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 µm 3 and trabecular thickness 0.05 ± 0.01 µm compared with control animals (4.3 ± 3.1 µm 3 , 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. © 2018 The Authors. Journal of Orthopaedic Research ® Published by Wiley Periodicals, Inc. on behalf of Orthopaedic Research Society. J Orthop Res 37:1294–1302, 2019.

A significant number of fractures develop nonunion. Stem cell homing is regulated through stromal cell-derived factor 1 (SDF1) and its receptor CXCR4. Stem/progenitor cell populations can be endogenously mobilized by administering growth factors with a pharmacological antagonist of CXCR4, AMD3100, which may be a means to improve fracture healing. A 1.5 mm femoral osteotomy in Wistar rats was stabilized with an external fixator. Rats were pretreated with phosphate buffered saline [PBS(P)], vascular endothelial growth factor [VEGF(V)], insulin-like growth factor-1 [IGF1(I)], or granulocyte colony stimulating factor [GCSF(G)] before AMD3100. A control group (C) did not receive growth factors or AMD3100. Bone formation after 5 weeks was analyzed. Group P had a significant increase in total bone volume (BV) (p = 0.01) and group I in percentage bone in the fracture gap (p = 0.035). Group G showed a decrease in BV. All treated groups had an increase in trabecular thickness. Histology showed decreased cartilage tissue associated with increased bone in groups with improved healing, and increased fibrous tissue in poorly performing groups. Antagonism of SDF1-CXCR4 axis can boost impaired fracture healing. AMD3100 given alone was the most effective means to boost healing, whereas pretreatment with GCSF reduced healing. AMD3100 is likely mobilizing stem cells into the blood stream that home to the fracture site enhancing healing.Impact StatementCurrently ∼10% of fractures progress to delayed or nonunion with significant morbidity and economic impact. Endogenous mobilization of stem cells by pharmacological antagonism of their homing and migration receptor CXCR4 with AMD3100 experimentally reduced delayed union development. Endogenous mobilization may, therefore, translate as a low risk means to boost healing and could potentially be given as a prophylaxis to patients with fractures at risk of delayed healing or nonunion. These patients may include fragility fractures, comminuted tibial fractures, or when treating established nonunions. This approach could have promise for other conditions that may benefit from stem cell treatments.