Background: Calcification and inflammation are atherosclerotic plaque compositional biomarkers that have both been linked to stroke risk. The aim of this study was to evaluate their co-existing prevalence in human carotid plaques with respect to plaque phenotype to determine the value of hybrid imaging for the detection of these biomarkers. Methods: Human carotid plaque segments, obtained from endarterectomy, were incubated in [111In]In-DOTA-butylamino-NorBIRT ([111In]In-Danbirt), targeting Leukocyte Function-associated Antigen-1 (LFA-1) on leukocytes. By performing SPECT/CT, both inflammation from DANBIRT uptake and calcification from CT imaging were assessed. Plaque phenotype was classified using histology. Results: On a total plaque level, comparable levels of calcification volume existed with different degrees of inflammation and vice versa. On a segment level, an inverse relationship between calcification volume and inflammation was evident in highly calcified segments, which classify as fibrocalcific, stable plaque segments. In contrast, segments with little or no calcification presented with a moderate to high degree of inflammation, often coinciding with the more dangerous fibrous cap atheroma phenotype. Conclusion: Calcification imaging alone can only accurately identify highly calcified, stable, fibrocalcific plaques. To identify high-risk plaques, with little or no calcification, hybrid imaging of calcification and inflammation could provide diagnostic benefit.

Purpose: Atherosclerotic plaque development and progression signifies a complex inflammatory disease mediated by a multitude of proinflammatory leukocyte subsets. Using single photon emission computed tomography (SPECT) coupled with computed tomography (CT), this study tested a new dual-isotope acquisition protocol to assess each radiotracer’s capability to identify plaque phenotype and inflammation levels pertaining to leukocytes expressing leukocyte function-associated antigen-1 (LFA-1) and the leukocyte subset of proinflammatory macrophages expressing somatostatin receptor subtype-2 (SST₂). Individual radiotracer uptake was quantified and the presence of corresponding immunohistological cell markers was assessed. Methods: Human symptomatic carotid plaque segments were obtained from endarterectomy. Segments were incubated in dual-isotope radiotracers [¹¹¹In]In-DOTA-butylamino-NorBIRT ([¹¹¹In]In-Danbirt) and [^99mTc]Tc-[N^0–1 ₄,Asp⁰,Tyr³]-octreotate ([^99mTc]Tc-Demotate 2) before scanning with SPECT/CT. Plaque phenotype was classified as pathological intimal thickening, fibrous cap atheroma or fibrocalcific using histology sections based on distinct morphological characteristics. Plaque segments were subsequently immuno-stained with LFA-1 and SST₂ and quantified in terms of positive area fraction and compared against the corresponding SPECT images. Results: Focal uptake of co-localising dual-radiotracers identified the heterogeneous distribution of inflamed regions in the plaques which co-localised with positive immuno-stained regions of LFA-1 and SST₂. [¹¹¹In]In-Danbirt and [^99mTc]Tc-Demotate 2 uptake demonstrated a significant positive correlation (r = 0.651; p = 0.001). Fibrous cap atheroma plaque phenotype correlated with the highest [¹¹¹In]In-Danbirt and [^99mTc]Tc-Demotate 2 uptake compared with fibrocalcific plaques and pathological intimal thickening phenotypes, in line with the immunohistological analyses. Conclusion: A dual-isotope acquisition protocol permits the imaging of multiple leukocyte subsets and the pro-inflammatory macrophages simultaneously in atherosclerotic plaque tissue. [¹¹¹In]In-Danbirt may have added value for assessing the total inflammation levels in atherosclerotic plaques in addition to classifying plaque phenotype.

Purpose: Intraplaque neovascularization (IPN) is an increasingly studied marker of the vulnerable atherosclerotic plaque, and contrast-enhanced ultrasound (CEUS) is an in vivo imaging technique for the assessment of IPN. The purpose of this study was to test novel quantification methods for the detection of carotid IPN using CEUS. Materials and Methods: 25 patients with established carotid atherosclerosis underwent bilateral carotid CEUS using a Philips iU-22 ultrasound system with an L9-3 transducer. Visual scoring of IPN was performed using a 3-point score. Quantification of IPN was performed using novel custom developed software. In short, regions of interest were drawn over the atherosclerotic plaques. After motion compensation, several IPN features were calculated. Statistical analysis was performed using Spearman's rho. Reproducibility of the quantification features was calculated using intra-class correlation coefficients and mean differences between calculations. Results: 45 carotid arteries were available for the quantification of IPN. The quantification of IPN was feasible in all 45 carotid plaques. The IPN area, IPN area ratio and neovessel count had a good correlation with the visual IPN score (respectively ρ=0.719, ρ=0.538, ρ=0.474 all p<0.01). The intra-observer and inter-observer agreement was good to excellent (p<0.01). The intra-observer and inter-observer variability was low. Conclusion: The quantification of carotid IPN on CEUS is feasible and provides multiple features on carotid IPN. Accurate quantitative assessment of IPN may be important to recognize and to monitor changes during therapy in vulnerable atherosclerotic plaques.