Pre-transplant kidney quality evaluation using photoacoustic imaging during normothermic machine perfusion.

Due to the shortage of kidneys donated for transplantation, surgeons are forced to use the organs with an elevated risk of poor function or even failure. Although the existing methods for pre-transplant quality evaluation have been validated over decades in population cohort studies across the world, new methods are needed as long as delayed graft function or failure in a kidney transplant occurs. In this study, we explored the potential of utilizing photoacoustic (PA) imaging during normothermic machine perfusion (NMP) as a means of evaluating kidney quality. We closely monitored twenty-two porcine kidneys using 3D PA imaging during a two-hour NMP session. Based on biochemical analyses of perfusate and produced urine, the kidneys were categorized into 'non-functional' and 'functional' groups. Our primary focus was to quantify oxygenation (sO2) within the kidney cortical layer of depths 2 mm, 4 mm, and 6 mm using two-wavelength PA imaging. Next, receiver operating characteristic (ROC) analysis was performed to determine an optimal cortical layer depth and time point for the quantification of sO2 to discriminate between functional and non-functional organs. Finally, for each depth, we assessed the correlation between sO2 and creatinine clearance (CrCl), oxygen consumption (VO2), and renal blood flow (RBF). We found that hypoxia of the renal cortex is associated with poor renal function. In addition, the determination of sO2 within the 2 mm depth of the renal cortex after 30 min of NMP effectively distinguishes between functional and non-functional kidneys. The non-functional kidneys can be detected with the sensitivity and specificity of 80% and 85% respectively, using the cut-off point of sO2 < 39%. Oxygenation significantly correlates with RBF and VO2 in all kidneys. In functional kidneys, sO2 correlated with CrCl, which is not the case for non-functional kidneys. We conclude that the presented technique has a high potential for supporting organ selection for kidney transplantation.

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.

Identifying lipid traces of atherogenic mechanisms in human carotid plaque.

BACKGROUND AND AIMS: Lipids play an important role in atherosclerotic plaque development and are interesting candidate predictive biomarkers. However, the link between circulating lipids, accumulating lipids in the vessel wall, and plaque destabilization processes in humans remains largely unknown. This study aims to provide new insights into the role of lipids in atherosclerosis using lipidomics and mass spectrometry imaging to investigate lipid signatures in advanced human carotid plaque and plasma samples. METHODS: We used lipidomics and desorption electrospray ionization mass spectrometry imaging (DESI-MSI) to investigate lipid signatures of advanced human carotid plaque and plasma obtained from patients who underwent carotid endarterectomy (n = 14 out of 17 whose plaque samples were analyzed by DESI-MSI). Multivariate data analysis and unsupervised clustering were applied to identify lipids that were the most discriminative species between different patterns in plaque and plasma. These patterns were interpreted by quantitative comparison with conventional histology. RESULTS: Lipidomics detected more than 300 lipid species in plasma and plaque, with markedly different relative abundances. DESI-MSI visualized the spatial distribution of 611 lipid-related m/z features in plaques, of which 330 m/z features could be assigned based on exact mass, comparison to the lipidomic data, and high mass resolution MSI. Matching spatial lipid patterns to histological areas of interest revealed several molecular species that were colocalized with pertinent disease processes in plaque including specific sphingomyelin and ceramide species with calcification, phospholipids and free fatty acids with inflammation, and triacylglycerols and phosphatidylinositols with fibrin-rich areas. CONCLUSIONS: By comparing lipid species in plaque and plasma, we identified those circulating species that were also prominently present in plaque. Quantitative comparison of lipid spectral patterns with histology revealed the presence of specific lipid species in destabilized plaque areas, corroborating previous in vitro and animal studies.

Spectroscopic optical coherence tomography at 1200 nm for lipid detection.

Significance: Spectroscopic analysis of optical coherence tomography (OCT) data can yield added information about the sample's chemical composition, along with high-resolution images. Typical commercial OCT systems operate at wavelengths that may not be optimal for identifying lipid-containing samples based on absorption features. Aim: The main aim of this study was to develop a 1200 nm spectroscopic OCT (SOCT) for the classification of lipid-based and water-based samples by extracting the lipid absorption peak at 1210 nm from the OCT data. Approach: We developed a 1200 nm OCT system and implemented a signal processing algorithm that simultaneously retrieves spectroscopic and structural information from the sample. In this study, we validated the performance of our OCT system by imaging weakly scattering phantoms with and without lipid absorption features. An orthogonal projections to latent structures-discriminant analysis (OPLS-DA) model was developed and applied to classify weakly scattering samples based on their absorption features. Results: The OCT system achieved an axial resolution of 7.2 µm and a sensitivity of 95 dB. The calibrated OPLS-DA model on weakly scattering samples with lipid and water-based absorption features correctly classified 19/20 validation samples. Conclusions: The 1200 nm SOCT system can discriminate the lipid-containing weakly scattering samples from water-based weakly scattering samples with good predictive ability.

Plaque burden is associated with minimal intimal coverage following drug-eluting stent implantation in an adult familial hypercholesterolemia swine model.

Safety and efficacy of coronary drug-eluting stents (DES) are often preclinically tested using healthy or minimally diseased swine. These generally show significant fibrotic neointima at follow-up, while in patients, incomplete healing is often observed. The aim of this study was to investigate neointima responses to DES in swine with significant coronary atherosclerosis. Adult familial hypercholesterolemic swine (n = 6) received a high fat diet to develop atherosclerosis. Serial OCT was performed before, directly after, and 28 days after DES implantation (n = 14 stents). Lumen, stent and plaque area, uncovered struts, neointima thickness and neointima type were analyzed for each frame and averaged per stent. Histology was performed to show differences in coronary atherosclerosis. A range of plaque size and severity was found, from healthy segments to lipid-rich plaques. Accordingly, neointima responses ranged from uncovered struts, to minimal neointima, to fibrotic neointima. Lower plaque burden resulted in a fibrotic neointima at follow-up, reminiscent of minimally diseased swine coronary models. In contrast, higher plaque burden resulted in minimal neointima and more uncovered struts at follow-up, similarly to patients' responses. The presence of lipid-rich plaques resulted in more uncovered struts, which underscores the importance of advanced disease when performing safety and efficacy testing of DES.

Photoacoustic imaging on its way toward clinical utility: a tutorial review focusing on practical application in medicine.

Significance: Photoacoustic imaging (PAI) enables the visualization of optical contrast with ultrasonic imaging. It is a field of intense research, with great promise for clinical application. Understanding the principles of PAI is important for engineering research and image interpretation. Aim: In this tutorial review, we lay out the imaging physics, instrumentation requirements, standardization, and some practical examples for (junior) researchers, who have an interest in developing PAI systems and applications for clinical translation or applying PAI in clinical research. Approach: We discuss PAI principles and implementation in a shared context, emphasizing technical solutions that are amenable to broad clinical deployment, considering factors such as robustness, mobility, and cost in addition to image quality and quantification. Results: Photoacoustics, capitalizing on endogenous contrast or administered contrast agents that are approved for human use, yields highly informative images in clinical settings, which can support diagnosis and interventions in the future. Conclusion: PAI offers unique image contrast that has been demonstrated in a broad set of clinical scenarios. The transition of PAI from a "nice-to-have" to a "need-to-have" modality will require dedicated clinical studies that evaluate therapeutic decision-making based on PAI and consideration of the actual value for patients and clinicians, compared with the associated cost.

LED-based photoacoustic imaging for preoperative visualization of lymphatic vessels in patients with secondary limb lymphedema.

Lymphedema is the accumulation of protein-rich fluid in the interstitium (i.e., dermal backflow (DBF)). Preoperative imaging of the lymphatic vessels is a prerequisite for lymphovenous bypass surgical planning. We investigated the visualization of lymphatic vessels and veins using light-emitting diode (LED)-based photoacoustic imaging (PAI). Indocyanine-green mediated near-infrared fluorescence lymphography (NIRF-L) was done in fifteen patients with secondary limb lymphedema. Photoacoustic images were acquired in locations where lymphatic vessels and DBF were observed with NIRF-L. We demonstrated that LED-based PAI can visualize and differentiate lymphatic vessels and veins even in the presence of DBF. We observed lymphatic and blood vessels up to depths of 8.3 and 8.6 mm, respectively. Superficial lymphatic vessels and veins can be visualized using LED-based PAI even in the presence of DBF showing the potential for pre-operative assessment. Further development of the technique is needed to improve its usability in clinical settings.

Intravascular Imaging-Derived Physiology-Basic Principles and Clinical Application.

Intravascular imaging-derived physiology is emerging as a promising tool allowing simultaneous anatomic and functional lesion assessment. Recently, several optical coherence tomography-based and intravascular ultrasound-based fractional flow reserve (FFR) indices have been developed that compute FFR through computational fluid dynamics, fluid dynamics equations, or machine-learning methods. This review aims to provide an overview of the currently available intravascular imaging-based physiologic indices, their diagnostic performance, and clinical application.

Imaging of the Lymphatic Vessels for Surgical Planning: A Systematic Review.

BACKGROUND: Secondary lymphedema is a common complication after surgical or radiotherapeutic cancer treatment. (Micro) surgical intervention such as lymphovenous bypass and vascularized lymph node transfer is a possible solution in patients who are refractory to conventional treatment. Adequate imaging is needed to identify functional lymphatic vessels and nearby veins for surgical planning. METHODS: A systematic literature search of the Embase, MEDLINE ALL via Ovid, Web of Science Core Collection and Cochrane CENTRAL Register of Trials databases was conducted in February 2022. Studies reporting on lymphatic vessel detection in healthy subjects or secondary lymphedema of the limbs or head and neck were analyzed. RESULTS: Overall, 129 lymphatic vessel imaging studies were included, and six imaging modalities were identified. The aim of the studies was diagnosis, severity staging, and/or surgical planning. CONCLUSION: Due to its utility in surgical planning, near-infrared fluorescence lymphangiography (NIRF-L) has gained prominence in recent years relative to lymphoscintigraphy, the current gold standard for diagnosis and severity staging. Magnetic resonance lymphography (MRL) gives three-dimensional detailed information on the location of both lymphatic vessels and veins and the extent of fat hypertrophy; however, MRL is less practical for routine presurgical implementation due to its limited availability and high cost. High frequency ultrasound imaging can provide high resolution imaging of lymphatic vessels but is highly operator-dependent and accurate identification of lymphatic vessels is difficult. Finally, photoacoustic imaging (PAI) is a novel technique for visualization of functional lymphatic vessels and veins. More evidence is needed to evaluate the utility of PAI in surgical planning.

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.

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.].

Label-free analytic histology of carotid atherosclerosis by mid-infrared optoacoustic microscopy.

Background and aims: Analysis of atherosclerotic plaque composition is a vital tool for unraveling the pathological metabolic processes that contribute to plaque growth. Methods: We visualize the constitution of human carotid plaques by mid-infrared optoacoustic microscopy (MiROM), a method for label-free analytic histology that requires minimal tissue preparation, rapidly yielding large field-of-view en-face images with a resolution of a few micrometers. We imaged endarterectomy specimens (n = 3, 12 sections total) at specific vibrational modes, targeting carbohydrates, lipids and proteins. Additionally, we recorded spectra at selected tissue locations. We identified correlations in the variability in this high-dimensional data set using non-negative matrix factorization (NMF). Results: We visualized high-risk plaque features with molecular assignment. Consistent NMF components relate to different dominant tissue constituents, dominated by lipids, proteins, and cholesterol and carbohydrates respectively. Conclusions: These results introduce MiROM as an innovative, stain-free, analytic histology technology for the biochemical characterization of complex human vascular pathology.

Nanosecond SRS fiber amplifier for label-free near-infrared photoacoustic microscopy of lipids.

Near-infrared photoacoustics receives increasing interest as an intravital modality to sense key biomolecules. One of the most central types of biomolecules of interest are lipids as they constitute essential bio-hallmarks of cardiovascular and metabolic diseases and their in-vivo detection holds insightful information about disease progression and treatment monitoring. However, the full potential of near-infrared photoacoustic for high-resolution and high-sensitivity biomedical studies of lipids has so far not been exploited due a lack of appropriate excitation sources delivering short-pulses at high-repetition-rate, high-pulse-energy, and wavelength around 1200 nm. Here, we demonstrate a custom-built SRS fiber amplifier that provides optical excitations at 1192.8 nm, repetition rates of 200 kHz, pulse durations below 2 ns, and pulse energies beyond 5 µJ. We capitalize on the performance of our excitation source and show near-infrared photoacoustics resolving intrinsic lipid contrast in biomedically relevant specimens ranging from single cells to lipid-rich tissue with subcellular resolution.

Micro Spectroscopic Photoacoustic (µsPA) imaging of advanced carotid atherosclerosis.

Atherosclerosis is a lipid-driven and an inflammatory disease of the artery walls. The composition of atherosclerotic plaque stratifies the risk of a specific plaque to cause a cardiovascular event. In an optical resolution photoacoustic microscopy setup, of 45 µm resolution, we extracted plaque lipid photoacoustic (PA) spectral signatures of human endarterectomy samples in the range of 1150-1240 nm, using matrix assisted laser desorption ionization mass spectrometry imaging as a reference. We found plaque PA signals to correlate best with sphingomyelins and cholesteryl esters. PA signal spectral variations within the plaque area were compared to reference molecular patterns and absorption spectra of lipid laboratory standards. Variability in the lipid spectroscopic features extracted by principal component analysis of all samples revealed three distinct components with peaks at: 1164, 1188, 1196 and 1210 nm. This result will guide the development of PA-based atherosclerosis disease staging capitalizing on lipidomics of atherosclerotic tissue.

Photoacoustic flow velocity imaging based on complex field decorrelation.

Photoacoustic (PA) imaging can be used to monitor flowing blood inside the microvascular and capillary bed. Ultrasound speckle decorrelation based velocimetry imaging was previously shown to accurately estimate blood flow velocity in mouse brain (micro-)vasculature. Translating this method to photoacoustic imaging will allow simultaneous imaging of flow velocity and extracting functional parameters like blood oxygenation. In this study, we use a pulsed laser diode and a quantitative method based on normalized first order field autocorrelation function of PA field fluctuations to estimate flow velocities in an ink tube phantom and in the microvasculature of the chorioallantoic membrane of a chicken embryo. We demonstrate how the decorrelation time of signals acquired over frames are related to the flow speed and show that the PA flow analysis based on this approach is an angle independent flow velocity imaging method.

The Prognostic Value of a Validated and Automated Intravascular Ultrasound-Derived Calcium Score.

BACKGROUND: Coronary calcification has been linked to cardiovascular events. We developed and validated an algorithm to automatically quantify coronary calcifications on intravascular ultrasound (IVUS). We aimed to assess the prognostic value of an IVUS-calcium score (ICS) on patient-oriented composite endpoint (POCE). METHODS: We included patients that underwent coronary angiography plus pre-procedural IVUS imaging. The ICS was calculated per patient. The primary endpoint was a composite of all-cause mortality, stroke, myocardial infarction, and revascularization (POCE). RESULTS: In a cohort of 408 patients, median ICS was 85. Both an ICS ≥ 85 and a 100 unit increase in ICS increased the risk of POCE at 6-year follow-up (adjusted hazard ratio (aHR) 1.51, 95%CI 1.05-2.17, p value = 0.026, and aHR 1.21, 95%CI 1.04-1.41, p value = 0.014, respectively). CONCLUSIONS: The ICS, calculated by a validated automated algorithm derived from routine IVUS pullbacks, was strongly associated with the long-term risk of POCE.

Lipid signature of advanced human carotid atherosclerosis assessed by mass spectrometry imaging.

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 high-risk 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 with morphological 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.

Real-Time Coded Excitation Imaging Using a CMUT-Based Side Looking Array for Intravascular Ultrasound.

Intravascular ultrasound (IVUS) is a well-established diagnostic method that provides images of the vessel wall and atherosclerotic plaques. We investigate the potential for phased-array IVUS utilizing coded excitation (CE) for improving the penetration depth and image signal-to-noise ratio (SNR). It is realized on a new experimental broadband capacitive micromachined ultrasound transducer (CMUT) array, operated in collapse mode, with 96 elements placed at the circumference of a catheter tip with a 1.2- mm diameter. We characterized the array performance for CE imaging and showed that the -6-dB device bandwidth at a 30-V dc biasing is 25 MHz with a 20-MHz center frequency, with a transmit sensitivity of 37 kPa/V at that frequency. We designed a linear frequency modulation code to improve penetration depth by compensating for high-frequency attenuation while preserving resolution by a mismatched filter reconstruction. We imaged a wire phantom and a human coronary artery plaque. By assessing the image quality of the reconstructed wire phantom image, we achieved 60- and 70- µm axial resolutions using the short pulse and coded signal, respectively, and gained 8 dB in SNR for CE. Our developed system shows 20-frames/s, pixel-based beam-formed, real-time IVUS images.