Closed Femur Fracture Model Research Articles

A segmental defect adaptation of the mouse closed femur fracture model for the analysis of severely impaired bone healing.

ObjectiveTo better characterize nonunion endochondral bone healing and evaluate novel therapeutic approaches for critical size defect healing in clinically challenging bone repair, a segmental defect model of bone injury was adapted from the three‐point bending closed fracture technique in the murine femur.MethodsThe mouse femur was surgically stabilized with an intramedullary threaded rod with plastic spacers and the defect adjusted to different sizes. Healing of the different defects was analyzed by radiology and histology to 8 weeks postsurgery. To determine whether this model was effective for evaluating the benefits of molecular therapy, BMP‐2 was applied to the defect and healing then examined.ResultsIntramedullary spacers were effective in maintaining the defect. Callus bone formation was initiated but was arrested at defect sizes of 2.5 mm and above, with no more progress in callus bone development evident to 8 weeks healing. Cartilage development in a critical size defect attenuated very early in healing without bone development, in contrast to the closed femur fracture healing, where callus cartilage was replaced by bone. BMP‐2 therapy promoted osteogenesis of the resident cells of the defect, but there was no further callus development to indicate that healing to pre‐surgery bone structure was successful.ConclusionsThis segmental defect adaptation of the closed femur fracture model of murine bone repair severely impairs callus development and bone healing, reflecting a challenging bone injury. It is adjustable and can be compared to the closed fracture model to ascertain healing deficiencies and the efficacy of therapeutic approaches.

Cordycepin promotes osteogenesis of bone marrow-derived mesenchymal stem cells and accelerates fracture healing via hypoxia in a rat model of closed femur fracture

Fracture is the most frequently encountered traumatic large-organ injury observed in human patients. Cordycepin possesses beneficial effects in osteogenesis of mesenchymal stem cells (MSCs), but its effect on fracture healing is largely unknown. A rat model of closed femur fracture was established, and treated with therapy using bone marrow-derived MSCs (BMMSCs). The effect of cordycepin on the osteogenic process of BMMSCs in vitro was evaluated by Alizarin Red S (ARS) staining and expressions of osteogenic marker genes. Radiographic evaluations and four-point bending mechanical testing were performed on model rats after BMMSC treatment, to assess the effect of cordycepin on fracture healing. Cordycepin promoted osteogenesis of BMMSCs in vitro, and enhanced radiographic parameters and mechanical properties in rat closed femur fracture model using BMMSC therapy in vivo. A hypoxia inhibitor echinomycin could negate the above-mentioned therapeutic effects of cordycepin, indicating that the beneficial effects of cordycepin were mediated via hypoxic response pathway. This study demonstrates that cordycepin promotes osteogenesis of BMMSCs and accelerates fracture healing via hypoxia in a rat model of closed femur fracture, and proposes the clinical potential of cordycepin in bone fracture treatments.

Inducible expression of Wnt7b promotes bone formation in aged mice and enhances fracture healing

There remain unmet clinical needs for safe and effective bone anabolic therapies to treat aging-related osteoporosis and to improve fracture healing in cases of nonunion or delayed union. Wnt signaling has emerged as a promising target pathway for developing novel bone anabolic drugs. Although neutralizing antibodies against the Wnt antagonist sclerostin have been tested, Wnt ligands themselves have not been fully explored as a potential therapy. Previous work has demonstrated Wnt7b as an endogenous ligand upregulated during osteoblast differentiation, and that Wnt7b overexpression potently stimulates bone accrual in the mouse. The earlier studies however did not address whether Wnt7b could promote bone formation when specifically applied to aged or fractured bones. Here we have developed a doxycycline-inducible strategy where Wnt7b is temporally induced in the bones of aged mice or during fracture healing. We report that forced expression of Wnt7b for 1 month starting at 15 months of age greatly stimulated trabecular and endosteal bone formation, resulting in a marked increase in bone mass. We further tested the effect of Wnt7b on bone healing in a murine closed femur fracture model. Induced expression of Wnt7b at the onset of fracture did not affect the initial cartilage formation but promoted mineralization of the subsequent bone callus. Thus, targeted delivery of Wnt7b to aged bones or fracture sites may be explored as a potential therapy.

Investigating the Therapeutic Potential of a Probiotic in a Rat Model for Infection Following Fracture Fixation

Background: Staphylococcus aureus (S. aureus) is the most common pathogen responsible for osteomyelitis. Objectives: Our objective was to investigate the potential of a probiotic as a treatment for S. aureus-induced infection following fracture fixation in a rat model. Methods: Fifty male Sprague-Dawley rats were assigned to five groups (Control, S. aureus, S. aureus +ceftriaxone, S. aureus + once weekly probiotic, and S. aureus + twice weekly probiotic). Lactobacillus casei subsp. casei (ATCC: 39392) was selected from eight strains of probiotic bacteria with anti-staphylococcal activity. Infection was induced by inoculation with106 colony-forming units (CFU) of S. aureus in a closed femur fracture model stabilized with an intramedullary pin. Three weeks after the surgery, the development of infection and response to the therapy was documented using radiographs, microbiological and histopathological analysis. Results: No bacteria were recovered from rats in the Control group. The analysis of variance revealed a significant difference in the CFU/femur (P < 0.001) and CFU/pin (P = 0.001) across all five treatment groups. When the results were compared, the CFU/femur was significantly lower in the S. aureus + Probiotic twice weekly in comparison with S. aureus (P = 0.008) and the S. aureus + ceftriaxone (P = 0.012) groups. Repeated measure ANOVA to test the radiographic scores during the follow-up time between the intervention groups revealed no significant differences (P = 0.179). Conclusions: Parenteral administration of viable L. casei inhibits S. aureus-induced infection as shown by the bacteriologic analysis, but makes no difference to the radiological union rates. This could be the first step towards developing an effective, biologic adjunctive therapy for the management of osteomyelitis following fracture fixation.

Obesity does not affect the healing of femur fractures in mice

Obesity is reported to be both protective and deleterious to bone. Lipotoxicity and inflammation might be responsible for bone loss through inhibition of osteoblasts and activation of osteoclasts. However, little is known whether obesity affects the process of fracture healing. Therefore, we studied the effect of high fat diet-induced (HFD) obesity on callus formation and bone remodelling in a closed femur fracture model in mice. Thirty-one mice were fed a diet containing 60kJ% fat (HFD) for a total of 20 weeks before fracture and during the entire postoperative observation period. Control mice (n=31) received a standard diet containing 10kJ% fat. Healing was analyzed using micro-CT, biomechanical, histomorphometrical, immunohistochemical, serum and protein biochemical analysis at 2 and 4 weeks after fracture. HFD-fed mice showed a higher body weight and increased serum concentrations of leptin and interleukin-6 compared to controls. Within the callus tissue Western blot analyses revealed a higher expression of transcription factor peroxisome proliferator-activated receptor y (PPARy) and a reduced expression of runt-related transcription factor 2 (RUNX2) and bone morphogenetic protein (BMP)-4. However, obesity did not affect the expression of BMP-2 and did not influence the receptor activator of nuclear factor κB (RANK)/RANK ligand/osteoprotegerin (OPG) pathway during fracture healing. Although the bones of HFD-fed animals showed an increased number of adipocytes within the bone marrow, HFD did not increase callus adiposity. In addition, radiological and histomorphometric analysis could also not detect significant differences in bone formation between HFD-fed animals and controls. Accordingly, HFD did not affect bending stiffness after 2 and 4 weeks of healing. These findings indicate that obesity does not affect femur fracture healing in mice.

Positive effect of tadalafil, a phosphodiesterase-5 inhibitor, on fracture healing in rat femur.

This study aims to investigate whether tadalafil accelerates fracture healing with an efficiency equal to that of sildenafil. Fracture healing was studied using a rat closed femur fracture model. Forty-eight male Wistar albino rats (mean age 13 weeks; range 12 to 14 weeks) were divided into three groups: 16 received sildenafil (5 mg/kg/day per oral), 16 received tadalafil (1 mg/kg/day per oral), and 16 (control group) received saline daily. After two and five weeks, eight rats from each group were euthanized and bone healing was evaluated using radiographic, histologic, and histomorphometric analyses. At second week, radiologic score of tadalafil group was higher than the sildenafil treated groups and the sildenafil and tadalafil groups both had higher radiographic scores than the control group, all groups had similar scores at fifth week. The sildenafil and tadalafil treated group exhibited smaller callus diameter at the second week comparing to control group but the difference was not statistically significant, while callus diameter was significantly smaller in tadalafil group when compared to the control group in fifth week. Compared with the control group, cartilage percentage of the callus was lower in the sildenafil group at two weeks and lower in both the tadalafil and sildenafil groups at five weeks. There were no differences between the tadalafil and sildenafil groups regarding the radiographic scores, callus size, cartilage and osseous callus percentage, and the qualitative bridging scores at second or fifth week. This study demonstrated that tadalafil accelerates fracture healing by enhancing osseous tissue formation similar to sildenafil.

Botulinum Toxin-induced Muscle Paralysis Inhibits Heterotopic Bone Formation.

Short-term muscle atrophy induced by botulinum toxin A (BTxA) has been observed to impair osteogenesis in a rat closed femur fracture model. However, it is unclear whether the underlying mechanism is a direct effect of BTxA on muscle-bone interactions or an indirect effect that is driven by skeletal unloading. Because skeletal trauma in the closed fracture model also leads to disuse atrophy, we sought to mitigate this confounding variable by examining BTxA effects on muscle-bone interactions in two complementary in vivo models in which osteogenesis is induced in the absence of skeletal unloading. The overall aim of this study was to identify a potential strategy to inhibit pathological bone formation and heterotopic ossification (HO). (1) Does muscle paralysis inhibit periosteal osteogenesis induced by a transcortical defect? (2) Does muscle paralysis inhibit heterotopic bone formation stimulated by intramuscular bone morphogenetic protein (BMP) injection? Focal osteogenesis was induced in the right hindlimb of mice through surgical initiation of a small transcortical defect in the tibia (fracture callus; n = 7/group) or intramuscular injection of BMP-2 (HO lesion; n = 6/group), both in the presence/absence of adjacent calf paralysis. High-resolution micro-CT images were obtained in all experimental groups 21 days postinduction and total volume (ie, perimeter of periosteal callus or HO lesion) and bone volume (calcified tissue within the total volume) were quantified as primary outcome measures. Finally, these outcome measures were compared to determine the effect of muscle paralysis on inhibition of local osteogenesis in both studies. After a transcortical defect, BTxA-treated mice showed profound inhibition of osteogenesis in the periosteal fracture callus 21 days postsurgery compared with saline-treated mice (total volume: 0.08 ± 0.06 versus 0.42 ± 0.11 mm(3), p < 0.001; bone volume: 0.07 ± 0.05 versus 0.32 ± 0.07 mm(3), p < 0.001). Similarly, BMP-2-induced HO formation was inhibited by adjacent muscle paralysis at the same time point (total volume: 1.42 ± 0.31 versus 3.42 ± 2.11 mm(3), p = 0.034; bone volume: 0.68 ± 0.18 versus 1.36 ± 0.79 mm(3), p = 0.045). Our data indicate that BTxA-induced neuromuscular inhibition mitigated osteogenesis associated with both a transcortical defect and BMP-2-induced HO. Focal neuromuscular inhibition represents a promising new approach that may lead to a new clinical intervention to mitigate trauma-induced HO, a healthcare challenge that is severely debilitating for civilian and war-wounded populations, is costly to both the patient and the healthcare system, and currently lacks effective treatments.

Mir-21 overexpressing mesenchymal stem cells accelerate fracture healing in a rat closed femur fracture model.

MicroRNAs are small noncoding RNAs involved in numerous biological processes. Emerging pieces of evidence suggest that microRNAs play important roles in osteogenesis and skeletal homeostasis. Recent studies indicated the significant regulation function of mir-21 in osteogenesis in vitro, but little information is known about its veritable functions in vivo. In the present study, we aimed to investigate the effect of mir-21 intervention on osteogenic differentiation of rats bone marrow derived mesenchymal stem cells (rBMSCs) and repair capacity in rats closed femur fracture model with internal fixation. The results showed that the upregulation of mir-21 not only increased the expression of osteopontin and alkaline phosphatase in rBMSCs but also promoted mineralization in the condition of osteogenic induction. Furthermore, the bone healing properties were also improved in fracture healing model according to the results of micro-CT, mechanical test, and histological analysis. The current study confirms that the overexpression of mir-21 could promote osteogenesis and accelerate bone fracture healing, which may contribute to a new therapeutic way for fracture repair.

Delayed Fracture Healing in Aged Senescence-Accelerated P6 Mice

ABSTRACTBackground: Osteoporosis is characterized by poor bone quality. However, it is still controversially discussed whether osteoporosis compromises fracture healing. Herein, we studied whether the course of healing of a femur fracture is affected by osteoporosis or age. Methods: Using the senescence-accelerated osteoporotic mouse, strain P6 (SAMP6), and a closed femur fracture model, we studied the process of fracture healing in 5- and 10-month-old animals, including biomechanical, histomorphometric, and protein biochemical analysis. Results: In five-month-old osteoporotic SAMP6 mice, bending stiffness, callus size, and callus tissue distribution as well as the concentrations of the bone formation marker osteocalcin and the bone resorption markers tartrate-resistant acid phosphatase form 5b (TRAP) and deoxypyridinoline (DPD) did not differ from that of non-osteoporotic, senescence-resistant, strain 1 (SAMR1) controls. In contrast, femur fractures in 10-month-old SAMP6 mice showed a significantly reduced bending stiffness and an increased callus size compared to fractures in age-matched SAMR1 controls. This indicates a delayed fracture healing in advanced age SAMP6 mice. The delay of fracture healing was associated with higher concentrations of TRAP and DPD. Significant differences in osteocalcin concentrations were not found between SAMP6 animals and SAMR1 controls. Conclusion: In conclusion, the present study indicates that fracture healing in osteoporotic SAMP6 mice is not affected in five-month-old animals, but delayed in animals with an age of 10 months. This is most probably due to the increased osteoclast activity in advanced age SAMP6 animals.

Increased exercise after stable closed fracture fixation does not affect fracture healing in mice

PurposeThe aim of the present study was to evaluate the systemic biological effect of increased exercise on bone repair after stable fracture fixation. MethodsTwo groups of SKH-1h mice were studied. Animals of the first group (n=36) were housed in cages supplied with a running wheel, while mice of the second group (n=37) were housed in standard cages for control. Using a closed femur fracture model, bone repair was analysed by histomorphometry and biomechanical testing at 2 and 5 weeks. At 2 weeks, we additionally evaluated the expression of the proliferation marker PCNA (proliferating cell nuclear antigen) and the angiogenic and osteogenic growth factor VEGF (vascular endothelial growth factor). To standardise the mechanical conditions in the fracture gap, we used an intramedullary compression screw for stable fracture fixation. ResultsEach mouse of the exercise group run a mean total distance of 23.5km after 2 weeks and 104.3km after 5 weeks. Histomorphometric analysis of the size and tissue composition of the callus could not reveal significant differences between mice undergoing exercise and controls. Accordingly, biomechanical testing showed a comparable torsional stiffness, peak rotation angle, and load at failure of the healing bones in the two groups. The expression of PCNA and VEGF did also not differ between mice of the exercise group and controls. ConclusionWe conclude that increased exercise does not affect bone repair after stable fracture fixation.

Sildenafil accelerates fracture healing in mice

Sildenafil, a cyclic guanosine monophosphate (cGMP)-dependent phospodiesterase-5 inhibitor, has been shown to be a potent stimulator of angiogenesis through upregulation of pro-angiogenic factors and control of cGMP concentration. Herein, we determined whether sildenafil also influences angiogenic growth factor expression and bone formation during the process of fracture healing. Bone healing was studied in a murine closed femur fracture model using radiological, biomechanical, histomorphometric, and protein biochemical analysis at 2 and 5 weeks after fracture. Thirty mice received 5 mg/kg body weight sildenafil p.o. daily. Controls (n = 30) received equivalent amounts of vehicle. After 2 weeks of fracture healing sildenafil significantly increased osseous fracture bridging, as determined radiologically and histologically. This resulted in an increased biomechanical stiffness compared to controls. A smaller callus area with a slightly reduced amount of cartilaginous tissue indicated an accelerated healing process. After 5 weeks the differences were found blunted, demonstrating successful healing in both groups. Western blot analysis showed a significantly higher expression of the pro-angiogenic and osteogenic cysteine-rich protein (CYR) 61, confirming the increase of bone formation. We show for the first time that sildenafil treatment accelerates fracture healing by enhancing bone formation, most probably by a CYR61-associated pathway.

Development of a fracture osteomyelitis model in the rat femur

Osteomyelitis contributes significantly to fracture morbidity. Our objective was to develop a model of induced implant-associated osteomyelitis following fracture repair by modifying an existing rat femur fracture model. Thirty male Sprague-Dawley rats were divided into three groups (Control, Staphylococcus aureus, S. aureus + ceftriaxone). The closed femur fracture model (right femur), stabilized with an intramedullary pin, was combined with inoculation of 10(4) colony-forming units (CFU) of S. aureus. Radiographs were obtained immediately after surgery and at weeks 1, 2, and 3 and were evaluated by individuals blinded to treatment group. At necropsy the CFU of S. aureus per femur and pin were determined and synovial tissue and blood were cultured. The fractured femur from two rats in each group was evaluated histologically. A statistically significant difference in the CFU/femur and CFU/pin was found across treatment groups, with the highest CFU in the S. aureus group and the lowest in the Control group. Cultures of synovial tissue were positive in 11/19 of inoculated limbs. Osteomyelitis was present both radiographically and histopathologically in both S. aureus groups but not in the controls. No rats were systemically ill or had positive blood cultures at the study endpoint. This model will be useful for the evaluation of treatments or prophylactics designed for use in implant-associated osteomyelitis.

Rapamycin affects early fracture healing in mice

The immunosuppressive drug rapamycin (RAPA) prevents rejection in organ transplantation by inhibiting interleukin-2-stimulated T-cell division. Rapamycin has also been suggested to possess strong anti-angiogenic activities linked to a decrease in production of vascular endothelial growth factor (VEGF). Angiogenesis and VEGF are thought to play a crucial role in fracture healing and as osteoporotic and traumatic fractures are common complications in immunosuppressed, organ transplantation patients, we conducted this study to analyze the effect of rapamycin treatment on bone repair. We investigated the effect of rapamycin treatment on bone repair in a murine closed femur fracture model using radiological, histomorphometric, immunohistochemical, biomechanical and protein biochemical analyses. X-ray analyses demonstrated that rapamycin treatment inhibits callus formation after two weeks of fracture healing. The radiologically observed lack of callus formation was confirmed by histomorphometric analyses, which revealed a significantly diminished callus size and a reduced amount of bone formation when compared with vehicle-treated controls. Biomechanical testing further demonstrated that rapamycin significantly reduces torsional stiffness of the callus. Interestingly, this effect was associated with decreased vessel formation; a diminished proliferation of osteoblasts, endothelial cells and periosteal cells; and a reduced VEGF expression in hypertrophic chondrocytes. After five weeks treatment, however, the negative impact of rapamycin on fracture healing was overcome. Thus, rapamycin initially delays fracture healing, most probably by inhibiting cell proliferation and neovascularization in the callus. These undesirable effects should be considered when rapamycin is administered to patients sustaining bone fractures.

A Fracture Pain Model in the Rat

Because of the relative lack of understanding of the mechanisms that drive skeletal pain, the purpose of this study was to adapt a previously validated closed femur fracture model to quantitatively evaluate skeletal pain in female and male rats. Three-month-old female and male Sprague-Dawley rats were anesthetized, and a stainless steel pin was inserted into the intramedullary space of the left femur. Three weeks later, the rats were reanesthetized, and left femoral diaphyses were fractured using a standardized impactor device. At 1-21 days after fracture, skeletal pain was measured by quantitatively assessing spontaneous guarding, spontaneous flinching, and weight bearing of the fractured hind limb. Females and males showed highly robust pain behaviors that were maximal at day 1 after fracture and returned gradually to normal nonfractured levels at days 14-21 after fracture. The magnitude of fracture pain was not significantly different at most time points between female and male rats. In both females and males, the pain-related behaviors were attenuated by subcutaneous morphine in a dose-dependent manner. This model may help in developing a mechanism-based understanding of the factors that generate and maintain fracture pain in both females and males and in translating these findings into new therapies for treating fracture pain.

Erythropoietin (EPO) — EPO-receptor signaling improves early endochondral ossification and mechanical strength in fracture healing

Beyond its role in the regulation of red blood cell proliferation, the glycoprotein erythropoietin (EPO) has been shown to promote cell regeneration and angiogenesis in a variety of different tissues. In addition, EPO has been indicated to share significant functional and structural homologies with the vascular endothelial growth factor (VEGF), a cytokine essential in the process of fracture healing. However, there is complete lack of information on the action of EPO in bone repair and fracture healing. Therefore, we investigated the effect of EPO treatment on bone healing in a murine closed femur fracture model using radiological, histomorphometric, immunohistochemical, biomechanical and protein biochemical analysis. Thirty-six SKH1-hr mice were treated with daily i.p. injections of 5000 U/kg EPO from day 1 before fracture until day 4 after fracture. Controls received equivalent amounts of the vehicle. After 2 weeks of fracture healing, we could demonstrate expression of the EPO-receptor (EPOR) in terminally differentiating chondrocytes within the callus. At this time point EPO-treated animals showed a higher torsional stiffness (biomechanical analysis: 39.6 ± 19.4% of the contralateral unfractured femur) and an increased callus density (X-ray analysis (callus density / spongiosa density): 110.5 ± 7.1%) when compared to vehicle-treated controls (14.3 ± 8.2% and 105.9 ± 6.6%; p < 0.05). Accordingly, the histomorphometric examination revealed an increased fraction of mineralized bone and osteoid (33.0 ± 3.0% versus 28.5 ± 3.6%; p < 0.05). Of interest, this early effect of the initial 6-day EPO treatment had vanished at 5 weeks after fracture. We conclude that EPO–EPOR signaling is involved in the process of early endochondral ossification, enhancing the transition of soft callus to hard callus.

Development of a locking femur nail for mice

We herein report on a novel locking intramedullary nail system in a murine closed femur fracture model. The nail system consists of a modified 24-gauge injection needle and a 0.1-mm-diameter tungsten guide wire. Rotation stability was accomplished by flattening the proximal and distal end of the needle. Torsional mechanical testing of the implants in osteotomized cadaveric femora revealed a superiority of the locking nail (3.9±1.0° rotation at a torque of 0.9 Nmm, n = 1 0 ) compared to the unmodified injection needle (conventional nail; 52.4±3.2°, n = 1 0 , p < 0.0 5 ). None of the implants, however, achieved the rotation stability of unfractured femora (0.3±0.5°, n = 1 0 ). In a second step, we tested the feasibility of the in vivo application of the locking nail to stabilize a closed femoral midshaft fracture in C57BL/6 mice. Of interest, none of the 10 animals showed a dislocation of the locking nail over a 5-week period, while 3 of 4 animals with conventional nail fracture stabilization showed a significant pin dislocation within the first 3 days ( p < 0.0 5 ). Mechanical testing after 5-weeks stabilization with the locking nail revealed an appropriate bone healing with a torque at failure of 71.6±3.4% and a peak rotation before failure of 68.4±5.3% relative to the unfractured contralateral femur. With the advantage that closed fractures can be fixed with rotation stability, the herein introduced model may represent an ideal tool to study bone healing in transgenic and knockout mice.

Characterization of a Closed Femur Fracture Model in Mice

The goal of this study was to develop and characterize a closed femur fracture model for mice that can be used for the molecular and genetic analysis of fracture healing. Longitudinal time study of species-specific fracture healing. A protocol was developed for creating reproducible, closed femur fractures in mice. Impending fractures were stabilized by retrograde insertion of a 0.01-inch-diameter, stainless steel wire into the intramedullary canal. The intramedullary wire was held in place with a wedge made from the first 2 mm of a 30-gauge needle. Fractures were produced by 3-point bending. Fracture healing was assessed by radiography, histology, and torsional mechanical testing. The mouse femur fracture technique produced good results with minimal loss of animals. Of the 246 mice used in the study, 22 mice were excluded due to poor fracture quality (8), loss of fracture stabilization (6), or to anesthesia death (8). Radiography showed a consistent pattern of fracture healing between mice with peak fracture callus volume evident at 10 (15 mice) to 14 days (18 mice) after fracture. Fracture bridging was apparent in all 3-week postfracture radiographs (35 mice). Histologic examination of 117 specimens at 9 time points showed chondrocyte differentiation within the fracture callus by 7 days after fracture, endochondral ossification occurring by 10 days after fracture, and bone remodeling evident as early as 3 weeks after fracture. Despite radiologic and histologic evidence of fracture bridging after 3 weeks, torsional mechanical testing of 68 mice at 3, 4, 6, and 12 weeks after fracture (group size of 15 to 18 mice at each time point) indicated that significant increases in structural or material strength did not occur until 6 to 12 weeks after fracture. Femur fracture healing in mice follows a typical endochondral ossification pathway with fracture bridging occurring approximately 1 week faster in mice than rats. This fracture model is amenable to the molecular and genetic analysis of fracture healing using different inbred, transgenic, and knockout strains of mice.

Testosterone is a Potent Accelerator of Fracture Healing: Early Structural Reconstruction and Improved Biomechanical Stability

Sex steroids (e. g., estrogen and testosterone) are known to have an integral function in skeletal homeostasis. Though they are known to improve bone strength, their role in fracture healing has not been well defined. The aim of this study was to examine the influence of testosterone on skeletal repair mechanisms, in order to test the hypothesis that sex steroids could be used to accelerate fracture healing after intramedullary nailing. Fracture healing was studied in 60 eugonadal female and male mice treated either with testosterone or placebo. To make mouse models accessible to fracture modulation studies that also allow biomechanical analysis, we modified the Einhorn tibial fracture model and developed a standardized closed femur fracture model. Mice treated with testosterone showed a significant acceleration of fracture healing. Callus size as assessed by contact-radiography and microCT was significantly enhanced. This beneficial effect could be demonstrated in female as well as in male mice. Furthermore, not only radiological and histological data demonstrated a positive morphological interference of testosterone with the fracture healing process, but also the functional competence, as assessed by three-point bending tests, was dramatically improved. This study shows that testosterone can be used to enhance and accelerate fracture healing.