Our previous rat study demonstrated an ex vivo-created “Biomimetic Hematoma” (BH) that mimics the intrinsic structural properties of normal fracture hematoma, consistently and efficiently enhanced the healing of large bone defects at extremely low doses of rhBMP-2 (0.33 μg). The aim of this study was to determine if an extremely low dose of rhBMP-2 delivered within BH can efficiently heal large bone defects in goats.

Goat 2.5 cm tibial defects were stabilized with circular fixators, and divided into groups (n=2-3): 2.1 mg rhBMP-2 delivered on an absorbable collagen sponge (ACS); 52.5 μg rhBMP-2 delivered within BH; and an empty group. BH was created using autologous blood with a mixture of calcium and thrombin at specific concentrations. Healing was monitored with X-rays. After 8 weeks, femurs were assessed using microCT.

Using 2.1 mg on ACS was sufficient to heal 2.5 cm bone defects. Empty defects resulted in a nonunion after 8 weeks. Radiographic evaluation showed earlier and more robust callus formation with 97.5 % (52.5 μg) less of rhBMP-2 delivered within the BH, and all tibias were fully bridged at 3 weeks. The bone mineral density was significantly higher in defects treated with BH than with ACS. Defects in the BH group had smaller amounts of intramedullary and cortical trabeculation compared to the ACS group, indicating advanced remodeling.

The results confirm that the delivery of rhBMP-2 within the BH was much more efficient than on an ACS. Not only did the large bone defects heal consistently with a 40x lower dose of rhBMP-2, but the quality of the defect regeneration was also superior in the BH group. These findings should significantly influence how rhBMP-2 is delivered clinically to maximize the regenerative capacity of bone healing while minimizing the dose required, thereby reducing the risk of adverse effects.

This study investigated concurrent talar dome injuries associated with tibial pilon fractures, mapping their distribution across the proximal talar dome articular surface. It compared the two main mechanisms of injury (MOI), falling from a height and motor vehicle accident (MVA), and whether the fractures were open or closed.

From a previously compiled database of acute distal tibial pilon fractures (AO/OTA 43B/C) in adults of 105 cases, 53 cases were identified with a concurrent injury to the talar dome with a known mechanism of injury and in 44 it was known if the fracture was open or closed. Case specific 2D injury maps were created using a 1×1mm grid, which were overlayed in an Excel document to allow for comparative analyses. A two-way ANOVA was conducted that examined the effect of both MOI and if the fracture was open or closed on what percentage of the talar dome surface was injured.

There was a statistically-significant difference between the average percentage of injured squares on the talar dome by both whether the fracture was open or closed (f(1)=5.27, p= .027) and the mechanism of injury (f(1)=8.08, p= .007), though the interaction between these was not significant (p= .156). Open injuries and injuries that occurred during an MVA were more likely to increase the surface area of the talar dome injuries.

We have identified both MOI and if the fracture was either open or closed impacts the size of the injury present on the talar dome. Future research will investigate the aetiology of the differences noted, highlighting the clinical implications.

Surgeons treating tibial pilon fractures caused by either a MVA or an open fracture, should be aware of an increased risk of large injuries to the surface of the talar dome.

Tibial pilon fractures are typically the result of high-energy axial loads, with complex intra- articular fractures that are often difficult to reconstruct anatomically. Only nine simultaneous pilon and talus fractures have been published previously, but we hypothesised the chondral surface of the dome is affected more frequently.

Data was acquired prospectively from 154 acute distal tibial pilon fractures (AO/OTA 43B/C) in adults. Radiographs, photographs, and intra-operative drawings of each case were utilised to document the presence of any macroscopic injuries of the talus. Detailed 1x1mm maps were created of the injuries in each case and transposed onto a statistical shape model of a talus; this enables the cumulative data to be analysed in Excel. Data was analysed using a Chi-squared test.

From 154 cases, 104 were considered at risk and their talar domes were inspected; of these, macroscopic injuries were identified in 55 (52.4%). The prevalence of talar dome injury was greater with B-type fractures (53.5%) than C-type fractures (31.5%) (ρ = .01). Injuries were more common in men than women and presented with different distribution of injuries (ρ = .032). A significant difference in the distribution of injuries was also identified when comparing falls and motor vehicle accidents (ρ = .007).

Concomitant injuries to the articular surface of the dome of the talus are relatively common, and this perhaps explains the discordance between the post-operative appearance following internal fixation and the clinical outcomes observed. These injuries were focused on the lateral third of the dome in men and MVAs, whereas women and fall mechanism were more evenly distributed.

Surgeons who operatively manage high-energy pilon fractures should consider routine inspection of the talar dome to assess the possibility of associated macroscopic osteochondral injuries.

Treatment of segmental bone defects remains a major clinical problem, and innovative strategies are often necessary to successfully reconstruct large volumes of bone. When fractures occur, the resulting hematoma serves as a reservoir for growth factors and a space for cell infiltration, both crucial to the initiation of bone healing. Our previous studies have demonstrated very clear ultrastructural differences between fracture hematomas formed in normally healing fractures and those formed in segmental bone defects. However, there is little information available regarding potential differences in the underlying gene expression between hematomas formed in normal fractures, which usually heal by themselves, and segmental bone defects, which do not. Therefore, the aim of this study was to identify differences in gene expression within hematomas collected from 0.5 mm (normal fracture) and 5 mm (segmental bone defect) fracture sites during the earliest stages of bone healing.

Osteotomies of 0.5 and 5 mm in the femur of Fisher 344 rats were stabilized with external fixators (RISystem AG). After 3 days the rats were sacrificed, and the fracture hematomas were collected for RNA-sequencing. Ingenuity pathway analysis (IPA) was used to identify upstream regulators and biological functions that were significantly enriched with differentially expressed genes from the RNA-sequencing analysis. Animal procedures were conducted following the IACUC protocol of the UT Health Science Center San Antonio.

Key upstream regulators of bone formation were less active (e.g. TGFB1, FGF2, SMAD3) or even inhibited (e.g. WNT3A, RUNX2, BMP2) in non-healing defects when compared to normally healing fractures. Many upstream regulators that were uniquely enriched in healing defects were molecules recently discovered to have osteogenic effects during fracture healing (e.g. GLI1, EZH2). Upstream regulators uniquely enriched in non-healing defects were mainly involved in an abnormal modulation of hematopoiesis, revealing evidence of impaired maturation of functional macrophages and cytokines (e.g. IL3, CEBPE), both essential for successful bone healing. In addition, the enrichment pattern suggested a dysregulation of megakaryopoiesis (e.g. MRTFA, MRTFB, GATA2), which directly affects platelet production, and therefore fracture hematoma formation. Remarkably, the organization of the ECM was the most significantly enriched biological function in the normally healing fractures, and implies that the defect size directly affected the structural properties within the fracture hematoma. Conversely, genes encoding important ECM components (e.g. BGN, various collagens, IBSP, TNC), cell adhesion molecules, MMPs (MMP2), and TIMPs were all significantly downregulated in non-healing defects.

Our most recent findings reveal new important key molecules that regulate defect size-dependent fracture healing. Combined with our previous results, which identified structural differences in fracture hematomas from both types of defects, current findings indicate that differential expression of genes is dictated by the structural properties of the hematomas formed during early fracture healing. Consequently, creating a bioscaffold that mimics the structure of normal fracture hematomas could be the first step towards developing new orthoregenerative treatment strategies that potentiate healing of large bone defects and non-healing fractures.

The management of bone defects and impaired fracture healing remains one of the most challenging clinical problems. Several treatments exist to aid in the healing of large bone defects, including biologics such as recombinant human bone morphogenetic protein-2 (BMP-2), yet all have met with limited success. Regeneration of bone requires a coordinated network of molecular signals where the local mechanical environment plays a major role in the success of the healing process. The mechanical environment itself is determined by the stiffness of the implant used to stabilize the fracture and weight-bearing, and if fixation is either too flexible or too rigid the healing might fail. The hypothesis is that the healing of large-segmental bone defects and fractures can be accelerated by the imposition of an appropriate mechanical environment. An overview of the progress made in this research area on how the amount of rhBMP-2 could be reduced and its effectiveness increased by providing an optimized mechanical environment to achieve bone union will be presented. Additionally, the latest findings of improved fracture healing through the manipulation of fixation stability introducing a potential clinical strategy to improve the healing outcome of unstable fractures, particularly for non-unions through increased stabilization, will be discussed.

Aims

This study describes the Osseointegration Group of Australia’s Accelerated Protocol two-stage strategy (OGAAP-1) for the osseointegrated reconstruction of amputated limbs.