In Vivo Molecular K-Edge Imaging of Atherosclerotic Plaque Using Photon-counting CT

Abstract

Background Macrophage burden is a major factor in the risk of atherosclerotic plaque rupture, and its evaluation remains challenging with molecular noninvasive imaging approaches. Photon-counting CT (PCCT) with k-edge imaging aims to allow for the specific detection of macrophages using gold nanoparticles. Purpose To perform k-edge imaging in combination with gold nanoparticles to detect and quantify the macrophage burden within the atherosclerotic aortas of rabbits. Materials and Methods Atherosclerotic and control New Zealand white rabbits were imaged before and at several time points up to 2 days after intravenous injection of gold nanoparticles (3.5 mL/kg, 65 mg gold per milliliter). Aortic CT angiography was performed at the end of the follow-up using an intravenous injection of an iodinated contrast material. Gold k-edge and conventional CT images were reconstructed for qualitative and quantitative assessment of the macrophage burden. PCCT imaging results were compared with findings at histologic examination, quantitative histomorphometry, transmission electron microscopy, and quantitative inductively coupled plasma optical emission spectrometry. Pearson correlations between the macrophage area measured in immunostained sections and the concentration of gold and attenuation measured in the corresponding PCCT sections were calculated. Results Seven rabbits with atherosclerosis and four control rabbits without atherosclerosis were analyzed. In atherosclerotic rabbits, calcifications were observed along the aortic wall before injection. At 2 days after injection of gold nanoparticles, only gold k-edge images allowed for the distinction of plaque enhancement within calcifications and for lumen enhancement during angiography. A good correlation was observed between the gold concentration measured within the wall and the macrophage area in 35 plaques (five per rabbit) (r = 0.82; 95% CI: 0.67, 0.91; P < .001), which was higher than that observed on conventional CT images (r = 0.41; 95% CI: 0.09, 0.65; P = .01). Transmission electron microscopy and inductively coupled plasma optical emission spectrometry analyses confirmed the gold k-edge imaging findings. Conclusion Photon-counting CT with gold nanoparticles allowed for the noninvasive evaluation of both molecular and anatomic information in vivo in rabbits with atherosclerotic plaques. Published under a CC BY 4.0 license. Online supplemental material is available for this article. See also the editorial by Leiner in this issue.

Graphical abstract

Figure 1:

Study design schematic. AuNP = gold nanoparticles, ICP-OES = inductively coupled plasma optical emission spectrometry, inj. = injection, PCCT = photon-counting CT, TEM = transmission electron microscopy.

Figure 2:

In vitro characterization and imaging of gold nanoparticles (AuNP). A, Schematic representation of a gold nanoparticle. B, Transmission electron photomicrograph of gold nanoparticle. Note that gold nanoparticles are represented by darker spots. C, Characteristic absorption spectrum of gold at energy-dispersive x-ray spectroscopy. Au = gold, Cu = copper. D, In vitro photon-counting CT images of tubes containing gold nanoparticles (4 mg/mL), iomeprol (4 mg/mL), or calcium phosphate (1800 HU).

Figure 3:

Photon-counting CT images of atherosclerotic rabbit aortas before and 2 days after injection (inj.) of gold nanoparticles. A, Images show noncalcified plaque with strong circumferential enhancement and mean wall concentration of 4.5 mg/mL of gold nanoparticles. Parietal calcifications (arrowheads) are also shown. Green line in first image indicates aortic section analyzed. Rectangles in Overlay Before inj. and Overlay Day 2 images represent close-up view of parietal wall for better analyzing enhancement. B, Images show calcified plaque with strong enhancement within and around calcified area and mean wall concentration of 2.74 mg/mL of gold nanoparticles. Parietal calcifications (arrowheads) are also shown. Green line in first image indicates aortic section analyzed. Spotty areas of enhancement were more discernable on gold k-edge image than on conventional image. Rectangles in Overlay Before inj. and Overlay Day 2 images represent a close-up view of parietal wall for better analyzing enhancement. C, Box-and-whisker plot shows quantitative analysis of mean gold concentration found in whole atherosclerotic rabbit abdominal aortic wall before injection (mean concentration, 0.28 mg/mL ± 0.22; range, 0.01–0.66 mg/mL) and 2 days after injection (mean, 1.07 mg/mL ± 0.36; range, 0.45–1.56 mg/mL). Lower and upper margins of each box indicate 25th and 75th percentiles. Cross represents mean, and line in box represents median. Outliers indicate minimal and maximal values.

Figure 4:

Photon-counting CT images of atherosclerotic rabbit aorta before and 2 days after injection (inj.) of gold nanoparticles. A, Coronal 5-mm-width maximum intensity projection conventional image before injection shows presence of focal hyperattenuation along aortic wall indicative of calcifications. B–D, Coronal 5-mm-width maximum intensity projection aortic CT angiograms of atherosclerotic rabbit injected with vascular contrast material (iomerprol, 400 mg/mL) at 2 days (D2) after injection of gold nanoparticles. Lumens of aortic and renal arteries and enhancement of kidney and dense wall lesions are apparent on, B, maximum intensity projection conventional and, D, iodine images. C, Coronal 5-mm-width maximum intensity projection gold k-edge image shows extensive heterogeneous enhancement of wall. E, Fusion of three-dimensional (3D) volume rendering of conventional images with gold k-edge and iodine images shows spatial distribution of atherosclerotic macrophage burden within aortic wall. 2D = two-dimensional.

Figure 5:

Molecular k-edge imaging approach using photon-counting CT in vivo for macrophage detection and quantification in calcified atherosclerotic plaque in combination with k-edge contrast material of gold nanoparticles (AuNP). A, Schematic of uptake of gold nanoparticles by macrophages within atherosclerotic plaque (gold nanoparticles are shown as yellow dots, macrophages and foam cells are shown in purple, lipid core is shown in white, and calcification is shown in red). Gray nodules in foam cells are lipid droplets. White dashed lines indicate field of view of close-up in B and C. B, Schematic of conventional CT image shows different high-attenuation materials (gold nanoparticles and calcifications) that hamper characterization and quantification of macrophage burden. C, Schematic of PCCT gold k-edge image for specific, noninvasive macrophage burden imaging, which permits quantification of gold nanoparticles.

Figure 6:

A, B, Photon-counting CT (PCCT) images of atherosclerotic macrophage burden before injection (inj.) of gold nanoparticles and 2 days after injection and corresponding photomicrographs obtained with immunostaining. A, Images of noncalcified plaque show matches of spotty areas of enhancement next to mesenteric artery branch. This section is represented by yellow dot in C and black dot in D. B, Images of calcified plaque show low enhancement on gold k-edge image, which is confirmed by low concentration of immunostained macrophages. This section is represented by yellow square in C and black square in D. C, D, Graphs show correlation between area immunostained for macrophages and, C, concentration of gold or, D, attenuation on conventional images. Dotted and dashed lines represent the linear regression line and 95% CI.