Radiomorphometric Quantitative Analysis of Vasculature Utilizing Micro–Computed Tomography and Vessel Perfusion in the Murine Mandible

Abstract

Purpose Biomechanical, densitometric, and histological analyses have been the mainstay for reproducible outcome measures for investigation of new bone formation and osseous healing. Here we report the addition of radiomorphometric vascular analysis as a quantitative measure of vascularity in the murine mandible. To our knowledge this is the first description of using micro–computed tomography (micro-CT) to evaluate the temporal and spatial pattern of angiogenesis in the craniofacial skeleton. Methods The vessel perfusion technique was performed on 10 Sprague-Dawley rats using Microfil (MV-122, Flow Tech; Carver, MA). After decalcification, hemimandibles were imaged using high-resolution micro-CT. Six separate radiomorphometric vascular metrics were calculated. Results Radiomorphometric values were analyzed using three different thresholds on micro-CT. Experimentally, 1000 Hounsfield units was found to be the optimal threshold for analysis to capture the maximal vascular content of the bone. Data from seven hemimandibles were analyzed. Minimal statistical variance in each of the quantitative measures of vascularity resulted in reproducible metrics for each of the radiomorphometric parameters. Conclusions We have demonstrated that micro-CT vascular imaging provides a robust methodology for evaluation of vascular networks in the craniofacial skeleton. This technique provides 3D quantitative data analysis that differs significantly from laser Doppler and microsphere methods, which simply measure flow. This technique is advantageous over labor-intensive 2D conventional analyses using histology and X-ray microangiography. Our data establish the appropriate thresholding for optimal vascular analyses and provide baseline measurements that can be used to analyze the role of angiogenesis in bone regeneration and repair in the craniofacial skeleton.