Quantitative measures of femoral fracture repair in rats derived by micro-computed tomography

Although fracture healing is frequently studied in pre-clinical models of long bone fractures using rodents, there is a dearth of objective quantitative techniques to assess successful healing. Biomechanical testing is possibly the most quantitative and relevant to a successful clinical outcome, but it is a destructive technique providing little insight into the cellular mechanisms associated with healing. The advent of X-ray computed tomography (CT) has provided the opportunity to quantitatively and non-destructively assess bone structure and density, but it is unknown how measurements derived using this technology relate to successful healing. To examine possible relationships, we used a pre-clinical model to test for statistically significant correlations between quantitative characteristics of the callus by micro-CT (microCT) and the bending strength, stiffness, and energy-to-failure of the callus as assessed by three-point bending of excised bones. A closed, transverse fracture was generated in the mid-shaft of rat femurs by impact loading. Shortly thereafter, the rats received a one-time, local injection of either the vehicle or one of four doses of lovastatin. Following sacrifice after 4 weeks of healing, fractured femurs were extracted for microCT analysis and then three-point bending. Setting the region of interest to be 3.2 mm above and below the fracture line, we acquired standard and new microCT-derived measurements. The mineralized callus volume and the mineral density of the callus correlated positively with callus strength (rxy = -0.315, p = 0.016 and rxy = 0.444, p<0.0005, respectively) and stiffness (rxy = -0.271, p = 0.040 and rxy = 0.325, p = 0.013, respectively), but the fraction of the callus that mineralized and the moment of inertia of the callus did not. This fraction did correlate with energy-to-failure (rxy = -0.343, p = 0.0085). Of the microCT-derived measurements, quantifying defects within the outer bridging cortices of the callus produced the strongest correlation with both callus strength (rxy = 0.557, p<0.0001) and stiffness (rxy = 0.468, p = 0.0002). By both reducing structural defects and increasing mineralization, lovastatin appears to increase the callus strength.