Age-related changes to macrophages are detrimental to fracture healing in mice.

The elderly population suffers from higher rates of complications during fracture healing that result in increased morbidity and mortality. Inflammatory dysregulation is associated with increased age and is a contributing factor to the myriad of age-related diseases. Therefore, we investigated age-related changes to an important cellular regulator of inflammation, the macrophage, and the impact on fracture healing outcomes. We demonstrated that old mice (24 months) have delayed fracture healing with significantly less bone and more cartilage compared to young mice (3 months). The quantity of infiltrating macrophages into the fracture callus was similar in old and young mice. However, RNA-seq analysis demonstrated distinct differences in the transcriptomes of macrophages derived from the fracture callus of old and young mice, with an up-regulation of M1/pro-inflammatory genes in macrophages from old mice as well as dysregulation of other immune-related genes. Preventing infiltration of the fracture site by macrophages in old mice improved healing outcomes, with significantly more bone in the calluses of treated mice compared to age-matched controls. After preventing infiltration by macrophages, the macrophages remaining within the fracture callus were collected and examined via RNA-seq analysis, and their transcriptome resembled macrophages from young calluses. Taken together, infiltrating macrophages from old mice demonstrate detrimental age-related changes, and depleting infiltrating macrophages can improve fracture healing in old mice

Impact of osteoporosis and osteoporosis medications on fracture healing: a narrative review

UNLABELLED Antiresorptive medications do not negatively affect fracture healing in humans. Teriparatide may decrease time to fracture healing. Romosozumab has not shown a beneficial effect on human fracture healing. BACKGROUND Fracture healing is a complex process. Uncertainty exists over the influence of osteoporosis and the medications used to treat it on fracture healing. METHODS Narrative review authored by the members of the Fracture Working Group of the Committee of Scientific Advisors of the International Osteoporosis Foundation (IOF), on behalf of the IOF and the Société Internationale de Chirurgie Orthopédique et de Traumatologie (SICOT). RESULTS Fracture healing is a multistep process. Most fractures heal through a combination of intramembranous and endochondral ossification. Radiographic imaging is important for evaluating fracture healing and for detecting delayed or non-union. The presence of callus formation, bridging trabeculae, and a decrease in the size of the fracture line over time are indicative of healing. Imaging must be combined with clinical parameters and patient-reported outcomes. Animal data support a negative effect of osteoporosis on fracture healing; however, clinical data do not appear to corroborate with this. Evidence does not support a delay in the initiation of antiresorptive therapy following acute fragility fractures. There is no reason for suspension of osteoporosis medication at the time of fracture if the person is already on treatment. Teriparatide treatment may shorten fracture healing time at certain sites such as distal radius; however, it does not prevent non-union or influence union rate. The positive effect on fracture healing that romosozumab has demonstrated in animals has not been observed in humans. CONCLUSION Overall, there appears to be no deleterious effect of osteoporosis medications on fracture healing. The benefit of treating osteoporosis and the urgent necessity to mitigate imminent refracture risk after a fracture should be given prime consideration. It is imperative that new radiological and biological markers of fracture healing be identified. It is also important to synthesize clinical and basic science methodologies to assess fracture healing, so that a convergence of the two frameworks can be achieved

VEGF with AMD3100 Endogenously Mobilizes Mesenchymal Stem Cells and Improves Fracture Healing

A significant number of fractures develop non‐union. Mesenchymal stem cell (MSC) therapy may be beneficial, however, this requires cell acquisition, culture and delivery. Endogenous mobilization of stem cells offers a non‐invasive alternative. The hypothesis was administration of VEGF and the CXCR4 antagonist AMD3100 would increase the circulating pool of available MSCs and improve fracture healing. Ex‐breeder female wistar rats received VEGF followed by AMD3100, or sham PBS. Blood prepared for culture and colonies were counted. P3 cells were analyzed by flow cytometry, bi‐differentiation. The effect of mobilization on fracture healing was evaluated with 1.5 mm femoral osteotomy stabilized with an external fixator in 12–14 week old female Wistars. The mobilized group had significantly greater number of cfus/ml compared to controls, p = 0.029. The isolated cells expressed 1.8% CD34, 35% CD45, 61% CD29, 78% CD90, and differentiated into osteoblasts but not into adipocytes. The fracture gap in animals treated with VEGF and AMD3100 showed increased bone volume; 5.22 ± 1.7 µm3 and trabecular thickness 0.05 ± 0.01 µm compared with control animals (4.3 ± 3.1 µm3, 0.04 ± 0.01 µm, respectively). Radiographic scores quantifying fracture healing (RUST) showed that the animals in the mobilization group had a higher healing score compared to controls (9.6 vs. 7.7). Histologically, mobilization resulted in significantly lower group variability in bone formation (p = 0.032) and greater amounts of bone and less fibrous tissue than the control group. Clinical significance: This pre‐clinical study demonstrates a beneficial effect of endogenous MSC mobilization on fracture healing, which may have translation potential to prevent or treat clinical fractures at risk of delayed or non‐union fractures

In the quest for the etiology of the delayed union of fractures: the inhibitory role of non-steroidal anti-inflammatory drugs - a complex pharmacological phenomenon?

Failure of fracture healing is one of the problems that clinical orthopaedics face in practice. This review will examine the role of non-steroidal anti-inflammatory drugs in fracture healing. It is believed that they are inhibitors of the early stages of fracture healing process, according to experimental models. Clinical studies in this area are few in number and without clear evidence regarding the inhibitory effect of non-steroidal anti-inflammatory drugs. Surprisingly the new substances of this class of drugs (COX-2 inhibitors) have the same action in fracture healing. Despite the possible adverse effects of non-steroidal anti-inflammatory drugs in gastrointestinal, cardiovascular systems and fracture healing, they are widely used for post-surgery orthopaedic pain and inflammation

The effectiveness and safety of parathyroid hormone in fracture healing: A meta-analysis

The very large economic and social burdens of fracture-related complications make rapid fracture healing a major public health goal. The role of parathyroid hormone (PTH) in treating osteoporosis is generally accepted, but the effect of PTH on fracture healing is controversial. This meta-analysis was designed to investigate the efficacy and safety of PTH in fracture healing. The EMBASE, PubMed, and Cochrane Library databases were systematically searched from the inception dates to April 26, 2018. The primary randomized clinical trials comparing PTH treatment for fracture healing with placebo or no treatment were identified. We did not gain additional information by contacting the authors of the primary studies. Two reviewers independently extracted the data and evaluated study quality. This meta-analysis was executed to determine the odds ratio, mean difference, standardized mean difference, and 95% confidence intervals with random-effects models. In total, 8 randomized trials including 524 patients met the inclusion criteria. There were significant differences in fracture healing time, pain relief and function improvement. There were no significant differences in the fracture healing rate or adverse events, including light-headedness, hypercalcemia, nausea, sweating and headache, except for slight bruising at the injection site. We determined that the effectiveness and safety of PTH in fracture healing is reasonably well established and credible

Application of Salubrinal for Bone Fracture Healing

abstractThe long-term objective of this project is to commercialize a novel synthetic chemical agent, salubrinal, for treatment of bone growth and fracture healing. Bone morphogenetic proteins (BMPs) are clinically administered as growth stimulators for bone fracture healing. However, BMPs are not only expensive, but also stimulate ectopic bone formation and potentially induce cancer. A synthetic chemical agent that permits facile storage and administration could reduce costs, and provide longer shelf-life, and better bone healing outcomes. Currently, no synthetic chemical agents as a stimulator of fracture healing are clinically available. The research team recently identified “salubrinal,” a synthetic chemical agent, as a potential therapeutic stimulator of bone growth and fracture healing. An invention disclosure and a U.S. patent were filed. In this FORCES project, we are examining efficacy of salubrinal using a mouse model of closed tibia fracture. The results strongly indicate that salubrinal can accelerate bone fracture healing

Effects of Eurycoma longifolia on fracture healing of androgen-deficient osteoporosis model: a micro computed tomograph analysis

Micro computed tomography (micro-CT) imaging is a useful tool to monitor fracture healing in osteoporosis model. It creates a 3-D image of the fracture callus which can be analysed to assess bone parameters quantitatively. In this study, micro-CT was used to assess the fracture healing of orchidectomised rats, an androgen-deficient osteoporosis model. The effects of Eurycoma longifolia, a medicinal plant with pro-androgenic effects, on fracture healing were assessed. The rats were grouped into orchidectomised-control (ORX), sham-operated (SHAM), orchidectomised and injected with testosterone intramuscularly once weekly (TEN) and orchidectomised and daily oral gavage of Eurycoma longifolia (EL). Treatment duration was six weeks following bone fracture. Fracture was induced in the right tibia of all the rats. A total of 100 axial slices above and below fracture line were scanned with a micro-CT. The micro-CT analysis was able to detect significant difference in the fracture healing rate of ORX and TEN groups. The bridging cortices and fraction of mineralized tissue of the bridging cortices of the callous of ORX group was significantly lower than TEN group. No significant micro-CT changes were seen in the fracture healing of the EL group. The effect of EL on fracture healing was not demonstrable in orchidectomised rat model

Transcriptional Analysis of Fracture Healing and the Induction of Embryonic Stem Cell–Related Genes

Fractures are among the most common human traumas. Fracture healing represents a unique temporarily definable post-natal process in which to study the complex interactions of multiple molecular events that regulate endochondral skeletal tissue formation. Because of the regenerative nature of fracture healing, it is hypothesized that large numbers of post-natal stem cells are recruited and contribute to formation of the multiple cell lineages that contribute to this process. Bayesian modeling was used to generate the temporal profiles of the transcriptome during fracture healing. The temporal relationships between ontologies that are associated with various biologic, metabolic, and regulatory pathways were identified and related to developmental processes associated with skeletogenesis, vasculogenesis, and neurogenesis. The complement of all the expressed BMPs, Wnts, FGFs, and their receptors were related to the subsets of transcription factors that were concurrently expressed during fracture healing. We further defined during fracture healing the temporal patterns of expression for 174 of the 193 genes known to be associated with human genetic skeletal disorders. In order to identify the common regulatory features that might be present in stem cells that are recruited during fracture healing to other types of stem cells, we queried the transcriptome of fracture healing against that seen in embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs). Approximately 300 known genes that are preferentially expressed in ESCs and ∼350 of the known genes that are preferentially expressed in MSCs showed induction during fracture healing. Nanog, one of the central epigenetic regulators associated with ESC stem cell maintenance, was shown to be associated in multiple forms or bone repair as well as MSC differentiation. In summary, these data present the first temporal analysis of the transcriptome of an endochondral bone formation process that takes place during fracture healing. They show that neurogenesis as well as vasculogenesis are predominant components of skeletal tissue formation and suggest common pathways are shared between post-natal stem cells and those seen in ESCs