Search

Thrombus computed tomography (CT) imaging characteristics may correspond with thrombus mechanical properties and thus predict thrombectomy success. The impact of red blood cell (RBC) content on these properties (imaging and mechanics) has been widely studied. However, the additional effect of platelets has not been considered. The objective of the current study was to examine the individual and combined effects of blood clot RBC and platelet content on resultant CT imaging and mechanical characteristics. Human blood clot analogues were prepared from a combination of preselected RBC volumes and platelet concentrations to decouple their contributions. The resulting clot RBC content (%) and platelet content (%) were determined using Martius Scarlet Blue and CD42b staining, respectively. Non-contrast and contrast-enhanced CT (NCCT and CECT) scans were performed to measure the clot densities. CECT density increase was taken as a proxy for clinical perviousness. Unconfined compressive mechanics were analysed by performing 10 cycles of 80% strain. RBC content is the major determinant of clot NCCT density. However, additional consideration of the platelet content improves the association. CECT density increase is influenced by clot platelet and not RBC content. Platelet content is the dominant component driving clot stiffness, especially at high strains. Both RBC and platelet content contribute to the clot's viscoelastic and plastic compressive properties. The current in vitro results suggest that CT density is reflective of RBC content and subsequent clot viscoelasticity and plasticity, and that perviousness reflects the clot's platelet content and subsequent stiffness. However, these indications should be confirmed in a clinical stroke cohort.

The rupture of an atherosclerotic plaque cap overlying a lipid pool and/or necrotic core can lead to thrombotic cardiovascular events. In essence, the rupture of the plaque cap is a mechanical event, which occurs when the local stress exceeds the local tissue strength. However, due to inter- and intra-cap heterogeneity, the resulting ultimate cap strength varies, causing proper assessment of the plaque at risk of rupture to be lacking. Important players involved in tissue strength include the load-bearing collagenous matrix, macrophages, as major promoters of extracellular matrix degradation, and microcalcifications, deposits that can exacerbate local stress, increasing tissue propensity for rupture. This review summarizes the role of these components individually in tissue mechanics, along with the interplay between them. We argue that to be able to improve risk assessment, a better understanding of the effect of these individual components, as well as their reciprocal relationships on cap mechanics, is required. Finally, we discuss potential future steps, including a holistic multidisciplinary approach, multifactorial 3D in vitro model systems, and advancements in imaging techniques. The obtained knowledge will ultimately serve as input to help diagnose, prevent, and treat atherosclerotic cap rupture.

Background Clot composition, contraction, and mechanical properties are likely determinants of endovascular thrombectomy success. A pre-interventional estimation of these properties is hypothesized to aid in selecting the most suitable treatment for different types of thrombi. Here we determined the association between the aforementioned properties and computed tomography (CT) characteristics using human blood clot analogues. Methods Clot analogues were prepared from the blood of 4 healthy human donors with 5 red blood cell (RBC) volume suspensions: 0%, 20%, 40%, 60% and 80% RBCs. Contraction was measured as the weight of the contracted clots as a percentage of the original suspension. The clots were imaged using CT with and without contrast to quantify clot density and density increase. Unconfined compression was performed to determine the high strain compressive stiffness. The RBC content was analysed using H&E staining. Results The 5 RBC suspensions formed only two groups of clots, fibrin-rich (0% RBCs) and RBC-rich (>90% RBCs), as determined by histology. The density of the fibrin-rich clots was significantly lower (31-38HU) compared to the RBC-rich clots (72-89HU), and the density increase of the fibrin-rich clots was significantly higher (82-127HU) compared to the RBC-rich clots (3-17HU). The compressive stiffness of the fibrin-rich clots was higher (178–1624 kPa) than the stiffness of the RBC-rich clots (6–526 kPa). Additionally, the degree of clot contraction was higher for the fibrin-rich clots (89–96%) compared to the RBC-rich clots (11–77%). Conclusions CT imaging clearly reflects clot RBC content and seems to be related to the clot contraction and stiffness. CT imaging might be a useful tool in predicting the thrombus characteristics. However, future studies should confirm these findings by analysing clots with intermediate RBC and platelet content.

Background The purpose of this study was to validate a technique for measuring mean calcium density and to determine associations of cardiovascular risk factors with carotid calcium density. Methods and Results We performed a cross-sectional study in a random sample of 100 stroke-free participants from the population-based Rotterdam Study. The mean calcium density of the combined left and right carotid bifurcations was quantified with a threshold of 130 Hounsfield Units (HU) using a novel density technique. To validate the methodology, carotid calcium volumes acquired using the technique in the current study were compared with measurements computed using dedicated clinical software (semiautomatic technique based on a threshold of ≥130 HU). Next, we investigated the associations of participant demographics, total calcium volume, and known cardiovascular risk factors (hypertension, diabetes, hypercholesterolemia, obesity, and smoking status) with the newly derived mean carotid calcium density measurement using linear regression analyses. Calcium volumes obtained with the 2 methods showed a high agreement (intraclass correlation coefficient=0.99, P<0.001), underlining the validity of the density technique. The total calcium volume was statistically significantly associated with the mean calcium density (cardiovascular risk factors adjusted model (B: 0.48 [95% CI, 0.30-0.66], P<0.001). We also found an association between hypercholesterolemia and mean calcium density (0.46 [0.09-0.83], P=0.017). No other significant associations were found between participant demographics or cardiovascular risk factors and mean carotid calcium density. Conclusions We demonstrated the feasibility of a carotid calcium density measurement technique. The data warrant a subsequent longitudinal study to determine the association between carotid calcium density and the risk of cerebrovascular events.

Rupture of the cap of an atherosclerotic plaque can lead to thrombotic cardiovascular events. It has been suggested, through computational models, that the presence of microcalcifications in the atherosclerotic cap can increase the risk of cap rupture. However, the experimental confirmation of this hypothesis is still lacking. In this study, we have developed a novel tissue-engineered model to mimic the atherosclerotic fibrous cap with microcalcifications and assess the impact of microcalcifications on cap mechanics. First, human carotid plaque caps were analyzed to determine the distribution, size, and density of microcalcifications in real cap tissue. Hydroxyapatite particles with features similar to real cap microcalcifications were used as microcalcification mimics. Injected clusters of hydroxyapatite particles were embedded in a fibrin gel seeded with human myofibroblasts which deposited a native-like collagenous matrix around the particles, during the 21-day culture period. Second harmonic multiphoton microscopy imaging revealed higher local collagen fiber dispersion in regions of hydroxyapatite clusters. Tissue-engineered caps with hydroxyapatite particles demonstrated lower stiffness and ultimate tensile stress than the control group samples under uniaxial tensile loading, suggesting increased rupture risk in atherosclerotic plaques with microcalcifications. This model supports previous computational findings regarding a detrimental role for microcalcifications in cap rupture risk and can further be deployed to elucidate tissue mechanics in pathologies with calcifying soft tissues.

Rupture of the cap of an atherosclerotic plaque can lead to thrombotic cardiovascular events. It has been suggested, through computational models, that the presence of microcalcifications in the atherosclerotic cap can increase the risk of cap rupture. However, the experimental confirmation of this hypothesis is still lacking. In this study, we have developed a novel tissue-engineered model to mimic the atherosclerotic fibrous cap with microcalcifications and assess the impact of microcalcifications on cap mechanics. First, human carotid plaque caps were analyzed to determine the distribution, size, and density of microcalcifications in real cap tissue. Hydroxyapatite particles with features similar to real cap microcalcifications were used as microcalcification mimics. Injected clusters of hydroxyapatite particles were embedded in a fibrin gel seeded with human myofibroblasts which deposited a native-like collagenous matrix around the particles, during the 21-day culture period. Second harmonic multiphoton microscopy imaging revealed higher local collagen fiber dispersion in regions of hydroxyapatite clusters. Tissue-engineered caps with hydroxyapatite particles demonstrated lower stiffness and ultimate tensile stress than the control group samples under uniaxial tensile loading, suggesting increased rupture risk in atherosclerotic plaques with microcalcifications. This model supports previous computational findings regarding a detrimental role for microcalcifications in cap rupture risk and can further be deployed to elucidate tissue mechanics in pathologies with calcifying soft tissues.

Endovascular thrombectomy procedures are significantly influenced by the mechanical response of thrombi to the multi-axial loading imposed during retrieval. Compression tests are commonly used to determine compressive ex vivo thrombus and clot analogue stiffness. However, there is a shortage of data in tension. This study compares the tensile and compressive response of clot analogues made from the blood of healthy human donors in a range of compositions. Citrated whole blood was collected from six healthy human donors. Contracted and non-contracted fibrin clots, whole blood clots and clots reconstructed with a range of red blood cell (RBC) volumetric concentrations (5–80%) were prepared under static conditions. Both uniaxial tension and unconfined compression tests were performed using custom-built setups. Approximately linear nominal stress–strain profiles were found under tension, while strong strain-stiffening profiles were observed under compression. Low- and high-strain stiffness values were acquired by applying a linear fit to the initial and final 10% of the nominal stress–strain curves. Tensile stiffness values were approximately 15 times higher than low-strain compressive stiffness and 40 times lower than high-strain compressive stiffness values. Tensile stiffness decreased with an increasing RBC volume in the blood mixture. In contrast, high-strain compressive stiffness values increased from 0 to 10%, followed by a decrease from 20 to 80% RBC volumes. Furthermore, inter-donor differences were observed with up to 50% variation in the stiffness of whole blood clot analogues prepared in the same manner between healthy human donors.

AIMS: Low wall shear stress (WSS) is acknowledged to play a role in plaque development through its influence on local endothelial function. Also, lipid-rich plaques (LRPs) are associated with endothelial dysfunction. However, little is known about the interplay between WSS and the presence of lipids with respect to plaque progression. Therefore, we aimed to study the differences in WSS-related plaque progression between LRPs, non-LRPs, or plaque-free regions in human coronary arteries.METHODS AND RESULTS: In the present single-centre, prospective study, 40 patients who presented with an acute coronary syndrome successfully underwent near-infrared spectroscopy intravascular ultrasound (NIRS-IVUS) and optical coherence tomography (OCT) of at least one non-culprit vessel at baseline and completed a 1-year follow-up. WSS was computed applying computational fluid dynamics to a three-dimensional reconstruction of the coronary artery based on the fusion of the IVUS-segmented lumen with a CT-derived centreline, using invasive flow measurements as boundary conditions. For data analysis, each artery was divided into 1.5 mm/45° sectors. Plaque growth based on IVUS-derived percentage atheroma volume change was compared between LRPs, non-LRPs, and plaque-free wall segments, as assessed by both OCT and NIRS. Both NIRS- and OCT-detected lipid-rich sectors showed a significantly higher plaque progression than non-LRPs or plaque-free regions. Exposure to low WSS was associated with a higher plaque progression than exposure to mid or high WSS, even in the regions classified as a plaque-free wall. Furthermore, low WSS and the presence of lipids had a synergistic effect on plaque growth, resulting in the highest plaque progression in lipid-rich regions exposed to low shear stress.CONCLUSION: This study demonstrates that NIRS- and OCT-detected lipid-rich regions exposed to low WSS are subject to enhanced plaque growth over a 1-year follow-up. The presence of lipids and

The rupture of atherosclerotic plaques in coronary and carotid arteries is the primary cause of fatal cardiovascular events. However, the rupture mechanics of the heterogeneous, highly collagenous plaque tissue, and how this is related to the tissue's fibrous structure, are not known yet. Existing pipelines to study plaque mechanics are limited to obtaining only gross mechanical characteristics of the plaque tissue, based on the assumption of structural homogeneity of the tissue. However, fibrous plaque tissue is structurally heterogeneous, arguably mainly due to local variation in the collagen fiber architecture. The mechano-imaging pipeline described here has been developed to study the heterogeneous structural and mechanical plaque properties. In this pipeline, the tissue's local collagen architecture is characterized using multiphoton microscopy (MPM) with second-harmonic generation (SHG), and the tissue's failure behavior is characterized under uniaxial tensile testing conditions using digital image correlation (DIC) analysis. This experimental pipeline enables correlation of the local predominant angle and dispersion of collagen fiber orientation, the rupture behavior, and the strain fingerprints of the fibrous plaque tissue. The obtained knowledge is key to better understand, predict, and prevent atherosclerotic plaque rupture events.

Objective: Plaque rupture in atherosclerotic carotid arteries is a main cause of ischemic stroke and it is correlated with high plaque stresses. Hence, analyzing stress patterns is essential for plaque specific rupture risk assessment. However, the critical information of the multicomponent material properties of atherosclerotic carotid arteries is still lacking greatly. This work aims to characterize component-wise material properties of atherosclerotic human carotid arteries under (almost) physiological loading conditions. Methods: An inverse finite element modeling (iFEM) framework was developed to characterize fibrous intima and vessel wall material properties of 13 cross sections from five carotids. The novel pipeline comprised ex-vivo inflation testing, pre-clinical high frequency ultrasound for deriving plaque deformations, pre-clinical high-magnetic field magnetic resonance imaging, finite element modeling, and a sample efficient machine learning based Bayesian Optimization. Results: The nonlinear Yeoh constants for the fibrous intima and wall layers were successfully obtained. The optimization scheme of the iFEM reached the global minimum with a mean error of 3.8% in 133 iterations on average. The uniqueness of the results were confirmed with the inverted Gaussian Process (GP) model trained during the iFEM protocol. Conclusion: The developed iFEM approach combined with the inverted GP model successfully predicted component-wise material properties of intact atherosclerotic human carotids ex-vivo under physiological-like loading conditions. Significance: We developed a novel iFEM framework for the nonlinear, component-wise material characterization of atherosclerotic arteries and utilized it to obtain human atherosclerotic carotid material properties. The developed iFEM framework has great potential to be advanced for patient-specific in-vivo application.

Thrombus composition and mechanical properties significantly impact the ease and outcomes of thrombectomy procedures in patients with acute ischemic stroke. A wide variation exists in the composition of thrombi between patients. If a relationship can be determined between the composition of a thrombus and its mechanical behaviour, as well as between the composition of a thrombus and its radiological imaging characteristics, then there is the potential to personalise thrombectomy treatment based on each individual thrombus. This review aims to give an overview of the current literature addressing this issue. Here, we present a scoping review detailing associations between thrombus composition, mechanical behaviour and radiological imaging characteristics. We conducted two searches 1) on the association between thrombus composition and the mechanical behaviour of the tissue and 2) on the association between radiological imaging characteristics and thrombus composition in the acute stroke setting. The review suggests that higher fibrin and lower red blood cell (RBC) content contribute to stiffer thrombi independent of the loading mode. Further, platelet-contracted thrombi are stiffer than non-contracted compositional counterparts. Fibrin content contributes to the elastic portion of viscoelastic behaviour while RBC content contributes to the viscous portion. It is possible to identify fibrin-rich or RBC-rich thrombi with computed tomography and magnetic resonance imaging vessel signs. Standardisation is required to quantify the association between thrombus density on non-contrast computed tomography and the RBC content. The characterisation of the thrombus fibrin network has not been addressed so far in radiological imaging but may be essential for the prediction of device-tissue interactions and distal thrombus embolization. The association between platelet-driven clot contraction and radiological imaging characteristics has not been explicitly investigated. Howeve

Intravascular ultrasound (IVUS) imaging offers accurate cross-sectional vessel information. To this end, registering temporal IVUS pullbacks acquired at two time points can assist the clinicians to accurately assess pathophysiological changes in the vessels, disease progression and the effect of the treatment intervention. In this paper, we present a novel two-stage registration framework for aligning pairs of longitudinal and axial IVUS pullbacks. Initially, we use a Dynamic Time Warping (DTW)-based algorithm to align the pullbacks in a temporal fashion. Subsequently, an intensity-based registration method, that utilizes a variant of the Harmony Search optimizer to register each matched pair of the pullbacks by maximizing their Mutual Information, is applied. The presented method is fully automated and only required two single global image-based measurements, unlike other methods that require extraction of morphology-based features. The data used includes 42 synthetically generated pullback pairs, achieving an alignment error of 0.1853 frames per pullback, a rotation error 0.93° and a translation error of 0.0161 mm. In addition, it was also tested on 11 baseline and follow-up, and 10 baseline and post-stent deployment real IVUS pullback pairs from two clinical centres, achieving an alignment error of 4.3±3.9 for the longitudinal registration, and a distance and a rotational error of 0.56±0.323 mm and 12.4°±10.5°, respectively, for the axial registration. Although the performance of the proposed method does not match that of the state-of-the-art, our method relies on computationally lighter steps for its computations, which is crucial in real-time applications. On the other hand, the proposed method performs even or better that the state-of-the-art when considering the axial registration. The results indicate that the proposed method can support clinical decision making and diagnosis based on sequential imaging examinations.

Objective: Atherosclerotic plaque rupture in carotid arteries is a major source of cerebrovascular events. Calcifications are highly prevalent in carotid plaques, but their role in plaque rupture remains poorly understood. This work studied the morphometric features of calcifications in carotid plaques and their effect on the stress distribution in the fibrous plaque tissue at the calcification interface, as a potential source of plaque rupture and clinical events. Methods: A comprehensive morphometric analysis of 65 histology cross-sections from 16 carotid plaques was performed to identify the morphology (size and shape) and location of plaque calcifications, and the fibrous tissue fiber organization around them. Calcification-specific finite element models were constructed to examine the fibrous plaque tissue stresses at the calcification interface. Statistical correlation analysis was performed to elucidate the impact of calcification morphology and fibrous tissue organization on interface stresses. Results: Hundred-seventy-one calcifications were identified on the histology cross-sections, which showed great variation in morphology. Four distinct patterns of fiber organization in the plaque tissue were observed around the calcification. They were termed as attached, pushed-aside, encircling and random patterns. The stress analyses showed that calcifications are correlated with high interface stresses, which might be comparable to or even above the plaque strength. The stress levels depended on the calcification morphology and fiber organization. Thicker calcification with a circumferential slender shape, located close to the lumen were correlated most prominently to high interface stresses. Conclusion: Depending on its morphology and the fiber organization around it, a calcification in an atherosclerotic plaque can act as a stress riser and cause high interface stresses. Significance: This study demonstrated the potential of calcifications in atherosclerotic

Treatment of acute ischemic stroke has been recently improved with the introduction of endovascular mechanical thrombectomy, a minimally invasive procedure able to remove a clot using aspiration devices and/or stent-retrievers. Despite the promising and encouraging results, improvements to the procedure and to the stent design are the focus of the recent efforts. Computational studies can pave the road to these improvements, providing their ability to describe and accurately reproduce a real procedure. A patient with ischemic stroke due to intracranial large vessel occlusion was selected and after the creation of the cerebral vasculature from computed tomography images and a histologic analysis to determine the clot composition, the entire thrombectomy procedure was virtually replicated. As in the real situation, the computational replica showed that two attempts were necessary to remove the clot, as a result of the position of the stent retriever with respect to the clot. Furthermore, the results indicated that clot fragmentation did not occur as the deformations were mainly in a compressive state without the possibility for clot cracks to propagate. The accurate representation of the procedure can be used as an important step for operative optimization planning and future improvements of stent designs.

Carotid atherosclerosis is a risk factor for ischemic stroke, one of the main causes of mortality and disability worldwide. The disease is characterized by plaques, heterogeneous deposits of lipids, and necrotic debris in the vascular wall, which grow gradually and may remain asymptomatic for decades. However, at some point a plaque can evolve to a highrisk plaque phenotype, which may trigger a cerebrovascular event. Lipids play a key role in the development and progression of atherosclerosis, but the nature of their involvement is not fully understood. Using matrix-assisted laser desorption/ionization mass spectrometry imaging, we visualized the distribution of approximately 200 different lipid signals, originating of >90 uniquely assigned species, in 106 tissue sections of 12 human carotid atherosclerotic plaques. We performed unsupervised classification of the mass spectrometry dataset, as well as a histology-directed multivariate analysis. These data allowed us to extract the spatial lipid patterns associated withmorphological plaque features in advanced plaques from a symptomatic population, revealing spatial lipid patterns in atherosclerosis and their relation to histological tissue type. The abundances of sphingomyelin and oxidized cholesteryl ester species were elevated specifically in necrotic intima areas, whereas diacylglycerols and triacylglycerols were spatially correlated to areas containing the coagulation protein fibrin. These results demonstrate a clear colocalization between plaque features and specific lipid classes, as well as individual lipid species in high-risk atherosclerotic plaques.

Abstract

High wall shear stress (WSS) and near-infrared spectroscopy (NIRS) detected lipid-rich plaque (LRP) are both known to be associated with plaque destabilization and future adverse cardiovascular events. However, knowledge of spatial co-localization of LRP and high WSS is lacking. This study investigated the co-localization of LRP based on NIRS and high WSS. Fifty-three patients presenting acute coronary syndrome underwent NIRS-intravascular-ultrasound (NIRS-IVUS) imaging of a non-culprit coronary artery. WSS was obtained using WSS profiling in 3D-reconstructions of the coronary arteries based on fusion of IVUS-segmented lumen and CT-derived 3D-centerline. Thirty-eight vessels were available for final analysis and divided into 0.5 mm/45° sectors. LRP sectors, as identified by NIRS, were more often colocalized with high WSS than sectors without LRP. Moreover, there was a dose-dependent relationship between lipid content and high WSS exposure. This study is a first step in understanding the evolution of LRPs to vulnerable plaques. [Figure not available: see fulltext.]

Wall shear stress (WSS), the frictional force of the blood on the vessel wall, plays a crucial role in atherosclerotic plaque development. Low WSS has been associated with plaque growth, however previous research used different approaches to define low WSS to investigate its effect on plaque progression. In this study, we used four methodologies to allocate low, mid and high WSS in one dataset of human coronary arteries and investigated the predictive power of low WSS for plaque progression. Coronary reconstructions were based on multimodality imaging, using intravascular ultrasound and CT-imaging. Vessel-specific flow was measured using Doppler wire and computational fluid dynamics was performed to calculate WSS. The absolute WSS range varied greatly between the coronary arteries. On the population level, the established pattern of most plaque progression at low WSS was apparent in all methodologies defining the WSS categories. However, for the individual patient, when using measured flow to determine WSS, the absolute WSS values range so widely, that the use of absolute thresholds to determine low WSS was not appropriate to identify regions at high risk for plaque progression.

Atherosclerotic plaques prone to rupture may cause acute myocardial infarction (MI) but can also heal without causing an event. Certain common histopathological features, including inflammation, a thin fibrous cap, positive remodelling, a large necrotic core, microcalcification, and plaque haemorrhage are commonly found in plaques causing an acute event. Recent advances in imaging techniques have made it possible to detect not only luminal stenosis and overall coronary atherosclerosis burden but also to identify such adverse plaque characteristics. However, the predictive value of identifying individual adverse atherosclerotic plaques for future events has remained poor. In this Position Paper, the relationship between vulnerable plaque imaging and MI is addressed, mainly for non-invasive assessments but also for invasive imaging of adverse plaques in patients undergoing invasive coronary angiography. Dynamic changes in atherosclerotic plaque development and composition may indicate that an adverse plaque phenotype should be considered at the patient level rather than for individual plaques. Imaging of adverse plaque burden throughout the coronary vascular tree, in combination with biomarkers and biomechanical parameters, therefore holds promise for identifying subjects at increased risk of MI and for guiding medical and invasive treatment.

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-[N0–14,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 fib