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Mice are extensively used for experimental bone-healing studies. However, there are few established nondestructive  in vivo  techniques for longitudinal fracture-healing analysis in mice, including  in vivo  micro-computed tomography (μCT) and radiography. Importantly, none of the established methods can discriminate between non-mineralized fibrous tissue and cartilage in the soft fracture callus. Therefore, the objective was to establish high-resolution  in vivo  magnetic resonance imaging (MRI) for the longitudinal assessment of soft callus formation during bone healing in mice. C57BL/6J mice received a femur osteotomy stabilized using an external fixator and were randomly assigned to five groups. Group 1 mice were scanned three times longitudinally during fracture healing using an optimized MRI scanning protocol to establish an algorithm to characterize the different fracture-callus tissues. Mice of groups 2–4 were scanned once on day 10, 14 or 21, respectively, euthanized after scanning and their femurs subjected to  ex vivo  μCT and histomorphometric analysis to compare the data assessed by MRI with μCT and histology. Control group 5 mice were not scanned. After 28 days, mice of groups 1 and 5 were euthanized and the fracture-healing outcome was evaluated by bending-test, μCT and histology to determine whether the repeated anesthesia, handling and the MRI measurements themselves influenced fracture healing. The callus-tissue values determined by MRI were mostly comparable to those obtained by μCT and histomorphometric analysis. However, at time points characterized by small relative bone or cartilage areas, MRI measurements were weakly comparable to histomorphometric data, possibly due to the inferior spatial resolution. Importantly, at the early and intermediate phases of healing, cartilage and fibrous-tissue values obtained by MRI were highly accurate. Furthermore, repeated anesthesia, handling and MRI scans did not impact bone healing. Therefore, we demonstrated the feasibility of high-resolution  in vivo  MRI for longitudinal assessment of soft callus formation during murine endochondral fracture healing.

Evaluation of high-resolution In Vivo MRI for longitudinal analysis of endochondral fracture healing in mice