Molecular imaging of phagocytic activity in abdominal aortic aneurysm

Objectives: : Imaging of biological processes associated with progression of abdominal aortic aneurysm (AAA), e.g., vessel wall inflammation, could lead to better AAA risk assessment and improve patient management. Phagocytosis is a key attribute of inflammatory cells, and imaging phagocytic activity, e.g., using ultrasmall superparamagnetic particles of iron oxide for magnetic resonance imaging (MRI), is a promising approach to assess inflammation. Compared to MRI, X-ray computed tomography (CT) has several advantages, including higher spatial resolution and lower cost, and is more often performed in combination with nuclear imaging to identify anatomic structures. Here, we sought to develop a CT-based approach for imaging vessel wall inflammation in preclinical models of AAA, which can be used alone or in conjunction with nuclear imaging. Methods: The potential of a nanoparticulated CT contrast agent (Np) to yield a measurable CT signal through phagocytosis was evaluated by incubation of Np with RAW 264.7 macrophage cells for 1 and 4 h prior to CT scanning of the cell pellet. Male apolipoprotein E-deficient mice (7-10-week-old, n = 22) were infused with angiotensin II (Ang II) to induce AAA. The surviving animals underwent CT scanning, performed immediately and 24 hours after intra-venous injection of Np suspension. CT images were acquired at 50 kV using a dedicated small animal CT camera (U-SPECT4/CT, MILabs). One group of animals (n = 8) was imaged at 4 and 5 days of Ang II infusion. A second group (n = 9) was imaged weekly during the 4 weeks of Ang II administration. The aorta was harvested after 4 weeks of Ang II infusion for ex vivo analysis in both groups. In a subgroup of animal euthanized at 4 weeks of Ang II infusion, the aneurysmal tissue was processed for transmission electron microscopy (TEM) and evaluated using a Lab6 emission, 80 kV microscope (G2 Spirit BioTWIN, FEI Tecnai).

Results: Np was taken up by RAW 264.7 cells, resulting in an increase of 21 and 64 Hounsfield units after 1 and 4 h of incubation, respectively, using a dose of Np equivalent to 20% of the initial blood concentration during in vivo imaging. 27% (6/22) of Ang II-infused mice died of aneurysm rupture within the first week. CT imaging performed right after Np injection showed contrast in the vascular pool. In 24% of animals, vascular contrast enabled the identification of a false lumen at the site of AAA, reflective of spontaneous aortic dissection. After 24 h, Np was cleared from the blood pool. In a subset of animals with AAA, including all animals with a false lumen, focally enhanced CT signal was observed in the aneurysm wall on delayed images. Accordingly, 5 out of 8 animals imaged at 4 days of Ang II infusion showed focal CT enhancement in the abdominal aortic wall. 7 out of these 8 animals had either AAA on ex vivo examination after 4 weeks of Ang II infusion or died of AAA rupture. The remaining animal without AAA did not show any Np uptake on early images. In the longitudinal study cohort, 56% (5/9) of animals showed Np uptake in the abdominal aorta at 1 week of Ang II infusion. Repeat Np-enhanced CT imaging during the follow up period detected additional areas of uptake in the aortic wall, reflecting newly developed areas of phagocytic activity. At 4 weeks, 89% (8/9) of animals showed Np uptake in the abdominal aorta and were confirmed to have AAA on ex vivo examination. TEM imaging showed phagosomal accumulation of X-ray-dense Np in inflammatory cells located in the aneurysmal aortic wall, primarily in the remodeled adventitia.

Conclusion: Vessel wall inflammation in AAA can be imaged using a novel Np-based CT imaging technique that targets phagocytosis. CT-detected aortic wall phagocytic signal is associated with AAA development and progression.