Radiomorphometric quantitative analysis of vasculature utilizing micro-computed tomography and vessel perfusion in the murine mandible.

To better understand the process of bone healing as well as the pathological conditions that impact bony union, a comprehensive grasp of blood supply is of paramount importance. The significance of the role angiogenesis plays in osteogenesis has been well appreciated since the 18th century when Albrecht von Haller wrote in his book Experimentorum de ossium formatione (1763): “…the origin of bone is the artery carrying the blood and in it the mineral elements.”1(p400) The blood supply of the mandible has certain unique features that are important for experimental procedures and for the study of bone healing. Our laboratory has been actively engaged in building a foundation of analytical and developmental research that provides both the intellectual and methodological basis for the study of new bone formation and osseous healing. Early works on angiogenesis during new bone formation and osseous healing have utilized a variety of techniques to evaluate vascular structures in animal models. Histology is the most commonly used method to analyze vascularity.2 Histological examination of blood vessels, however, is limited in its accuracy and reproducibility because of the subjectivity of the examiner, the problem of precisely accounting for the cross sectioning of the serpentine course of vasculature throughout the tissue samples, and the incomplete sampling involved in histologic analysis. Verdonck et al utilized laser Doppler flow to assess bone vascularity in the edentulous human mandible after oral implant.3 Laser Doppler imaging (LDI) uses a laser scan to provide a color-coded, 2D image of tissue perfusion without disturbing the tissue of interest. This technique provides semiquantitative data and is more a measure of functionality. LDI, however, does not provide anatomic information at all and is limited by the fact that only the most superficial blood flow is measured. LDI cannot be used dynamically to image high-frequency blood flow fluctuations and also takes a long time to scan. Finally, LDI is operator dependent and provides low image resolution. Another popular technique, X-ray microangiography, provides high-resolution 2D angiograms of the vascular network but lacks the ability to employ a quantitative, volumetric analysis.4,5 Micro–computed tomography (micro-CT) imaging with perfused contrast agents has emerged as a promising imaging technology to visualize 3D vascular networks in organs or tumors of small animals.6 Previous studies from our laboratory have focused on utilizing biomechanical, densitometric, and histological analysis as the mainstay for reproducible outcome measures for investigation of new bone formation and osseous healing.7 Our goal now is to investigate the feasibility of using micro-CT imaging with a perfused contrast agent as a quantitative measure of vascularity and vasculogenesis in the murine mandible. To our knowledge this is the first description of this technique in the mandible and will lay down the groundwork for using micro-CT to quantitatively evaluate the temporal and spatial pattern of angiogenesis during distraction osteogenesis (DO) as well as other forms of bone regeneration and repair in the craniofacial skeleton.