A New Method for Segmentation and Analysis of Bone Callus in Rodent Fracture Models using micro-CT.

Fracture burden has created a need to better understand bone repair processes under different pathophysiological states. Evaluation of structural and material properties of the mineralised callus, which is integral to restoring biomechanical is therefore, vital. Micro-CT can facilitate non-invasive imaging of fracture repair, however, current methods for callus segmentation are only semi-automated, restricted to defined regions, time/labour intensive and prone to user-variation. Herein, we share a new automatic method for segmenting callus in micro-CT tomograms that will allow for objective, quantitative analysis of the bone fracture microarchitecture. Fractured and non-fractured mouse femurs were scanned and processed by both manual and automated segmentation of fracture callus from cortical bone after which microarchitectural parameters were analysed. All segmentation and analysis steps were performed using CTAn (Bruker, Belgium) with automatic segmentation performed using the software's image processing plugins. Results showed automatic segmentation reliably and consistently segmented callus from cortical bone, demonstrating good agreement with manual methods with low bias: tissue volume: -0.320mm 3 , bone volume: 0.0358mm 3 and bone volume/tissue volume: -3.52%, and was faster and eliminated user-bias and variation. Method scalability and translatability across rodent models was verified in scans of fractured rat femora showing good agreement with manual methods with low bias: tissue volume: -3.654mm 3 , bone volume: 0.830mm 3 and bone volume/tissue volume: 7.81%. Together, these data validate a new automated method for segmentation of callus and cortical bone in micro-CT tomograms that we share as a fast, reliable and less user-dependent tool for application to study bone callus in fracture, and potentially elsewhere. This article is protected by copyright. All rights reserved.