The Masquelet technique is a variable method for treating critical-sized bone defects, but there is a need to develop a technique for promoting bone regeneration. In recent studies of bone fracture healing promotion, macrophage-mesenchymal stem cell (MSC) cross-talk has drawn attention. This study aimed to investigate macrophage expression in the induced membrane (IM) of the Masquelet technique using a mouse critical-sized bone defect model.

The study involved a 3-mm bone defect created in the femur of mice and fixed with a mouse locking plate. The Masquelet (M) group, in which a spacer was inserted, and the Control (C) group, in which the defect was left intact, were established. Additionally, a spacer was inserted under the fascia of the back (B group) to form a membrane due to the foreign body reaction. Tissues were collected at 1, 2, and 4 weeks after surgery (n=5 in each group), and immunostaining (CD68, CD163: M1, M2 macrophage markers) and RT-qPCR were performed to investigate macrophage localization and expression in the tissues.

The study found that CD68-positive cells were present in the IM of the M group at all weeks, and RT-qPCR showed the highest CD68 expression at 1 week. In addition, there was similar localization and expression of CD163. The C group showed lower expression of CD68 and CD163 than the M group at all weeks. The B group exhibited CD68-positive cells in the fibrous capsule and CD163-positive cells in the connective tissue outside the capsule, with lower expression of both markers compared to the M group at all weeks.

Macrophage expression in IM in M group had different characteristics compared to C group and B group. These results suggest that the IM differs from the fibrous capsules due to the foreign body reaction, and the macrophage-MSC cross-talk may be involved in Masquelet technique.

Recent researches indicate that both M1 and M2 macrophages play vital roles in tissue repair and foreign body reaction processes. In this study, we investigated the dynamics of M1 macrophages in the induced membrane using a mouse femur critical-sized bone defect model.

The Masquelet method (M) and control (C) groups were established using C57BL/6J male mice (n=24). A 3mm-bone defect was created in the right femoral diaphysis followed by a Kirschner wire fixation, and a cement spacer was inserted into the defect in group M. In group C, the bone defect was left uninserted. Tissues around the defect were harvested at 1, 2, 4, and 6 weeks after surgery (n=3 in each group at each time point). Following Hematoxylin and eosin (HE) staining, immunohistochemical staining (IHC) was used to evaluate the CD68 expression as a marker of M1 macrophage. Iron staining was performed additionally to distinguish them from hemosiderin-phagocytosed macrophages.

In group M, HE staining revealed a hematoma-like structure, and CD68-positive cells were observed between the spacer and fibroblast layer at 1 week. The number of CD68-positive cells decreased at 2 weeks, while they were observed around the new bone at 4 and 6 weeks. In group C, fibroblast infiltration and fewer CD68-positive cells were observed in the bone defect without hematoma-like structure until 2 weeks, and no CD68-positive cells were observed at 4 and 6 weeks. Iron staining showed hemosiderin deposition in the surrounding area of the new bone in both groups at 4 and 6 weeks. The location of hemosiderin deposition was different from that of macrophage aggregation.

This study suggests that M1 macrophage aggregation is involved in the formation of induced membranes and osteogenesis and may be facilitated by the presence of spacers.

Segmental bone transport (SBT) using an external fixator is currently a standard treatment for large-diameter bone defects at the donor site with low morbidity. However, long-term application of the device is needed for bone healing. In addition, patients who received SBT treatment sometimes fail to show bone repair and union at the docking site, and require secondary surgery. The objective of this study was to investigate whether a single injection of recombinant human bone morphogenetic protein 2 (rhBMP-2)-loaded artificial collagen-like peptide gel (rhBMP-2/ACG) accelerates consolidation and bone union at the docking site in a mouse SBT model.

Six-month-old C57BL/6J mice were reconstructed by SBT with external fixator that has transport unit, and a 2.0-mm bone defect was created in the right femur. Mice were divided randomly into four treatment groups with eight mice in each group, Group CONT (immobile control), Group 0.2mm/d, Group 1.0mm/d, and Group BMP-2. Mice in Group 0.2mm/d and 1.0mm/d, bone segment was moved 0.2 mm per day for 10 days and 1.0 mm per day for 2 days, respectively. Mice in Group BMP-2 received an injection of 2.0 μg of rhBMP-2 dissolved in ACG into the bone defect site immediately after the defect-creating surgery and the bone segment was moved 1.0 mm/day for 2 days.

All animals were sacrificed at eight weeks after surgery. Consolidation at bone defect site and bone union at docking site were evaluated radiologically and histologically.

At the bone defect site, seven of eight mice in Group 0.2mm/d and two of eight mice in Group 1.0mm/d showed bone union. In contrast, all mice in Group CONT showed non-union at the bone defect site. At the docking site, four of eight mice in Group 0.2 mm/d and three of eight mice in Group 1.0 mm/d showed non-union. Meanwhile, all mice in Group BMP-2 showed bone union at the bone defect and docking sites. Bone volume and bone mineral content were significantly higher in Group 0.2mm/d and Group BMP-2 than in Group CONT. HE staining of tissue from Group 0.2mm/d and Group BMP-2 showed large amounts of longitudinal trabecular bone and regenerative new bone at eight weeks after surgery at the bone defect site. Meanwhile, in Group CONT and Group 1.0mm/d, maturation of regenerative bone at the bone defect site was poor. Differences between groups were analyzed using one-way ANOVA and a subsequent Bonferroni's post-hoc comparisons test. P < 0.05 was considered significant.

rhBMP-2/ACG combined with SBT may be effective for enhancing bone healing in large bone defects without the need for secondary procedures.

Segmental bone transport (SBT) with an external fixator has become a standard method for treatment of large bone defect. However, a long time-application of devices can be very troublesome and complications such as nonunion is sometimes seen at docking site. Although there have been several studies on SBT with large animal models, they were unsuitable for conducting drug application to improve SBT. The purpose of this study was to establish a bone transport model in mice. Six-month-old C57BL/6J mice were divided randomly into bone transport group (group BT) and an immobile control group (group EF). In each group, a 2-mm bone defect was created in the right femur. Group BT was reconstructed by SBT with external fixator (MouseExFix segment transport, RISystem, Switzerland) and group EF was fixed simply with unilateral external fixator (MouseExFix simple). In group BT, a bone segment was transported by 0.2 mm per day. Radiological and histological studies were conducted at 3 and 8 weeks after the surgery. In group BT, radiological data showed regenerative new bone consolidation at 8 weeks after the surgery, whereas high rate of nonunion was observed at the docking site. Histological data showed intramembranous and endochondral ossification. Group EF showed no bone union. In this study, experimental group showed good regenerative new bone formation and was similar ossification pattern to previous large animal models. Thus, the utilization of this bone defect mice model allows to design future studies with standardized mechanical conditions for analyzing mechanisms of bone regeneration induced by SBT.

The reduction for unstable femoral intertrochanteric fracture should be extramedullary, which means that the proximal fragment protrudes for the distal fragment. However, only few articles have compared extramedullary and intramedullary reductions in a biomechanical study. Thus, we created unstable femoral intertrochanteric fracture models using imitational bone (extramedullary and intramedullary groups, each with 12 cases) and evaluated their biomechanical stabilities. The fracture type was 31-A2 according to the AO-OTA Classification of Fractures and Dislocations and greatly lacked bone on the posterior side. We performed compression examination and evaluated stiffness. The implant used for fixation was TFNA (DePuy Synthes). We applied axial compression with 20 adduction in the standing position. Statistical analysis was performed using the Mann-Whitney U test. No significant difference in initial loading force was found between the two groups. However, the axial stiffness of the extramedullary bone showed a significant increase (p < 0.05) in high loading force (800–1000 N). This means that the stability of the extramedullary reduction was superior to that of the intramedullary reduction in terms of high loading force in the standing position. We suggest that antero-medial bony buttress is important for unstable femoral intertrochanteric fractures. These data indicate that extramedullary reduction and fixation for unstable femoral intertrochanteric fractures increase stability.

To elucidate the molecular biology of fracture healing, murine models are preferred. We performed a study with the first internal fixation system that allows studying murine fracture healing in a controlled mechanical environment, to characterise the timing of the fracture healing cascade with this model, based on a histological evaluation.

Femoral osteotomies were performed in 68 male C57BL/six mice and stabilised with locking internal fixation plates in either stiff, or defined, flexible configurations. Healing progression was studied at 10 time points between 3 and 42 days post- surgery. After surgery, mice were radiographed to confirm the correct implant positioning. After sacrifice, the extracted femora were processed for decalcified histology. Thin sections were taken as serial transverse sections and stained for subsequent histomorphometric analysis and three-dimensional reconstruction of the different fracture callus tissues.

The surgery was successful in 62 animals. Only six6 (8.8%) animals had to be sacrificed due to complications during surgery. The post-operative radiographs demonstrated a high reproducibility of implant positioning and no implant failure or screw loosening occurred during the experimental period. The improved consistency in surgical technique leading to more uniform results represents a key advantage of this system over other mouse fracture healing models. As such, it may allow a reduction in the sample size needed in future murine fracture healing studies. The histological evaluation confirmed the lack of a periosteal callus, and exclusively endosteal, intramembraneous bone formation in the bones stabilised with the stiff implants. The bones that were stabilised with the more flexible internal fixation plates showed additional endochondral ossification with extensive, highly asymmetrical, periosteal callus formation.

Our results demonstrate that this murine fracture model leads to different healing patterns depending on the flexibility of the chosen plate system. This allows researchers to investigate the molecular biology of fracture healing in different ossification modes by selection of the appropriate fixation.

The aim of this study is to compare the clinical results of the Pin-Sleeve System (AI Wiring System; AIWS) with the Dall-Miles Cable Grip System (DMCGS) for reattachment after dissection of the greater trochanter in hip arthroplasty.

The DMCGS was used in 33 cases 35 hips from 1994 to 1998 and AIWS in 40 cases 42 hips from 1998 to 2001. The age at operation was 61.3 years (24 to 85 years) in the DMCGS group and 67 years (24 to 86 years) in the AIWS group. The postoperative follow-up period was 24 months (4 months to 54 months) in the DMCGS group and 30 months (11 months to 42 months) in the AIWS group.

Bone union failure of the great trochanter occurred in the DMCGS group eight hips (22.9%) and AIWS group five hips (11.9%). The DMCGS group four hips (11.4%) had broken cables, while not even one case of the AIWS group had them (p< 0.05). Fragments from the cable were found in the DMCGS group seven hips (20%) and AIWS group two hips (4.8%). Bone resorption around the cable, grip or sleeve occurred in the DMCGS group 19 hips (54.3%) and AIWS group five hips (11.9%) (p< 0.05). Clinically, the DMCGS group 13 hips (37.1%) and AIWS group seven hips (16.7%) had dysphoria at the greater trochanter; the DMCGS group 17 hips (48.6%) and AIWS group eight hips (19%) had pain at the greater trochanter in the recumbent position with the affected side down (p< 0.05); the DMCGS group 13 hips (37.1%) and AIWS group six hips (14.3%) had pain on exertion.

The AIWS is considered to be a useful implant for reattachment of the greater trochanter compared with the DMCGS.