months, the 24-mm screws (initial volume: 174 ± 0.2 mm3) degraded by 51% (86 ± 21 mm3, mean difference 88 [95% CI 52 to 123]; p = 0.004), and after 25 months by 72% (49 ± 25 mm3, mean difference 125 [95% CI 83 to 167]; p = 0.003). After 13 months, the 16-mm screws (initial volume: 112 ± 5 mm3) degraded by 57% (49 ± 8 mm3, mean difference 63 [95% CI 50 to 76]; p < 0.001), and after 25 months by 61% (45 ± 10 mm3, mean difference 67 [95% CI 52 to 82]; p < 0.001). Histologic evaluation qualitatively showed ongoing resorption with new bone formation closely connected to the resorbing screw without an inflammatory reaction. In patients treated with Mg-alloy screws after a mean of 2.5 years, the implants were radiographically not visible in 17 of 18 patients and the bone had homogenous texture in 15 of 18 patients. No clinical or patient-reported complications were observed.
Conclusion
In this sheep model, Mg-alloy (ZX00) screws showed a resorption to one-third of the original volume after 25 months, without eliciting adverse immunologic reactions, supporting biocompatibility during this period. Mg-alloy (ZX00) implants were not detectable on radiographs after a mean of 2.5 years, suggesting full resorption, but further studies are needed to assess environmental changes regarding bone quality at the implantation site after implant resorption.
Clinical Relevance
The study demonstrated successful healing of medial malleolar fractures using bioresorbable Mg-alloy screws without clinical complications or revision surgery, resulting in pain-free ankle function after 2.5 years. Future prospective studies with larger samples and extended follow-up periods are necessary to comprehensively assess the long-term effectiveness and safety of ZX00 screws, including an exploration of limitations when there is altered bone integrity, such as in those with osteoporosis. Additional use of advanced imaging techniques, such as high-resolution CT, can enhance evaluation accuracy.
Introduction
The basic goal of a trauma surgeon during fracture fixation is to achieve reduction and stabilize the fractured bone to promote bony healing and enable early mobility, ultimately leading to full functional recovery. A new development in fracture fixation is the use of bioresorbable screws. Unlike traditional surgical steel and titanium alloy screws, these bioresorbable screws are made from magnesium (Mg)-based materials. These screws provide initial stability during the healing process and gradually resorb over time, eliminating the need for screw removal. Recent developments have focused on the search for a suitable alloy [1-4, 6, 14, 21, 22] and testing in animals [7, 19, 24]. Several Mg-based alloys have already been used in patients [13, 16, 23, 25, 27, 28], including the bioresorbable magnesium-zinc-calcium (Mg-Zn-Ca) alloy, designated as ZX00. ZX00 is composed of three elements that are naturally found in the human body, which is a novel approach to Mg-based implants. In contrast, other Mg-based implants usually incorporate alloying systems containing rare-earth elements. When these implants degrade, they are not fully absorbed because they build chelate complexes that form precipitates that may cause long-term side effects [1].
The biocompatibility and resorption of ZX00 has been investigated in mouse and sheep models [7, 20], including proof-of-concept for successful fixation of osteotomies [9]. The first in-human Mg-alloy (ZX00) screw implantations were performed in 2018 and 2019 [8, 10], when feasibility in the clinical environment was demonstrated with a medial malleolar fracture model. Successful healing of the medial malleolar fracture was achieved with Mg-alloy (ZX00) screws, avoiding screw removal. We have published results for 12-week [10] and 1-year [8] follow-up, where Mg-alloy (ZX00) screws were partially degraded. However, knowledge about the state of Mg-alloy (ZX00) screws after the fracture healing period is lacking. Until now, there has been no radiographic monitoring extending beyond 1 year, and the impact of Mg-alloy (ZX00) on local bone tissue beyond 1 year has not been described.
We therefore asked: (1) What is the behavior of implanted Mg-alloy (ZX00) screws in terms of resorption (implant volume, implant surface, and gas volume) and bone response (histologic evaluation) in a sheep model after 13 months and 25 months? (2) What are the radiographic changes and clinical outcomes, including patient-reported outcome measures, at a mean of 2.5 years after Mg-alloy (ZX00) screw fixation in patients with medial malleolar fractures?
Materials and Methods
Background and Overview of the Study Design
The sheep animal model was designed to examine the resorption properties and histologic behavior of Mg-alloy (ZX00) up to 25 months. This study is a follow-up to a study that was approved by the Ethics Committee of the Medical University of Graz (registration number 28 to 071 ex 15/16). Two reports on the study have been published, one with a 12-week follow-up period [10] and another with 1 year of follow-up [8]. Similar animal experiments with shorter observation periods have been performed [7]. However, the findings of this current study are original.
Implant Material and Screws
Mg-alloy (ZX00, a designation according to ASTM B275) screws with a nominal composition of Mg-0.45Zn-0.45Ca (wt%) was fabricated from rods. The production process included the extrusion of Mg-alloy (ZX00) rods by computer numerical control machining. Polycrystalline diamond tools without lubrication were used to manufacture preclinical and clinical study screws to avoid possible contamination and corrosion. The Mg-alloy (ZX00) screws were further cleaned with acetone in an ultrasonic bath and air-dried in a clean room atmosphere, followed by sterilization with gamma irradiation (29.2 kGy), as described by our group [10]. The Mg-alloy (ZX00) has shown promising in vivo behavior in terms of resorption, stability for bone fixation [9], and biocompatibility based on the composition of magnesium, zinc, and calcium [2] without the addition of rare-earth elements or other ions.
In the preclinical sheep study, Mg-alloy (ZX00) screws with a 3.5-mm diameter with three different lengths (29 mm, 24 mm, and 16 mm) were used. As state-of-the-art controls, 18-mm titanium alloy (Ti-6Al-4V) screws with a diameter of 3.5 mm were used. In total, 18 Mg-alloy (ZX00) screws (four 29-mm, four 24-mm, and ten 16-mm screw) and six Ti-6Al-4V screws were implanted into two sheep.
For the clinical study, partially threaded and noncannulated Mg-alloy (ZX00) screws with a diameter of 3.5 mm and a length of 40 mm were used in all patients.
Preclinical Animal Study
Ethical Approval
All animal experiments for this study were approved by the Austrian Federal Ministry for Science and Research under the following permits: BMWFW-66.010/0073-WF/V/3b/2015 and BMWFW-66.010/0049-WF/II/3b/14. The animal experiments were performed in accordance with the “European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes.”
Animal Surgery
The Core Facility of Biomedical Research at the Medical University of Graz provided animal housing and husbandry, including an acclimatization period of 2 weeks. Animals were housed in groups of two with free access to lawn and meadow, with water ad libitum. Female sheep were chosen because they are lighter and easier to handle than male sheep over a longer period. The sample size (two) was chosen based on the 3R rule (replacement, reduction, and refinement) because this was a pilot preclinical study.
Two female domestic sheep (Ovis aries) aged 3 months at the time of implantation were randomly assigned to the 13-month or 25-month model. Screws were surgically implanted with the sheep under general anesthesia and clinical conditions in March 2017. The implantation method was described in our previous work [7, 9]. Both tibiae of each sheep were implanted with a set of Mg-alloy (ZX00) and Ti-6Al-4V screws (Fig. 1). The sheep were euthanized with sodium thiopental (Thiopental Sandoz, Sandoz GmbH), one at 13 months and the other at 25 months postoperatively, to extract the tibiae for further analysis. No adverse events were reported and no animals were excluded during the study period. No blinding protocol was applied during the experiments.
Fig. 1.
This illustration shows the screw location in sheep tibiae. The right leg of both animals had a ZX00 screw in the proximal epiphysis (29-mm monocortical) and proximal metaphysis (24-mm bicortical), and three screws were placed in the diaphysis (16-mm bicortical) as well as one in the distal metaphysis (16-mm monocortical) and distal epiphysis (16-mm monocortical). The left legs of both animals received ZX00 screws in the proximal epiphysis (29-mm monocortical) and proximal metaphysis (24-mm bicortical), and three Ti-6Al-4V screws were placed bicortically in the diaphysis as a control, except for the 25-month animal, in which most distal diaphysis Ti screw was monocortical.
Primary and Secondary Study Outcomes of the Preclinical Study
We primarily aimed to evaluate the Mg-alloy (ZX00) resorption properties (implant volume, surface area, and gas volume) in the sheep model. Therefore, ex vivo, high-resolution micro-CT (Siemens Inveon, Siemens Healthcare Diagnostics GmbH) scans were performed for all bones. Settings used for ex vivo scans were 51.75 µm voxel size with binning setting, and segmentation and 3D evaluation to measure related parameters were performed using Mimics Software (version 21.0 Materialise). For this, bone and implant masks were created with a range threshold from -620 HU to 2166 HU within the Digital Imaging and Communications in Medicine data of ex vivo micro-CT bone scans, separately for all evaluated implants. Further segmentation to acquire implant volume and surface was performed by using “multiple slice editing.” Hydrogen gas, which is a degradation product of magnesium-based implants, was also evaluated as a primary outcome. To evaluate the hydrogen gas volume and surface, masks with a minimum of -1024 HU and a maximum of 299 HU were created. Further segmentation was performed with the “multiple slice editing” and “dynamic region grow” options in the software. The same method was used to evaluate the initial volume and surface of Mg-alloy (ZX00) screws.
The secondary study goal was to perform a histologic analysis to provide information on the bone-to-implant interface and its potential effect on the surrounding bone cellular components at 13 months and 25 months post-implantation. Samples were fixed in 4% formaldehyde, pH 7.0, for 1 week at 4°C, followed by dehydration in ascending grades of ethanol. Infiltration and embedding were done with Technovit 9100 according to the manufacturer’s protocol (Kulzer GmbH). The region of interest for cutting the samples was determined using two-dimensional and three-dimensional images of the bones. Thin ground sections approximately 100-μm thick were created using EXAKT cutting and grinding equipment (EXAKT Advanced Technologies GmbH). Finally, the slices were stained with Laczko-Levai (1975) staining, and scans were performed with the Olympus Microscope scanning system (Olympus Europa SE & Co.). Scanning was performed for one section per implant: in total, four ZX00 and two Ti-6Al-4V screws from the 13-month sheep tibiae and five ZX00 and two Ti-6Al-4V screws from the 25-month sheep tibiae.
Clinical Study
Study Design and Setting
The clinical part of the present study was designed as a first-in-human clinical trial. Briefly, we evaluated the safety and efficacy of bioresorbable Mg-alloy (ZX00) screws in the surgical fixation of medial malleolar fractures in 20 patients. The trial was conducted in accordance with the principles of Good Clinical Practice and the Declaration of Helsinki and was approved by the ethics committee of the University of Graz (registration number 28 to 071 ex 15/16). The current follow-up study investigated the performance of Mg-alloy (ZX00) screws using radiographic and clinical follow-up data at a mean of 2.5 ± 0.6 years postoperatively.
Patients
Between July 2018 and October 2019, 20 patients were treated for medial malleolar fractures with Mg-alloy (ZX00) screws at our institution, a tertiary urban trauma center. Inclusion criteria were met when patients were otherwise healthy and between 18 and 65 years old with either isolated, bimalleolar, or trimalleolar fractures. Written consent was required preoperatively. Exclusion criteria included pathologic fractures, systemic pathologies (such as a diagnosed bone disease, kidney disease, or diabetes mellitus), polytrauma, and pregnant or breastfeeding women. Of the 20 initial patients considered for follow-up, one female patient discontinued participation 17 months after a second ankle surgery to address an old osteochondral lesion that was asymptomatic around the time of the medial malleolar fracture and Mg-alloy (ZX00) implantation [11]. Another female patient was lost to follow-up; she was unreachable 12 weeks after surgery [8]. Thus, the study included 18 patients with complete radiographic and clinical follow-up, with a mean follow-up duration of 2.5 ± 0.6 years and a median follow-up of 2.4 years (range 18 to 43 months).
Descriptive Data
There were 11 men and nine women, with a mean age of 40 ± 14 years. The injuries comprised three isolated medial malleolar fractures, two bimalleolar fractures, 10 trimalleolar fractures, and five Maisonneuve fractures.
Surgery
The surgical procedure included open reduction of the medial malleolar fracture, placement of parallel guide wires, 2.7-mm cannulated drilling, and fracture stabilization with one or two 3.5 x 40–mm noncannulated Mg-alloy (ZX00) screws. A stable construct with compression at the medial malleolar fracture was achieved. Any concurrent fractures of the lateral malleolus, posterior malleolus, and syndesmotic instabilities were managed using standard (stainless steel, titanium, or titanium alloy) implants per the established protocol for trauma surgery. The ankle was immobilized in an under-knee plaster cast for 4 to 6 weeks postoperatively.
Primary and Secondary Study Outcomes
The primary goal was to assess radiographic changes at the implant site on postoperative radiographs. Radiographs of the ankle were obtained at 6 weeks, 1 year, and the final follow-up at a mean of 2.5 ± 0.6 years and analyzed using semiquantitative and qualitative approaches. The semiquantitative analysis evaluated the visibility of Mg-alloy (ZX00) screws using “not visible (-),” “weakly or partially visible (+/++),” and “clearly visible (+++).” For the qualitative analysis, the bone texture at the implant site was evaluated based on its radiographic opacity or lucency and compared with adjacent unaffected bone. Its bony texture was rated as either “homogeneous” or “slightly inhomogeneous” (substantial inhomogeneity was not considered because no cases were rated as such). The radiographic assessments were conducted independently by two evaluators (VL and PLH), and there was 96% agreement between the evaluators. In case of a disagreement, a third evaluator (OS) was consulted for assessment until a consensus was reached. Additionally, radiographic assessment of the ankle included an evaluation of osteoarthritic changes, including subchondral sclerosis, osteophytes, joint space narrowing, cyst formation, and malalignment. Moreover, radiographs were scrutinized for nonunion or refracture. A clinical CT scan was performed at the last follow-up visit to visualize the local situation after ZX00 resorption.
The secondary goal was to assess the clinical outcome, including patient-reported outcome measures. Clinical evaluation assessed the occurrence of various adverse events such as skin or scar complications, infection, hypersensitivity reactions, complex regional pain syndrome, malalignment, and revision surgery. ROM of the ankle was evaluated by measuring dorsal and plantar flexion with a standard goniometer. Patient-reported outcome measures included pain at rest as assessed through the VAS (range 0 to 100), with higher scores indicating greater pain levels, and the American Orthopaedic Foot and Ankle Society hindfoot score, which evaluates pain and function, with higher scores (maximum 100) indicating lower pain levels and better function. All patient-reported outcome measures were collected after a mean of 2.5 years.
Statistical Analysis
The statistical analysis was performed using GraphPad Prism software (version 9.3.1, GraphPad Software). Implant volume, surface area, and gas volume are expressed as the mean ± SD. The mean differences in implant volume and surface values between the peri-implantation and post-implantation stages at 13 and 25 months within the groups of same-sized screws were compared using an unpaired t-test (95% CI, p value). Gas volume values were compared between the two post-implantation timepoints. Differences were considered significant if p values were less than 0.05.
Results
Preclinical Evaluation in Sheep
Implant Volume Resorption, Surface Area, and Gas Formation
After 13 months, the 29-mm screws (initial volume: 198 ± 1 mm3) degraded by approximately 41% to a mean volume of 116 ± 6 mm3 (mean difference 82 [95% CI 71 to 92]; p < 0.001), and after 25 months by approximately 65% to a mean volume of 69 ± 7 mm3 (mean difference 130 [95% CI 117 to 142]; p < 0.001). Moreover, the 24-mm screws (initial volume: 174 ± 0.2 mm3) degraded by approximately 51% to a mean volume of 86 ± 21 mm3 by 13 months (mean difference 88 [95% CI 52 to 123]; p = 0.004), and after 25 months degraded by approximately 72% to a mean volume of 49 ± 25 mm3 (mean difference 125 [95% CI 83 to 167]; p = 0.003). Regarding the 16-mm screw (initial volume: 112 ± 5 mm3), after 13 months, 57% of the initial volume degraded to a mean volume of 49 ± 8 mm3 (mean difference 63 [95% CI 50 to 76]; p < 0.001), whereas after 25 months, approximately 61% was degraded, reaching an end mean volume of 45 ± 10 mm3 (mean difference 67 [95% CI 52 to 82]; p < 0.001) (Fig. 2A).
Fig. 2.
This figure shows the evaluation and comparison of ZX00 and Ti-6Al-4V implant properties obtained by ex vivo high-resolution micro-CT. Different lengths of ZX00 and Ti-6Al-4V screws were implanted in sheep tibiae. (A) Implant volume, (B) implant surface area, and (C) gas volume were obtained by ex vivo micro-CT and previewed with mean and standard deviation analyzed by Mimics software. In comparison to initial values, all three types of implanted ZX00 screws showed a decrease in (A) volume and (B) surface area after 13 and 25 months. The coronal view of ex vivo micro-CT images shows (D) the 16-mm ZX00 screw after 13 months, (E) the 18-mm Ti-6Al-4V screw after 25 months, and (F) the 16-mm ZX00 screw after 25 months. (D, F) The black arrows indicate new bone formation, with qualitatively increased endosteal and periosteal bone volume, and bone formation around the ZX00 screw in the medullary cavity, the black arrowhead indicates a narrow bone-implant interface of ZX00 screws, and the black asterisk indicates gas formation around the ZX00 screw. (E) The white arrow indicates absence of osteoconduction in the medullary cavity, the white arrowhead indicates the visible area between the bone-implant interface, and the white asterisk shows common artefacts after Ti-6Al-4V imaging. (G) 3D images of different length screws after degradation at 13 and 25 months are shown. A color image accompanies the online version of this article.
The surface area of the 29-mm screws (initial surface area: 360 ± 1 mm2) decreased by 26% to a mean value of 267 ± 12 mm2 after 13 months (mean difference 93 [95% CI 73 to 114]; p < 0.001) and by 41% to a mean value of 212 ± 8 mm2 after 25 months (mean difference 148 [95% CI 134 to 162]; p < 0.001) (Fig. 2B). Moreover, the surface area of the 24-mm screws (initial surface: 319 ± 0.4 mm2) decreased by 28% to a mean of 232 ± 29 mm2 after 13 months (mean difference 88 [95% CI 40 to 136]; p = 0.010) and by 53% to a mean of 149 ± 49 mm2 after 25 months (mean difference 170 [95% CI 89 to 252]; p = 0.007). Regarding the 16-mm screws, after 13 months, the surface area (initial surface area: 233 ± 0.2 mm2) decreased by 37% to a mean of 147 ± 12 mm2 (mean difference 80 [95% CI 60 to 100]; p < 0.001), whereas it decreased by 35% after 25 months to 152 ± 56 mm2 (mean difference 74 [95% CI -7 to 156]; p = 0.068) (Fig 2B).
The mean gas volume around the 29-mm screws was 41 ± 42 mm3 after 13 months and 23 ± 3 mm3 after 25 months (mean difference 19 [95% CI -109 to 146]; p = 0.596). In the 24-mm screws, the mean gas volume was 48 ± 7 mm3 and 47 ± 23 mm3 after 13 and 25 months, respectively (mean difference 1 [95% CI -72 to 74]; p = 0.965). The mean gas volume in the 16-mm screws was 73 ± 92 mm3 after 13 months and 11 ± 13 mm3 after 25 months (mean difference 63 [95% CI -33 to 159]; p = 0.170) (Fig. 2C).
Ex vivo micro-CT images demonstrated ongoing Mg-alloy screw degradation and showed the Mg-alloy and Ti-6Al-4V screw positions after 13 and 25 months in sheep tibiae (Fig. 2D-F). The software evaluation of implant degradation allowed for a 3D preview of the screws at the initial and final timepoints. (Fig. 2G).
Histologic Evaluation
A qualitative analysis was performed on thin ground sections of the 24-mm Mg-alloy (ZX00) screw after 25 months with signs of ongoing screw resorption, with visible gas cavities predominantly formed in the cortical bone and around the screw head (Fig. 3). Newly formed bone with an emphasis on “bony bridge” formation along with a narrow bone-implant interface was visible in the medullary cavity. The area around the screw head appeared to be completely covered with newly formed cortical bone with a normal morphologic appearance. The formation of reactive woven bone with osteoblasts and osteoclasts was adjacent to the cortical bone near the screw head. Additionally, areas of fat necrosis were focally present around the degraded parts of the screw head. No inflammation or foreign body giant cells were observed.
Fig. 3.
(A-C) Histologic evaluation of the 24-mm ZX00 screw after the 25-month period in sheep tibiae with (D) respective micro-CT. (A) New bone formation, “bony bridges,” within the medullary cavity is marked with a black arrow, and reactive woven bone formation on the periosteum adjacent to the screw head is marked with a white asterisk. (B) Osteocytes and osteoblasts are marked with black and white arrowheads, respectively. (C) The fat necrosis area is shown with white arrows, and air pockets are marked with black asterisks (Laczko-Levai stain; original magnification, x10).
Histologic sections of the 18-mm Ti-6Al-4V screw after 25 months in the tibia diaphysis (Fig. 4) showed a narrow bone-implant interface in the cortical bone. The region around the screw head was covered with newly formed bone, and adjacent to the cortical bone in the same region, there was formation of reactive woven bone. Moreover, areas of fat necrosis were focally present around the screw head, whereas no inflammatory cells were observed.
Fig. 4.
(A-C) Histologic evaluation of the 18-mm Ti-6Al-4V screw after the 25-month period in sheep tibiae with (D) respective micro-CT. (A) Newly formed bone in the medullary cavity is marked with a black arrow, and reactive woven bone formation on the periosteum adjacent to the screw head is labeled with a white asterisk. (B) Osteocytes and osteoblasts are marked with black and white arrowheads, respectively, whereas the (C) fat necrosis area is shown with a white arrow (Laczko-Levai stain; original magnification, x10).
Radiographic and Clinical Evaluation of Patients
Radiographic Findings
A representative case with radiographs after medial malleolar fixation using ZX00 screws and fibulo-tibial stabilization (with a titanium screw) demonstrated no visible screws and a homogeneous bone texture at the ZX00 implantation site at the final follow-up (Fig. 5). Similarly, in another case involving ZX00 fixation at the medial malleolus, along with plating at the lateral and posterior malleoli, there were no visible screws at the previous ZX00 implantation site at the final follow-up (Fig. 6). The pattern of decreasing screw visibility is consistently observed across patients (examples of such patients can be seen in Supplemental Figs. 1-4; http://links.lww.com/CORR/B212).
Fig. 5.
(A-D) Radiographic evaluation of a 47-year-old female patient who underwent surgery for a Maisonneuve fracture. Postoperative radiographs were obtained at (A) 6 weeks, (B) 1 year, and (C) 3 years and 7 months after surgical fixation. The patient received ZX00 screws for the medial malleolus fracture and a fibulo-tibial positioning screw (titanium) for syndesmotic instability, which was removed 6 weeks postoperatively (white arrow indicating screw hole). (D) Axial CT images at four levels reveal implant sites after ZX00 screw resorption at the medial malleolus and the titanium positioning screw site (white arrow) at the distal tibia. Both sites show changes or a lack of trabecular structures. The visibility of the ZX00 screws was rated as (+++) at 6 weeks, (+/++) at 1 year, and (-) at the final follow-up (reprinted with permission from the authors [10]).
Fig. 6.
(A-C) Radiographic evaluation of a 20-year-old male patient who was surgically treated for a trimalleolar fracture. Postoperative radiographs were obtained at (A) 6 weeks, (B) 1 year, and (C) 2 years and 10 months after injury and surgery. The patient received two ZX00 screws for the medial malleolus fracture and locking compression plates for the lateral and posterior malleolus fractures. The locking compression plates were removed 1 year and 6 months postoperatively. The visibility of the ZX00 screws was rated as (+++) at 6 weeks, (+/++) at 1 year, and (-) at the final follow-up.
Regarding the semiquantitative radiographic assessment, screw visibility was rated as “not visible” in 17 of 18 patients, “partially or weakly visible” in one of 18, and “clearly visible” in 0 of 18 at the final follow-up at a mean of 2.5 years. Gas formation was rated as “not visible” in 17 of 18 patients, “partially or weakly visible” in one of 18, and “clearly visible” in 0 of 18 at the final follow-up. The bone texture at the implant site was “homogeneous” in 15 of 18 and “slightly inhomogeneous” in three of 18 patients at the final follow-up.
Qualitatively, radiographs showed a progressive increase in homogeneity at the implantation site at the final follow-up compared with 1 year, resembling the texture of the adjacent nonimplanted bone. In all 18 ankle radiographs, evidence of fracture union was observed without indications of nonunion or refracture. Radiographs were further assessed for signs of osteoarthritis. Mild changes with subchondral sclerosis were seen in five patients. Moderate changes with progressive subchondral sclerosis and initial osteophyte formation were detected in two patients. On visual examination of a CT image from a single patient, the absence of trabeculae was observed at the site of ZX00 screw resorption and where a titanium positioning screw had been implanted (subsequently removed 6 weeks postsurgery) (Fig. 5D).
Clinical Findings
None of the 18 patients experienced any complications or underwent additional surgery during the study. The mean dorsal flexion was 22º ± 6°, and the mean plantar flexion was 47º ± 5°. The mean VAS score was 1 ± 0 at rest, ranging from 0 to 100, with higher scores indicating greater pain. The mean American Orthopaedic Foot and Ankle Society score was 94 ± 5 points, ranging from 0 to 100, with higher scores indicating better function and less pain (Table 1).
Table 1.
Clinical findings at the final follow-up
| Variable | Value |
| Ankle dorsal flexion, ° | 22 ± 6 |
| Ankle plantar flexion, ° | 47 ± 5 |
| VAS score (range 0 to 100) | 1 ± 0 |
| American Orthopaedic Foot and Ankle Society score (range 0 to 100) | 94 ± 5 |
Data are presented as mean ± SD at mean follow-up of 2.5 ± 0.6 years
Discussion
Bioresorbable implants provide adequate compression and stability for bone healing while gradually being resorbed, rendering hardware removal unnecessary. Therefore, they could potentially reduce disease burden and costs. The Mg-alloy (ZX00) screw used in this study comprises three elements naturally present in the human body and has already been shown to resorb without the formation of precipitates or adverse immunological reactions [1, 7]. However, a temporal evaluation of the resorption and bone interaction of Mg-alloy-based screws for fracture fixation is missing and must be performed in terms of long-term outcomes. Therefore, we examined the resorption of implanted Mg-alloy (ZX00) screws via micro-CT data and the bone response via histologic evaluation at 13 and 25 months postsurgery in a sheep model. Furthermore, we reported the radiographic changes and clinical performance at a mean of 2.5 years after Mg-alloy (ZX00) screw fixation in patients with medial malleolar fractures. We observed extensive resorption of the Mg-alloy (ZX00) screws, with radiographic disappearance in 17 of 18 patients and partial disappearance in one patient, while achieving good functional outcomes. Furthermore, the study demonstrated a high degree of resorption in sheep, with good biocompatibility. However, longer-term radiologic and clinical assessments are important to pursue because bone remodeling can be expected to continue.
Limitations
This study has several limitations, including the small sample size in the preclinical and clinical studies. Despite the limited power, our observations in the sheep model indicated adequate resorption of the Mg-alloy (ZX00) screws, with comparable implant volume loss and gas volume as in previous preclinical studies conducted over shorter observation periods [7, 19]. In vivo studies remain crucial for understanding the long-term tissue response to implant degradation. Simultaneously, it is important to always strive to minimize animal suffering during such studies. Another limitation is the difference in age between the study animals and the study patients between the preclinical and clinical studies. The resorption study was performed on growing sheep, although the histologic evaluation at 25 months reflects an adult sheep model. In contrast, the clinical study was conducted with adult patients. We chose our established model of female, juvenile sheep [7, 9, 19] to investigate the resorption behavior and bone response to Mg-alloy (ZX00) in a long-term study throughout the maturation of bone. This decision was based on the similarities in bone size between sheep and humans, as opposed to conducting the study using small-animal models such as rodents. The time period is also a limitation. The final evaluation of Mg-alloy (ZX00) screw resorption in the sheep model occurred at 25 months, with approximately one-third of the original size of the screws remaining. In contrast, the final evaluation in patients at a mean of 2.5 years (30 months) showed that the implants were undetectable on radiographs, suggesting full resorption. An ideal study would span a longer period with multiple sample points to determine the complete resorption timeline for the Mg-alloy (ZX00) implant in the sheep model. However, based on the combined preclinical and clinical evaluations, it is expected that the Mg-alloy (ZX00) will eventually fully resorb in sheep. The absence of inflammation in the histology of Mg-alloy (ZX00) screws in sheep at 25 months, along with favorable radiographic findings and positive clinical outcomes in patients, suggest the tissue reaction in humans may be as advantageous as observed in sheep. The difference in bone regions between the preclinical and clinical studies is also a limitation because gas formation during screw resorption differed among different regions of bone, with the diaphysis exhibiting the highest levels owing to the space of the medullary cavity [19]. Although the screws used in sheep were allocated to the diaphysis and diametaphysis, in humans, medial malleolar fractures were fixed in the epiphysis. We did not specifically address differences in screw resorption behavior across different bone regions.
The clinical study also has several limitations, including the loss of two patients who were not part of the current follow-up. The loss to follow-up of one female patient after 12 weeks [10] and the discontinuation of another female patient after 17 months [11] could introduce transfer bias. Furthermore, this contributes to the inherent gender bias in a mixed-gender study group; it cannot be assumed that the findings are equally applicable to men and women separately. Despite the defined inclusion criteria, there is a likelihood of selection bias based on the surgeon's decision and the patient's willingness to participate in the study, which could lead to an overrepresentation of fitter and more adherent patients. Additionally, assessment bias might have occurred because we conducted the clinical follow-up without external validation. The absence of advanced imaging techniques is notable. Although radiographs revealed a homogeneous bone texture, CT slices revealed gaps at the implant site in the trabecular bone, which may be because of recent resorption that has not been remodeled yet and may only be resolved with a longer bone remodeling period. Even for titanium alloy screws in this study, this phenomenon is seen after removal surgery (Fig. 5D). Further investigation is necessary, including CT or even HR-pQCT studies in a larger sample, to address this knowledge gap. The lack of comparability between imaging modalities is clearly a limitation of this study. Finally, in the clinical study, we did not compare the Mg-alloy (ZX00) screws with conventional standard implants (screws made of stainless steel, titanium, or titanium alloy) or other biodegradable products.
Preclinical Evaluation in Sheep
The results from our sheep experiments demonstrated a decrease in Mg-alloy (ZX00) screw volume, reaching approximately half of the original volume at 13 months and further decreasing to approximately one-third of the original volume at 25 months. Additionally, we observed a concurrent reduction in implant surface and gas volume, indicating an ongoing dynamic process during the study period. In previous work, our study group reported that the resorption rate of Mg-alloy (ZX00) implants in growing sheep ranged between 0.23 and 0.75 mm/year [19]. Higher resorption rates were found in the epiphysis than in the metaphysis of the distal tibia, potentially because of differences in blood supply to these regions in growing bone [19].
The presence of gas formation in the medullary cavity and its absence at the screw head, surrounded by newly formed bone, were seen with micro-CT and histologic analysis. Histology revealed the biocompatibility of Mg-alloy (ZX00) screws with the physiologic bone-cell environment, as indicated by the presence of osteoclasts and osteoblasts and the absence of foreign body giant cells or inflammation. The analysis also revealed new bone bridges along the implant surface and the presence of fat necrosis around the degraded parts of the screw head, indicating physiologic bone healing processes. Compared with the Ti-alloy implant, Mg-alloy (ZX00) screws showed less-enhanced new bone formation in the medullary cavity but maintained a physiologic bone cell environment without foreign body giant cells. In contrast, in the cortical bone area, the screw head of the Mg-alloy (ZX00) screws showed more-pronounced new bone formation. Additionally, resorption of Mg-alloy (ZX00) does not result in the formation of chelate complexes, giving rise to precipitates in the surrounding tissue that do not resorb, as occurs with alloys containing rare-earth elements. Overall, the results suggest that Mg-alloy (ZX00) screws demonstrate biocompatibility and promote osseointegration and osteoconduction in a sheep model, comparable to our previous study [7].
Clinical Evaluation in Patients
The key finding in the clinical study is the radiologic disappearance of the Mg-alloy (ZX00) screws on radiographs at a mean of 2.5 years postoperatively, suggesting successful resorption. Biopolymers based on poly-L-lactic acid, with a high lactic acid content and high molecular weight, exhibit considerably longer resorption times [5, 26]. One year after Mg-alloy (ZX00) implantation, the screws were still weakly or partly visible, and trabecular bone that was not fully remodeled could be discerned. Substantial resorption of the Mg-alloy (ZX00) screws continued past 1 year until they became increasingly invisible on radiographs and disappeared at approximately 2.5 years, with increasingly homogeneous bone texture at the implant site. This increased radiopacity can be attributed to resorption of the screw and formation of new bone at the implant site, or the persistence of a defect in the trabecular bone with surrounding trabeculae becoming stronger (and thus more radiopaque) in response to changes in mechanical stress and force transmission. The question of how these changes in the trabecular bone are managed by the body after resorption leaves room for further investigation. A final judgment on the end state can only be made after further observation and longer follow-up.
Other studies reporting outcomes with degradable Mg screws in patients have not disclosed any long-term resorption data. Apart from radiographs, no advanced imaging techniques (such as CT or HR-pQCT [17]) have been used or reported [13, 15, 18, 20, 23, 25, 28] to describe the long-term resorption performance at the implant site.
Remarkably, the CT images in one patient demonstrated a noteworthy absence of trabeculae at the previous implantation site of the titanium screw (a fibulotibial positioning screw that was removed 6 weeks postoperatively) and at the implantation site of the ZX00 screws after resorption (Fig. 5D). Evidently, the adjacent trabecular network must have undergone adaptation to accommodate and distribute the load or stress in these regions. It is crucial to determine whether these findings are associated with any clinical complications, such as pain or refracture, through long-term observations.
Thus far, the clinical findings beyond 1 year are positive; patients are pain-free with good ankle function and without complications or revision surgery. Despite a high prevalence of severe injuries, such as Maisonneuve and trimalleolar fractures [12], patients show excellent functional outcomes, even the seven patients with radiographic signs of mild or moderate post-traumatic osteoarthritis.
Conclusion
This sheep study confirmed resorption of Mg-alloy (ZX00) screws to one-third of the original volume after 25 months with no histologic signs of inflammation, suggesting good biocompatibility. Radiographs of patients suggest full resorption of Mg-alloy (ZX00) screws and bony integrity after a mean follow-up of 2.5 years, with pain-free ankle function after medial malleolar fracture fixation. To fully evaluate the local effect at the implant site post-resorption, further investigation using advanced imaging techniques is required. Additionally, comparative trials are necessary to demonstrate the noninferiority of Mg-alloy (ZX00) implants compared with standard implants. To accurately interpret postoperative radiographs, clinicians should become familiar with radiographic changes during the resorption of Mg-alloy (ZX00) screws because it is a dynamic process; the current work may serve as a useful guide. Based on current insights, the use of Mg-alloy (ZX00) screws for the surgical fixation of medial malleolar fractures is promising.
Supplementary Material
Acknowledgments
We thank the team of the emergency unit, ward, and surgical unit of the Department of Orthopaedics and Trauma at the Medical University of Graz for taking care of the patients.
Footnotes
The institution of one or more of the authors (AL) has received funding, during the study period, in an amount of USD 10,000 to USD 100,000 from Johnson & Johnson, Alphamed, Implantec, and Medacta. One of the authors (PLH) certifies receipt of personal payments or benefits, during the study period, in an amount of USD 10,000 to USD 100,000 from the Lorenz Böhler Fond and holds shares in Bioretec Ltd.
All ICMJE Conflict of Interest Forms for authors and Clinical Orthopaedics and Related Research® editors and board members are on file with the publication and can be viewed on request.
Clinical Orthopaedics and Related Research® neither advocates nor endorses the use of any treatment, drug, or device. Readers are encouraged to always seek additional information, including FDA approval status, of any drug or device before clinical use.
Ethical approval for the human study was approved and registered by the local ethics committee of Medical University of Graz under number: 28 to 071 ex 15/16. All animal experiments for this study were approved by the Austrian Federal Ministry for Science and Research under the following permits: BMWFW-66.010/0073-WF/V/3b/2015 and BMWFW-66.010/0049-WF/II/3b/14.
This work was performed at the Medical University of Graz, Graz, Austria.
Contributor Information
Viktor Labmayr, Email: viktor.labmayr@medunigraz.at.
Omer Suljevic, Email: omer.suljevic@medunigraz.at.
Nicole Gabriele Sommer, Email: nicole.sommer@medunigraz.at.
Uwe Yacine Schwarze, Email: uwe.schwarze@medunigraz.at.
Romy Linda Marek, Email: romy.marek@medunigraz.at.
Iva Brcic, Email: iva.brcic@medunigraz.at.
Ines Foessl, Email: ines.foessl@medunigraz.at.
Andreas Leithner, Email: andreas.leithner@medunigraz.at.
Franz Josef Seibert, Email: franz.seibert@medunigraz.at.
Valentin Herber, Email: valentin.herber@unibas.ch.
Patrick Lukas Holweg, Email: p.holweg@medunigraz.at.
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