Microfocal CT: a method for evaluating murine cranial sutures in situ

Introduction. The murine model is a well- established surrogate for studying human cranial su- ture biology. In mice, all sutures with the exception of the posterior frontal (PF) suture remain patent throughout life. Histology is regarded as the gold stan- dard for analyzing sutures. On this basis, PF suture fusion begins on day of life 25 and is complete by day 45. Cranial suture histology, however, requires sacri- fice of the animal to obtain tissue for analysis. As a result, knowledge of the kinetics of cranial suture fu- sion is based on a patchwork analysis of many sutures from many different animals. The behavior of a single suture through time is unknown. Our goal is to de- velop a noninvasive means to repeatedly image mouse cranial sutures in vivo. As a first step, the present study was performed to evaluate microfocal computer tomography (micro-CT) technology for the use of cap- turing images of a mouse cranium in situ. Methods. The micro-CT system consists of a microfo- cal X-ray source and a large format CCD camera opti- cally coupled to a high-resolution X-ray image inten- sifier, digitally linked to a computer. The PF and sagittal sutures lie in continuity along the midline of the skull. Holes were drilled in the calvaria on both sides of the PF and sagittal sutures of a 45-day-old euthanized mouse. A micro-CT scan of this animal was performed and hundreds of cross-sectional images were generated for the cranium. These images were used to reconstruct three-dimensional volumetric im- ages of the entire cranium. Comparisons were made between (1) the gross specimen and the three dimen- sional reconstructions; (2) two-dimensional coronal images obtained by micro-CT and those obtained by histology. Results. Analysis of PF and sagittal sutures demon- strated the following: (1) The drilled holes were accu- rately rendered by micro-CT, when compared to both the gross specimen and the histology. (2) The sagittal suture was found to be patent by both micro-CT and histology. (3) The PF suture is fused by histology, but unexpectedly, the PF suture appears incompletely fused by micro-CT. By micro-CT, however, the anterior and endocranial regions appear more extensively fused than the remainder of the PF suture, a finding consistent with published histologic analysis. Conclusions. We successfully imaged 45-day-old mouse cranial sutures in situ using micro-CT technol- ogy. Precise correlation between histologic sections and radiologic images is difficult, but convincing sim- ilarities exist between the gross specimen and images from micro-CT and histology. PF suture fusion in a 45-day-old animal appears different by micro-CT than by histology. One possible explanation for this appar- ent discrepancy is that suture fusion in histology is determined based on the appearance of bone morphol- ogy and not tissue density, as the specimens are nec- essarily decalcified to section the bone. Micro-CT, on the other hand, distinguishes tissues on the basis of density. Newly forming bone may require bone matrix formation prior to complete calcification; PF suture in 45-day-old mice may be morphologically complete but incompletely ossified. Studies correlating histologic and micro-CT assessment of suture development are underway. Micro-CT appears to be a promising method for noninvasive imaging of mouse cranial