Spatial lipidomics of coronary atherosclerotic plaque development in a familial hypercholesterolemia swine model.

Coronary atherosclerosis is caused by plaque build-up, with lipids playing a pivotal role in its progression. However, lipid composition and distribution within coronary atherosclerosis remain unknown. This study aims to characterize lipids and investigate differences in lipid composition across disease stages to aid in the understanding of disease progression. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) was used to visualize lipid distributions in coronary artery sections (n = 17) from hypercholesterolemic swine. We performed histology on consecutive sections to classify the artery segments and to investigate colocalization between lipids and histological regions of interest in advanced plaque, including necrotic core and inflammatory cells. Segments were classified as healthy (n = 6), mild (n = 6), and advanced disease (n = 5) artery segments. Multivariate data analysis was employed to find differences in lipid composition between the segment types, and the lipids' spatial distribution was investigated using non-negative matrix factorization (NMF). Through this process, MALDI-MSI detected 473 lipid-related features. NMF clustering described three components in positive ionization mode: triacylglycerides (TAG), phosphatidylcholines (PC), and cholesterol species. In negative ionization mode, two components were identified: one driven by phosphatidylinositol(PI)(38:4), and one driven by ceramide-phosphoethanolamine(36:1). Multivariate data analysis showed the association between advanced disease and specific lipid signatures like PC(O-40:5) and cholesterylester(CE)(18:2). Ether-linked phospholipids and LysoPC species were found to colocalize with necrotic core, and mostly CE, ceramide, and PI species colocalized with inflammatory cells. This study, therefore, uncovers distinct lipid signatures correlated with plaque development and their colocalization with necrotic core and inflammatory cells, enhancing our understanding of coronary atherosclerosis progression.

Mechanical wall stress and wall shear stress are associated with atherosclerosis development in non-calcified coronary segments.

BACKGROUND AND AIMS: Atherosclerotic plaque onset and progression are known to be affected by local biomechanical factors. While the role of wall shear stress (WSS) has been studied, the impact of another biomechanical factor, namely mechanical wall stress (MWS), remains poorly understood. In this study, we investigated the association of MWS, independently and combined with WSS, towards atherosclerosis in coronary arteries. METHODS: Thirty-four human coronary arteries were analyzed using near-infrared spectroscopy intravascular ultrasound (NIRS-IVUS) and optical coherence tomography (OCT) at baseline and after 12 months. Baseline WSS and MWS were calculated using computational models, and wall thickness (ΔWT) and lipid-rich necrotic core size (ΔLRNC) change were measured in non-calcified coronary segments. The arteries were further divided into 1.5 mm/45° sectors and categorized as plaque-free or plaque sectors. For each category, associations between biomechanical factors (WSS & MWS) and changes in coronary wall (ΔWT & ΔLRNC) were studied using linear mixed models. RESULTS: In plaque-free sectors, higher MWS (p < 0.001) was associated with greater vessel wall growth. Plaque sectors demonstrated wall thickness reduction over time, likely due to medical therapy, where higher levels of WSS and WMS, individually and combined, (p < 0.05) were associated with a greater reduction. Sectors with low MWS combined with high WSS demonstrated the highest LRNC increase (p < 0.01). CONCLUSIONS: In this study, we investigated the association of the (largely-overlooked) biomechanical factor MWS with coronary atherosclerosis, individually and combined with WSS. Our results demonstrated that both MWS and WSS significantly correlate with atherosclerotic plaque initiation and development.

Continuous shear wave measurements for dynamic cardiac stiffness evaluation in pigs.

Ultrasound-based shear wave elastography is a promising technique to non-invasively assess the dynamic stiffness variations of the heart. The technique is based on tracking the propagation of acoustically induced shear waves in the myocardium of which the propagation speed is linked to tissue stiffness. This measurement is repeated multiple times across the cardiac cycle to assess the natural variations in wave propagation speed. The interpretation of these measurements remains however complex, as factors such as loading and contractility affect wave propagation. We therefore applied transthoracic shear wave elastography in 13 pigs to investigate the dependencies of wave speed on pressure-volume derived indices of loading, myocardial stiffness, and contractility, while altering loading and inducing myocardial ischemia/reperfusion injury. Our results show that diastolic wave speed correlates to a pressure-volume derived index of operational myocardial stiffness (R = 0.75, p < 0.001), suggesting that both loading and intrinsic properties can affect diastolic wave speed. Additionally, the wave speed ratio, i.e. the ratio of systolic and diastolic speed, correlates to a pressure-volume derived index of contractility, i.e. preload-recruitable stroke work (R = 0.67, p < 0.001). Measuring wave speed ratio might thus provide a non-invasive index of contractility during ischemia/reperfusion injury.

Higher Order Singular Value Decomposition Filter for Contrast Echocardiography.

Assessing the coronary circulation with contrast-enhanced echocardiography has high clinical relevance. However, it is not being routinely performed in clinical practice because the current clinical tools generally cannot provide adequate image quality. The contrast agent's visibility in the myocardium is generally poor, impaired by motion and nonlinear propagation artifacts. The established multipulse contrast schemes (MPCSs) and the more experimental singular value decomposition (SVD) filter also fall short to solve these issues. Here, we propose a scheme to process amplitude modulation/amplitude-modulated pulse inversion (AM/AMPI) echoes with higher order SVD (HOSVD) instead of conventionally summing the complementary pulses. The echoes from the complementary pulses form a separate dimension in the HOSVD algorithm. Then, removing the ranks in that dimension with dominant coherent signals coming from tissue scattering would provide the contrast detection. We performed both in vitro and in vivo experiments to assess the performance of our proposed method in comparison with the current standard methods. A flow phantom study shows that HOSVD on AM pulsing exceeds the contrast-to-background ratio (CBR) of conventional AM and an SVD filter by 10 and 14 dB, respectively. In vivo porcine heart results also demonstrate that, compared to AM, HOSVD improves CBR in open-chest acquisition (up to 19 dB) and contrast ratio (CR) in closed-chest acquisition (3 dB).

High-Frame-Rate Volumetric Porcine Renal Vasculature Imaging.

OBJECTIVE: The aim of this study was to assess the feasibility and imaging options of contrast-enhanced volumetric ultrasound kidney vasculature imaging in a porcine model using a prototype sparse spiral array. METHODS: Transcutaneous freehand in vivo imaging of two healthy porcine kidneys was performed according to three protocols with different microbubble concentrations and transmission sequences. Combining high-frame-rate transmission sequences with our previously described spatial coherence beamformer, we determined the ability to produce detailed volumetric images of the vasculature. We also determined power, color and spectral Doppler, as well as super-resolved microvasculature in a volume. The results were compared against a clinical 2-D ultrasound machine. RESULTS: Three-dimensional visualization of the kidney vasculature structure and blood flow was possible with our method. Good structural agreement was found between the visualized vasculature structure and the 2-D reference. Microvasculature patterns in the kidney cortex were visible with super-resolution processing. Blood flow velocity estimations were within a physiological range and pattern, also in agreement with the 2-D reference results. CONCLUSION: Volumetric imaging of the kidney vasculature was possible using a prototype sparse spiral array. Reliable structural and temporal information could be extracted from these imaging results.

Local characterization of collagen architecture and mechanical failure properties of fibrous plaque tissue of atherosclerotic human carotid arteries.

Atherosclerotic plaque rupture in carotid arteries is a major cause of cerebrovascular events. Plaque rupture is the mechanical failure of the heterogeneous fibrous plaque tissue. Local characterization of the tissue's failure properties and the collagen architecture are of great importance to have insights in plaque rupture for clinical event prevention. Previous studies were limited to average rupture properties and global structural characterization, and did not provide the necessary local information. In this study, we assessed the local collagen architecture and failure properties of fibrous plaque tissue, by analyzing 30 tissue strips from 18 carotid plaques. Our study framework entailed second harmonic generation imaging for local collagen orientation and dispersion, and uniaxial tensile testing and digital image correlation for local tissue mechanics. The results showed that 87% of the imaged locations had collagen orientation close to the circumferential direction (0°) of the artery, and substantial dispersion locally. All regions combined, median [Q1:Q3] of the predominant angle measurements was -2° [-16°:16°]. The stretch ratio measurements clearly demonstrated a nonuniform stretch ratio distribution in the tissue under uniaxial loading. The rupture initiation regions had significantly higher stretch ratios (1.26 [1.15-1.40]) than the tissue average stretch ratio (1.11 [1.10-1.16]). No significant difference in collagen direction and dispersion was identified between the rupture regions and the rest of the tissue. The presented study forms an initial step towards gaining better insights into the characterization of local structural and mechanical fingerprints of fibrous plaque tissue in order to aid improved assessment of plaque rupture risk. STATEMENT OF SIGNIFICANCE: Plaque rupture risk assessment, critical to prevent cardiovascular events, requires knowledge on local failure properties and structure of collagenous plaque tissue. Our current knowledge is unfortunately limited to tissue's overall ultimate failure properties with scarce information on collagen architecture. In this study, local failure properties and collagen architecture of fibrous plaque tissue were obtained. We found predominant circumferential alignment of collagen fibers with substantial local dispersion. The tissue showed nonuniform stretch distribution under uniaxial tensile loading, with high stretches at rupture spots. This study highlights the significance of local mechanical and structural assessment for better insights into plaque rupture and the potential use of local stretches as risk marker for plaque rupture for patient-specific clinical applications.

Coupling Two Ultra-high-Speed Cameras to Elucidate Ultrasound Contrast-Mediated Imaging and Therapy.

Ultrasound contrast-mediated medical imaging and therapy both rely on the dynamics of micron- and nanometer-sized ultrasound cavitation nuclei, such as phospholipid-coated microbubbles and phase-change droplets. Ultrasound cavitation nuclei respond non-linearly to ultrasound on a nanosecond time scale that necessitates the use of ultra-high-speed imaging to fully visualize these dynamics in detail. In this study, we developed an ultra-high-speed optical imaging system that can record up to 20 million frames per second (Mfps) by coupling two small-sized, commercially available, 10-Mfps cameras. The timing and reliability of the interleaved cameras needed to achieve 20 Mfps was validated using two synchronized light-emitting diode strobe lights. Once verified, ultrasound-activated microbubble responses were recorded and analyzed. A unique characteristic of this coupled system is its ability to be reconfigured to provide orthogonal observations at 10 Mfps. Acoustic droplet vaporization was imaged from two orthogonal views, by which the 3-D dynamics of the phase transition could be visualized. This optical imaging system provides the temporal resolution and experimental flexibility needed to further elucidate the dynamics of ultrasound cavitation nuclei to potentiate the clinical translation of ultrasound-mediated imaging and therapy developments.

Intravascular optical coherence elastography.

Optical coherence elastography (OCE), a functional extension of optical coherence tomography (OCT), visualizes tissue strain to deduce the tissue's biomechanical properties. In this study, we demonstrate intravascular OCE using a 1.1 mm motorized catheter and a 1.6 MHz Fourier domain mode-locked OCT system. We induced an intraluminal pressure change by varying the infusion rate from the proximal end of the catheter. We analysed the pixel-matched phase change between two different frames to yield the radial strain. Imaging experiments were carried out in a phantom and in human coronary arteries in vitro. At an imaging speed of 3019 frames/s, we were able to capture the dynamic strain. Stiff inclusions in the phantom and calcification in atherosclerotic plaques are associated with low strain values and can be distinguished from the surrounding soft material, which exhibits elevated strain. For the first time, circumferential intravascular OCE images are provided side by side with conventional OCT images, simultaneously mapping both the tissue structure and stiffness.

Carotid Plaque Characteristics Predict Recurrent Ischemic Stroke and TIA: The PARISK (Plaque At RISK) Study.

BACKGROUND: Patients with symptomatic carotid stenosis are at high risk for recurrent stroke. The decision for carotid endarterectomy currently mainly relies on degree of stenosis (cutoff value >50% or 70%). Nevertheless, also, patients with mild-to-moderate stenosis still have a considerable recurrent stroke risk. Increasing evidence suggests that carotid plaque composition rather than degree of stenosis determines plaque vulnerability; however, it remains unclear whether this also provides additional information to improve clinical decision making. OBJECTIVES: The PARISK (Plaque At RISK) study aimed to improve the identification of patients at increased risk of recurrent ischemic stroke using multimodality carotid imaging. METHODS: The authors included 244 patients (71% men; mean age, 68 years) with a recent symptomatic mild-to-moderate carotid stenosis in a prospective multicenter cohort study. Magnetic resonance imaging (carotid and brain) and computed tomography angiography (carotid) were performed at baseline and after 2 years. The clinical endpoint was a recurrent ipsilateral ischemic stroke or transient ischemic attack (TIA). Cox proportional hazards models were used to assess whether intraplaque hemorrhage (IPH), ulceration, proportion of calcifications, and total plaque volume in ipsilateral carotid plaques were associated with the endpoint. Next, the authors investigated the predictive performance of these imaging biomarkers by adding these markers (separately and simultaneously) to the ECST (European Carotid Surgery Trial) risk score. RESULTS: During 5.1 years follow-up, 37 patients reached the clinical endpoint. IPH presence and total plaque volume were associated with recurrent ipsilateral ischemic stroke or TIA (HR: 2.12 [95% CI: 1.02-4.44] for IPH; HR: 1.07 [95% CI: 1.00-1.15] for total plaque volume per 100 µL increase). Ulcerations and proportion of calcifications were not statistically significant determinants. Addition of IPH and total plaque volume to the ECST risk score improved the model performance (C-statistics increased from 0.67 to 0.75-0.78). CONCLUSIONS: IPH and total plaque volume are independent risk factors for recurrent ipsilateral ischemic stroke or TIA in patients with mild-to-moderate carotid stenosis. These plaque characteristics improve current decision making. Validation studies to implement plaque characteristics in clinical scoring tools are needed. (PARISK: Validation of Imaging Techniques [PARISK]; NCT01208025).

Sparse 2-D PZT-on-PCB Arrays With Density Tapering.

Two-dimensional (2-D) arrays offer volumetric imaging capabilities without the need for probe translation or rotation. A sparse array with elements seeded in a tapering spiral pattern enables one-to-one connection to an ultrasound machine, thus allowing flexible transmission and reception strategies. To test the concept of sparse spiral array imaging, we have designed, realized, and characterized two prototype probes designed at 2.5-MHz low-frequency (LF) and 5-MHz high-frequency (HF) center frequencies. Both probes share the same electronic design, based on piezoelectric ceramics and rapid prototyping with printed circuit board substrates to wire the elements to external connectors. Different center frequencies were achieved by adjusting the piezoelectric layer thickness. The LF and HF prototype probes had 88% and 95% of working elements, producing peak pressures of 21 and 96 kPa/V when focused at 5 and 3 cm, respectively. The one-way -3-dB bandwidths were 26% and 32%. These results, together with experimental tests on tissue-mimicking phantoms, show that the probes are viable for volumetric imaging.

Erratum: Spectroscopic thermo-elastic optical coherence tomography for tissue characterization: publisher's note.

[This corrects the article on p. 1430 in vol. 13, PMID: 35414978.].

Dispersing and Sonoporating Biofilm-Associated Bacteria with Sonobactericide.

Bacteria encased in a biofilm poses significant challenges to successful treatment, since both the immune system and antibiotics are ineffective. Sonobactericide, which uses ultrasound and microbubbles, is a potential new strategy for increasing antimicrobial effectiveness or directly killing bacteria. Several studies suggest that sonobactericide can lead to bacterial dispersion or sonoporation (i.e., cell membrane permeabilization); however, real-time observations distinguishing individual bacteria during and directly after insonification are missing. Therefore, in this study, we investigated, in real-time and at high-resolution, the effects of ultrasound-induced microbubble oscillation on Staphylococcus aureus biofilms, without or with an antibiotic (oxacillin, 1 µg/mL). Biofilms were exposed to ultrasound (2 MHz, 100-400 kPa, 100-1000 cycles, every second for 30 s) during time-lapse confocal microscopy recordings of 10 min. Bacterial responses were quantified using post hoc image analysis with particle counting. Bacterial dispersion was observed as the dominant effect over sonoporation, resulting from oscillating microbubbles. Increasing pressure and cycles both led to significantly more dispersion, with the highest pressure leading to the most biofilm removal (up to 83.7%). Antibiotic presence led to more variable treatment responses, yet did not significantly impact the therapeutic efficacy of sonobactericide, suggesting synergism is not an immediate effect. These findings elucidate the direct effects induced by sonobactericide to best utilize its potential as a biofilm treatment strategy.

A concise history of echocardiography: timeline, pioneers, and landmark publications.

Echocardiography is less than 70 years old, and many major advances have occurred within living memory, but already some pioneering contributions may be overlooked. In order to consider what circumstances have been common to the most successful innovations, we have studied and here provide a timeline and summary of the most important developments in transthoracic and transoesophageal ultrasound imaging and Doppler techniques, as well as in intravascular ultrasound and imaging in paediatric cardiology. The entries are linked to a comprehensive list of first publications and to a collection of first-hand historical accounts published by early investigators. Review of the original manuscripts highlights that it is difficult to establish unequivocal precedence for many new imaging methods, since engineers were often working independently but simultaneously on similar problems. Many individuals who are prominently linked with particular developments were not the first in their field. Developments in echocardiography have been highly dependent on technological advances, and most likely to be successful when engineers and clinicians were able to collaborate with open exchange between centres and disciplines. As with many other new medical technologies, initial responses were sceptical and introduction into clinical practice required persistence and substantial energy from the first adopters. Current developments involve advances in software as much as in equipment, and progress will depend on continuing collaborations between engineers and clinical scientists, for example to identify unmet needs and to investigate the clinical impact of particular imaging approaches.

Internalization of targeted microbubbles by endothelial cells and drug delivery by pores and tunnels.

Ultrasound insonification of microbubbles can locally enhance drug delivery by increasing the cell membrane permeability. To aid development of a safe and effective therapeutic microbubble, more insight into the microbubble-cell interaction is needed. In this in vitro study we aimed to investigate the initial 3D morphology of the endothelial cell membrane adjacent to individual microbubbles (n = 301), determine whether this morphology was affected upon binding and by the type of ligand on the microbubble, and study its influence on microbubble oscillation and the drug delivery outcome. High-resolution 3D confocal microscopy revealed that targeted microbubbles were internalized by endothelial cells, while this was not the case for non-targeted or IgG1-κ control microbubbles. The extent of internalization was ligand-dependent, since αvß3-targeted microbubbles were significantly more internalized than CD31-targeted microbubbles. Ultra-high-speed imaging (~17 Mfps) in combination with high-resolution confocal microscopy (n = 246) showed that microbubble internalization resulted in a damped microbubble oscillation upon ultrasound insonification (2 MHz, 200 kPa peak negative pressure, 10 cycles). Despite damped oscillation, the cell's susceptibility to sonoporation (as indicated by PI uptake) was increased for internalized microbubbles. Monitoring cell membrane integrity (n = 230) showed the formation of either a pore, for intracellular delivery, or a tunnel (i.e. transcellular perforation), for transcellular delivery. Internalized microbubbles caused fewer transcellular perforations and smaller pore areas than non-internalized microbubbles. In conclusion, studying microbubble-mediated drug delivery using a state-of-the-art imaging system revealed receptor-mediated microbubble internalization and its effect on microbubble oscillation and resulting membrane perforation by pores and tunnels.

Independent Component Analysis Filter for Small Vessel Contrast Imaging During Fast Tissue Motion.

Suppressing tissue clutter is an essential step in blood flow estimation and visualization, even when using ultrasound contrast agents. Blind source separation (BSS)-based clutter filter for high-framerate ultrasound imaging has been reported to perform better in tissue clutter suppression than the conventional frequency-based wall filter and nonlinear contrast pulsing schemes. The most notable BSS technique, singular value decomposition (SVD) has shown compelling results in cases of slow tissue motion. However, its performance degrades when the tissue motion is faster than the blood flow speed, conditions that are likely to occur when imaging the small vessels, such as in the myocardium. Independent component analysis (ICA) is another BSS technique that has been implemented as a clutter filter in the spatiotemporal domain. Instead, we propose to implement ICA in the spatial domain where motion should have less impact. In this work, we propose a clutter filter with the combination of SVD and ICA to improve the contrast-to-background ratio (CBR) in cases where tissue velocity is significantly faster than the flow speed. In an in vitro study, the range of fast tissue motion velocity was 5-25 mm/s and the range of flow speed was 1-12 mm/s. Our results show that the combination of ICA and SVD yields 7-10 dB higher CBR than SVD alone, especially in the tissue high-velocity range. The improvement is crucial for cardiac imaging where relatively fast myocardial motions are expected.

Spectroscopic thermo-elastic optical coherence tomography for tissue characterization.

Optical imaging techniques that provide free space, label free imaging are powerful tools in obtaining structural and biochemical information in biological samples. To date, most of the optical imaging technologies create images with a specific contrast and require multimodality integration to add additional contrast. In this study, we demonstrate spectroscopic Thermo-elastic Optical Coherence Tomography (TE-OCT) as a potential tool in tissue identification. TE-OCT creates images based on two different forms of contrast: optical reflectance and thermo-elastic deformation. TE-OCT uses short laser pulses to induce thermo-elastic tissue deformation and measures the resulting surface displacement using phase-sensitive OCT. In this work we characterized the relation between thermo-elastic displacement and optical absorption, excitation, fluence and illumination area. The experimental results were validated with a 2-dimensional analytical model. Using spectroscopic TE-OCT, the thermo-elastic spectra of elastic phantoms and tissue components in coronary arteries were extracted. Specific tissue components, particularly lipid, an important biomarker for identifying atherosclerotic lesions, can be identified in the TE-OCT spectral response. As a label-free, free-space, dual-contrast, all-optical imaging technique, spectroscopic TE-OCT holds promise for biomedical research and clinical pathology diagnosis.

Theranostic Microbubbles with Homogeneous Ligand Distribution for Higher Binding Efficacy.

Phospholipid-coated targeted microbubbles are used for ultrasound molecular imaging and locally enhanced drug delivery, with the binding efficacy being an important trait. The use of organic solvent in microbubble production makes the difference between a heterogeneous or homogeneous ligand distribution. This study demonstrates the effect of ligand distribution on the binding efficacy of phospholipid-coated ανß3-targeted microbubbles in vitro using a monolayer of human umbilical-vein endothelial cells and in vivo using chicken embryos. Microbubbles with a homogeneous ligand distribution had a higher binding efficacy than those with a heterogeneous ligand distribution both in vitro and in vivo. In vitro, 1.55× more microbubbles with a homogeneous ligand distribution bound under static conditions, while this was 1.49× more under flow with 1.25 dyn/cm2, 1.56× more under flow with 2.22 dyn/cm2, and 1.25× more in vivo. The in vitro dissociation rate of bound microbubbles with homogeneous ligand distribution was lower at low shear stresses (1-5 dyn/cm2). The internalized depth of bound microbubbles was influenced by microbubble size, not by ligand distribution. In conclusion, for optimal binding the use of organic solvent in targeted microbubble production is preferable over directly dispersing phospholipids in aqueous medium.

Multicomponent material property characterization of atherosclerotic human carotid arteries through a Bayesian Optimization based inverse finite element approach.

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.

Vancomycin-decorated microbubbles as a theranostic agent for Staphylococcus aureus biofilms.

Bacterial biofilms are a huge burden on our healthcare systems worldwide. The lack of specificity in diagnostic and treatment possibilities result in difficult-to-treat and persistent infections. The aim of this in vitro study was to investigate if microbubbles targeted specifically to bacteria in biofilms could be used both for diagnosis as well for sonobactericide treatment and demonstrate their theranostic potential for biofilm infection management. The antibiotic vancomycin was chemically coupled to the lipid shell of microbubbles and validated using mass spectrometry and high-axial resolution 4Pi confocal microscopy. Theranostic proof-of-principle was investigated by demonstrating the specific binding of vancomycin-decorated microbubbles (vMB) to statically and flow grown Staphylococcus aureus (S. aureus) biofilms under increasing shear stress flow conditions (0-12 dyn/cm2), as well as confirmation of microbubble oscillation and biofilm disruption upon ultrasound exposure (2 MHz, 250 kPa, and 5,000 or 10,000 cycles) during flow shear stress of 5 dyn/cm2 using time-lapse confocal microscopy combined with the Brandaris 128 ultra-high-speed camera. Vancomycin was successfully incorporated into the microbubble lipid shell. vMB bound significantly more often than control microbubbles to biofilms, also in the presence of free vancomycin (up to 1000 µg/mL) and remained bound under increasing shear stress flow conditions (up to 12 dyn/cm2). Upon ultrasound insonification biofilm area was reduced of up to 28%, as confirmed by confocal microscopy. Our results confirm the successful production of vMB and support their potential as a new theranostic tool for S. aureus biofilm infections by allowing for specific bacterial detection and biofilm disruption.

Multicomponent Mechanical Characterization of Atherosclerotic Human Coronary Arteries: An Experimental and Computational Hybrid Approach.

Atherosclerotic plaque rupture in coronary arteries, an important trigger of myocardial infarction, is shown to correlate with high levels of pressure-induced mechanical stresses in plaques. Finite element (FE) analyses are commonly used for plaque stress assessment. However, the required information of heterogenous material properties of atherosclerotic coronaries remains to be scarce. In this work, we characterized the component-wise mechanical properties of atherosclerotic human coronary arteries. To achieve this, we performed ex vivo inflation tests on post-mortem human coronary arteries and developed an inverse FE modeling (iFEM) pipeline, which combined high-frequency ultrasound deformation measurements, a high-field magnetic resonance-based artery composition characterization, and a machine learning-based Bayesian optimization (BO) with uniqueness assessment. By using the developed pipeline, 10 cross-sections from five atherosclerotic human coronary arteries were analyzed, and the Yeoh material model constants of the fibrous intima and arterial wall components were determined. This work outlines the developed pipeline and provides the knowledge of non-linear, multicomponent mechanical properties of atherosclerotic human coronary arteries.