Comparison of micro-CT image enhancement after use of different vascular casting agents

Microvascular visualization is crucial in understanding the mechanisms of several pathologies. For instance, visualization of the tumor microenvironment is important in understanding angiogenesis and role in cancer progression. Visualization would provide insights to cancer diagnosis, predicting metastatic growth, and evaluating therapeutic protocols. Similarly, understanding the microvascular network could be beneficial for study of degenerative diseases and tissue repair. The use of microscale computed tomography (micro-CT) and vascular casting agents provides high-resolution images of tissue vasculature in volumetric space. The purpose of this research was to compare a selection of commercially available contrast agents to determine the optimal solution for vascular visualization.

A population of 16 female nude athymic mice (Charles Rivers Laboratories) were implanted with MDA-MB-231 breast cancer cells (ATCC) orthotopically in the lower left mammary fat pad to investigate the tumor microenvironment. Once tumors reach sufficient size, animals were equally divided into four groups based on the micro-CT agent to be administered, namely, control (no contrast agent), barium sulfate (BaSO₄), Vascupaint, or Microfil. Animals were anesthetized prior to transcarotid micro-cannulation to infuse 2 mL of the specific contrast agent for intravascular distribution throughout the animal. The jugular vein on the other side of the carotid artery was opened to drain blood flow. Following successful perfusion, animals and extracted organs underwent high-resolution micro-CT scanning (OI/CT, MILabs). Images were reconstructed and analyzed using analysis software to extract mean intensity signals.

Preliminary post-mortem micro-CT results reveal Vascupaint and BaSO₄ are useful for microvascular visualization. Both Vascupaint and BaSO₄ produced significant contrast-enhanced micro-CT image enhancement in the brain (3.39±0.93 and 6.27±3.78, respectively) and kidney (12.85±1.98 and 32.87±10.03, respectively) as compared to Microfil (0.22±0.07 and 0.91±0.63, respectively; P<0.05). For the various contrast agents, there were no differences in image enhancement from the liver, spleen, or tumor tissue (P>0.21). Moreover, use of Vascupaint and BaSO₄ allowed for visualization of smaller microvascular structures with average diameters of 20.54±4.15 and 25.82±3.75 µm, which were smaller compared to the 91.66±24.91 µm measurements from Microfil-enhanced micro-CT images (P<0.004).

Our study suggests that the use of Vascupaint and BaSO₄ is more than sufficient for ex vivo visualization of microvascular structures with contrast-enhanced micro-CT imaging as these contrast agents more effectively perfused smaller blood vessels.