Pulse-echo ultrasound attenuation tomography.

Objective.We present the first fully two-dimensional attenuation imaging technique developed for pulse-echo ultrasound systems. Unlike state-of-the-art techniques, which use line-by-line acquisitions, our method uses steered emissions to constrain attenuation values at each location with multiple crossing wave paths, essential to resolve the spatial variations of this tissue property.Approach.At every location, we compute normalized cross-correlations between the beamformed images that are obtained from emissions at different steering angles. We demonstrate that their log-amplitudes provide the changes between attenuation-induced amplitude losses undergone by the different incident waves. This allows us to formulate a linear tomographic problem, which we efficiently solve via a Tikhonov-regularized least-squares approach.Main results.The performance of our tomography technique is first validated in numerical examples and then experimentally demonstrated in custom-made tissue-mimicking phantoms with inclusions of varying size, echogenicity, and attenuation. We show that this technique is particularly good at resolving lateral variations in tissue attenuation and remains accurate in media with varying echogenicity.Significance.Based on a similar principle, this method can be easily combined with computed ultrasound tomography in echo mode for speed-of-sound imaging, paving the way towards a multi-modal ultrasound tomography framework characterizing multiple acoustic tissue properties simultaneously.

Fluorescence lifetime imaging and phasor analysis of intracellular porphyrinic photosensitizers applied with different polymeric formulations.

The fluorescence lifetime of a porphyrinic photosensitizer (PS) is an important parameter to assess the aggregation state of the PS even in complex biological environments. Aggregation-induced quenching of the PS can significantly reduce the yield of singlet oxygen generation and thus its efficiency as a medical drug in photodynamic therapy (PDT) of diseased tissues. Hydrophobicity and the tendency to form aggregates pose challenges on the development of efficient PSs and often require carrier systems. A systematic study was performed to probe the impact of PS structure and encapsulation into polymeric carriers on the fluorescence lifetime in solution and in the intracellular environment. Five different porphyrinic PSs including chlorin e6 (Ce6) derivatives and tetrakis(m-hydroxyphenyl)-porphyrin and -chlorin were studied in free form and combined with polyvinylpyrrolidone (PVP) or micelles composed of triblock-copolymers or Cremophor. Following incubation of HeLa cells with these systems, fluorescence lifetime imaging combined with phasor analysis and image segmentation was applied to study the lifetime distribution in the intracellular surrounding. The data suggest that for free PSs, the structure-dependent cell uptake pathways determine their state and emission lifetimes. PS localization in the plasma membrane yielded mostly monomers with long fluorescence lifetimes whereas the endocytic pathway with subsequent lysosomal deposition adds a short-lived component for hydrophilic anionic PSs. Prolonged incubation times led to increasing contributions from short-lived components that derive from aggregates mainly localized in the cytoplasm. Encapsulation of PSs into polymeric carriers led to monomerization and mostly fluorescence emission decays with long fluorescence lifetimes in solution. However, the efficiency depended on the binding strength that was most pronounced for PVP. In the cellular environment, PVP was able to maintain monomeric long-lived species over prolonged incubation times. This was most pronounced for Ce6 derivatives with a logP value around 4.5. Micellar encapsulation led to faster release of the PSs resulting in multiple components with long and short fluorescence lifetimes. The hydrophilic hardly aggregating PS exhibited a mostly stable invariant lifetime distribution over time with both carriers. The presented data are expected to contribute to optimized PDT treatment protocols and improved PS-carrier design for preventing intracellular fluorescence quenching. In conclusion, amphiphilic and concurrent hydrophobic PSs with high membrane affinity as well as strong binding to the carrier have best prospects to maintain their photophysical properties in vivo and serve thus as efficient photodynamic diagnosis and PDT drugs.

First-in-human diagnostic study of hepatic steatosis with computed ultrasound tomography in echo mode.

BACKGROUND: Non-alcoholic fatty liver disease is rapidly emerging as the leading global cause of chronic liver disease. Efficient disease management requires low-cost, non-invasive techniques for diagnosing hepatic steatosis accurately. Here, we propose quantifying liver speed of sound (SoS) with computed ultrasound tomography in echo mode (CUTE), a recently developed ultrasound imaging modality adapted to clinical pulse-echo systems. CUTE reconstructs the spatial distribution of SoS by measuring local echo phase shifts when probing tissue at varying steering angles in transmission and reception. METHODS: In this first-in-human phase II diagnostic study, we evaluated the liver of 22 healthy volunteers and 22 steatotic patients. We used conventional B-mode ultrasound images and controlled attenuation parameter (CAP) to diagnose the presence (CAP≥ 280 dB/m) or absence (CAP < 248 dB/m) of steatosis in the liver. A fully integrated convex-probe CUTE implementation was developed on the ultrasound system to estimate liver SoS. We investigated its diagnostic value via the receiver operating characteristic (ROC) analysis and correlation to CAP measurements. RESULTS: We show that liver CUTE-SoS estimates correlate strongly (r = -0.84, p = 8.27 × 10-13) with CAP values and have 90.9% (95% confidence interval: 84-100%) sensitivity and 95.5% (81-100%) specificity for differentiating between normal and steatotic livers (area under the ROC curve: 0.93-1.0). CONCLUSIONS: Our results demonstrate that liver CUTE-SoS is a promising quantitative biomarker for diagnosing liver steatosis. This is a necessary first step towards establishing CUTE as a new quantitative add-on to diagnostic ultrasound that can potentially be as versatile as conventional ultrasound imaging.

Non-alcoholic fatty liver disease (NAFLD), characterized by fat accumulation in the liver, is rapidly becoming the most common cause of chronic liver disease worldwide. Therefore, there is an urgent need to develop accurate diagnostic techniques that are inexpensive, non-invasive, and broadly available. Ultrasound imaging systems, which use sound waves to produce images of internal body structures, possess these qualities but cannot currently diagnose NAFLD accurately. Here, we propose to use a recently developed technique called computed ultrasound tomography in echo mode (CUTE). It measures the speed at which ultrasound waves propagate in tissues, a property that substantially varies with the fat content. We show that CUTE measurements allow us to accurately distinguish the livers of healthy people from those of individuals diagnosed with NAFLD. This promising finding encourages the integration of CUTE into standard ultrasound systems.

Wide-field time-of-flight measurements of highly scattering media.

We present a setup that makes use of a time-resolved single-photon camera to determine the scattering parameters of media. The measurement is realized in a non-contact way, both for the illumination laser and the detection. By fitting the time-of-flight acquired distributions at different spatial positions with the diffusion equation, we retrieve the reduced scattering coefficients of a highly diffusive isotropic reference media for wavelengths in the range from 540 to 840 nm.

Compromised Blood-Brain Barrier Junctions Enhance Melanoma Cell Intercalation and Extravasation.

Melanoma frequently metastasises to the brain, and a detailed understanding of the molecular and cellular mechanisms underlying melanoma cell extravasation across the blood-brain barrier (BBB) is important for preventing brain metastasis formation. Making use of primary mouse brain microvascular endothelial cells (pMBMECs) as an in vitro BBB model, we imaged the interaction of melanoma cells into pMBMEC monolayers. We observed exclusive junctional intercalation of melanoma cells and confirmed that melanoma-induced pMBMEC barrier disruption can be rescued by protease inhibition. Interleukin (IL)-1ß stimulated pMBMECs or PECAM-1-knockout (-ko) pMBMECs were employed to model compromised BBB barrier properties in vitro and to determine increased melanoma cell intercalation compared to pMBMECs with intact junctions. The newly generated brain-homing melanoma cell line YUMM1.1-BrM4 was used to reveal increased in vivo extravasation of melanoma cells across the BBB of barrier-compromised PECAM-1-deficient mice compared to controls. Taken together, our data indicate that preserving BBB integrity is an important measure to limit the formation of melanoma-brain metastasis.

The Cilialyzer - A freely available open-source software for the analysis of mucociliary activity in respiratory cells.

BACKGROUND AND OBJECTIVE: Primary ciliary dyskinesia (PCD) is a rare genetic disorder causing a defective ciliary structure, which predominantly leads to an impaired mucociliary clearance and associated airway disease. As there is currently no single diagnostic gold standard test, PCD is diagnosed by a combination of several methods comprising genetic testing and the examination of the ciliary structure and function. Among the approved diagnostic methods, only high-speed video microscopy (HSVM) allows to directly observe the ciliary motion and therefore, to directly assess ciliary function. In the present work, we present our recently developed freely available open-source software - termed "Cilialyzer", which has been specifically designed to support and facilitate the analysis of the mucociliary activity in respiratory epithelial cells captured by high-speed video microscopy. METHODS: In its current state, the Cilialyzer software enables clinical PCD analysts to load, preprocess and replay recorded image sequences as well as videos with a feature-rich replaying module facilitating the commonly performed qualitative visual assessment of ciliary function (including the assessment of the ciliary beat pattern). The image processing methods made accessible through an intuitive user interface allow clinical specialists to comfortably compute the ciliary beating frequency (CBF), the activity map and the "frequency correlation length" - an observable getting newly introduced. Furthermore, the Cilialyzer contains a simple-to-use particle tracking interface to determine the mucociliary transport speed. RESULTS: Cilialyzer is fully written in the Python programming language and freely available under the terms of the MIT license. The proper functioning of the computational analysis methods constituting the Cilialyzer software is demonstrated by using simulated and representative sample data from clinical practice. Additionally, the software was used to analyze high-speed videos showing samples obtained from healthy controls and genetically confirmed PCD cases (DNAI1 and DNAH11 mutations) to show its clinical applicability. CONCLUSIONS: Cilialyzer serves as a useful clinical tool for PCD analysts and provides new quantitative information awaiting to be clinically evaluated using cohorts of PCD. As Cilialyzer is freely available under the terms of a permissive open-source license, it serves as a ground frame for further development of computational methods aiming at the quantification and automation of the analysis of mucociliary activity captured by HSVM.

Comparing PVP and Polymeric Micellar Formulations of a PEGylated Photosensitizing Phthalocyanine by NMR and Optical Techniques.

Phthalocyanines are ideal candidates as photosensitizers for photodynamic therapy (PDT) of cancer due to their favorable chemical and photophysical properties. However, their tendency to form aggregates in water reduces PDT efficacy and poses challenges in obtaining efficient forms of phthalocyanines for therapeutic applications. In the current work, polyvinylpyrrolidone (PVP) and micellar formulations were compared for encapsulating and monomerizing a water-soluble zinc phthalocyanine bearing four non-peripheral triethylene glycol chains (Pc1). 1H NMR spectroscopy combined with UV-vis absorption and fluorescence spectroscopy revealed that Pc1 exists as a mixture of regioisomers in monomeric form in dimethyl sulfoxide but forms dimers in an aqueous buffer. PVP, polyethylene glycol castor oil (Kolliphor RH40), and three different triblock copolymers with varying proportions of polyethylene and polypropylene glycol units (termed P188, P84, and F127) were tested as micellar carriers for Pc1. 1H NMR chemical shift analysis, diffusion-ordered spectroscopy, and 2D nuclear Overhauser enhancement spectroscopy was applied to monitor the encapsulation and localization of Pc1 at the polymer interface. Kolliphor RH40 and F127 micelles exhibited the highest affinity for encapsulating Pc1 in the micellar core and resulted in intense Pc1 fluorescence emission as well as efficient singlet oxygen formation along with PVP. Among the triblock copolymers, efficiency in binding and dimer dissolution decreased in the order F127 > P84 > P188. PVP was a strong binder for Pc1. However, Pc1 molecules are rather surface-attached and exist as monomer and dimer mixtures. The results demonstrate that NMR combined with optical spectroscopy offer powerful tools to assess parameters like drug binding, localization sites, and dynamic properties that play key roles in achieving high host-guest compatibility. With the corresponding adjustments, polymeric micelles can offer simple and easily accessible drug delivery systems optimizing phthalocyanines' properties as efficient photosensitizers.

Excluding Echo Shift Noise in Real-Time Pulse-Echo Speed-of-Sound Imaging.

Computed ultrasound tomography in echo mode (CUTE) allows real-time imaging of the tissue speed of sound (SoS) using handheld ultrasound. The SoS is retrieved by inverting a forward model that relates the spatial distribution of the tissue SoS to echo shift maps detected between varying transmit and receive angles. Despite promising results, in vivo SoS maps often show artifacts due to elevated noise in echo shift maps. To minimize artifacts, we propose a technique where an individual SoS map is reconstructed for each echo shift map separately, as opposed to a single SoS map from all echo shift maps simultaneously. The final SoS map is then obtained as a weighted average over all SoS maps. Due to the partial redundancy between different angle combinations, artifacts that appear only in a subset of the individual maps can be excluded via the averaging weights. We investigate this real-time capable technique in simulations using two numerical phantoms, one with a circular inclusion and one with two layers. Our results demonstrate that the SoS maps reconstructed using the proposed technique are equivalent to the ones using simultaneous reconstruction when considering uncorrupted data but show significantly reduced artifact level for data that are corrupted by noise.

Investigation of the Influence of Pulse Duration and Application Mode on Microsecond Laser Microsurgery of the Retinal Pigment Epithelium.

Optical microsurgery confined to the retinal pigment epithelium (RPE) requires locally optimized laser parameters and reliable real-time feedback dosimetry (RFD) to prevent unwanted neuroretinal overexposure. This study aimed to compare pulses of different durations and application modes (single, ramp, burst). Moreover, optical coherence tomography (OCT)-based RFD was investigated in an ex vivo experiment, utilizing nine porcine eyes that were exposed to laser pulses of 8, 12, 16 and 20 µs duration (wavelength: 532 nm, exposure area: 90 × 90 µm2, radiant exposure: 247 to 1975 mJ/µm2). Simultaneously, time-resolved OCT M-scans were recorded (central wavelength: 870 nm, scan rate: 85 kHz) for RFD. Post irradiation, retinal changes were assessed with color fundus photography (CFP) and cross-sectional OCT B-scans. RPE cell damage was quantified via fluorescence-based cell viability assay and compared to the OCT dosimetry feedback. Our experiments indicate cumulative RPE damage for pulse bursts of 16 µs and 20 µs, whereas no cumulative effects were found for pulse durations of 8 µs and 12 µs applied in ramp mode. According to statistical analysis, OCT-RFD correctly detected RPE cell damage with 96% sensitivity and 97% specificity using pulses of 8 µs duration in ramp mode.

In Vitro Effect of Combined Hypertonic Saline and Salbutamol on Ciliary Beating Frequency and Mucociliary Transport in Human Nasal Epithelial Cells of Healthy Volunteers and Patients with Cystic Fibrosis.

Background: Inhalation of hypertonic saline (HS) is standard of care in patients with cystic fibrosis (CF). However, it is unclear if adding salbutamol has-besides bronchodilation-further benefits, for example, on the mucociliary clearance. We assessed this in vitro by measuring the ciliary beating frequency (CBF) and the mucociliary transport rate (MCT) in nasal epithelial cells (NECs) of healthy volunteers and patients with CF. Aims: To investigate the effect of HS, salbutamol, and its combination on (muco)ciliary activity of NECs in vitro, and to assess potential differences between healthy controls and patients with CF. Methods: NECs obtained from 10 healthy volunteers and 5 patients with CF were differentiated at the air-liquid interface and aerosolized with 0.9% isotonic saline ([IS] control), 6% HS, 0.06% salbutamol, or combined HS and salbutamol. CBF and MCT were monitored over 48-72 hours. Results: In NECs of healthy controls, the absolute CBF increase was comparable for all substances, but CBF dynamics were different: HS increased CBF slowly and its effect lasted for an extended period, salbutamol and IS increased CBF rapidly and the effect subsided similarly fast, and HS and salbutamol resulted in a rapid and long-lasting CBF increase. Results for CF cells were comparable, but less pronounced. Similar to CBF, MCT increased after the application of all the tested substances. Conclusion: CBF and MCT of NECs of healthy participants and CBF of patients with CF increased upon treatment with aerosolized IS, HS, salbutamol, or HS and salbutamol, showing a relevant effect for all tested substances. The difference in the CBF dynamics can be explained by the fact that the properties of the mucus are changed differently by different saline concentrations.

Real-time OCT feedback-controlled RPE photodisruption in ex vivo porcine eyes using 8 microsecond laser pulses.

Selective retinal pigment epithelium (RPE) photodisruption requires reliable real-time feedback dosimetry (RFD) to prevent unwanted overexposure. In this study, optical coherence tomography (OCT) based RFD was investigated in ex vivo porcine eyes exposed to laser pulses of 8 µs duration (wavelength: 532 nm, exposure area: 90 × 90 µm2, radiant exposure: 247 to 1975 mJ/µm2). For RFD, fringe washouts in time-resolved OCT M-scans (central wavelength: 870 nm, scan rate: 85 kHz) were compared to an RPE cell viability assay. Statistical analysis revealed a moderate correlation between RPE lesion size and applied treatment energy, suggesting RFD adaptation to inter- and intraindividual RPE pigmentation and ocular transmission.

Pulse-echo speed-of-sound imaging using convex probes.

Computed ultrasound tomography in echo mode (CUTE) is a new ultrasound (US)-based medical imaging modality with promise for diagnosing various types of disease based on the tissue's speed of sound (SoS). It is developed for conventional pulse-echo US using handheld probes and can thus be implemented in state-of-the-art medical US systems. One promising application is the quantification of the liver fat fraction in fatty liver disease. So far, CUTE was using linear array probes where the imaging depth is comparable to the aperture size. For liver imaging, however, convex probes are preferred since they provide a larger penetration depth and a wider view angle allowing to capture a large area of the liver. With the goal of liver imaging in mind, we adapt CUTE to convex probes, with a special focus on discussing strategies that make use of the convex geometry in order to make our implementation computationally efficient. We then demonstrate in an abdominal imaging phantom that accurate quantitative SoS using convex probes is feasible, in spite of the smaller aperture size in relation to the image area compared to linear arrays. A preliminaryin vivoresult of liver imaging confirms this outcome, but also indicates that deep quantitative imaging in the real liver can be more challenging, probably due to the increased complexity of the tissue compared to phantoms.

Machine learning enabled multiple illumination quantitative optoacoustic oximetry imaging in humans.

Optoacoustic (OA) imaging is a promising modality for quantifying blood oxygen saturation (sO2) in various biomedical applications - in diagnosis, monitoring of organ function, or even tumor treatment planning. We present an accurate and practically feasible real-time capable method for quantitative imaging of sO2 based on combining multispectral (MS) and multiple illumination (MI) OA imaging with learned spectral decoloring (LSD). For this purpose we developed a hybrid real-time MI MS OA imaging setup with ultrasound (US) imaging capability; we trained gradient boosting machines on MI spectrally colored absorbed energy spectra generated by generic Monte Carlo simulations and used the trained models to estimate sO2 on real OA measurements. We validated MI-LSD in silico and on in vivo image sequences of radial arteries and accompanying veins of five healthy human volunteers. We compared the performance of the method to prior LSD work and conventional linear unmixing. MI-LSD provided highly accurate results in silico and consistently plausible results in vivo. This preliminary study shows a potentially high applicability of quantitative OA oximetry imaging, using our method.

A quantitative interspecies comparison of the respiratory mucociliary clearance mechanism.

Collectively coordinated ciliary activity propels the airway mucus, which lines the luminal surface of the vertebrate respiratory system, in cranial direction. Our contemporary understanding on how the quantitative characteristics of the metachronal wave field determines the resulting mucociliary transport is still limited, partly due to the sparse availability of quantitative observational data. We employed high-speed video reflection microscopy to image and quantitatively characterize the metachronal wave field as well as the mucociliary transport in excised bovine, porcine, ovine, lapine, turkey and ostrich samples. Image processing techniques were used to determine the ciliary beating frequency (CBF), the velocity and wavelength of the metachronal wave and the mucociliary transport velocity. The transport direction was found to strongly correlate with the mean wave propagation direction in all six species. The CBF yielded similar values (10-15 Hz) for all six species. Birds were found to exhibit higher transport speeds (130-260 [Formula: see text]m/s) than mammals (20-80 [Formula: see text]m/s). While the average transport direction significantly deviates from the tracheal long axis in mammals, no significant deviation was found in birds. The metachronal waves were found to propagate at about 4-8 times the speed of mucociliary transport in mammals, whereas in birds they propagate at about the transport speed. The mucociliary transport in birds is fast and roughly follows the TLA, whereas the transport is slower and proceeds along a left-handed spiral in mammals. The longer wavelengths and the lower ratio between the metachronal wave speed and the mucociliary transport speed provide evidence that the mucociliary clearance mechanism operates differently in birds than in mammals.

Selective Large-Area Retinal Pigment Epithelial Removal by Microsecond Laser in Preparation for Cell Therapy.

Purpose: Cell therapy is a promising treatment for retinal pigment epithelium (RPE)-associated eye diseases such as age-related macular degeneration. Herein, selective microsecond laser irradiation targeting RPE cells was used for minimally invasive, large-area RPE removal in preparation for delivery of retinal cell therapeutics. Methods: Ten rabbit eyes were exposed to laser pulses 8, 12, 16, and 20 µs in duration (wavelength, 532 nm; top-hat beam profile, 223 × 223 µm²). Post-irradiation retinal changes were assessed with fluorescein angiography (FA), indocyanine green angiography (ICGA), and optical coherence tomography (OCT). RPE viability was evaluated with an angiographic probit model. Following vitrectomy, a subretinal injection of balanced salt solution was performed over a lasered (maximum 13.6 mm2) and untreated control area. Bleb retinal detachment (bRD) morphology was then evaluated by intraoperative OCT. Results: Within 1 hour after irradiation, laser lesions showed FA and ICGA leakage. OCT revealed that large-area laser damage was limited to the RPE. The angiographic median effective dose irradiation thresholds (ED50) were 45 µJ (90 mJ/cm2) at 8 µs, 52 µJ (104 mJ/cm2) at 12 µs, 59 µJ (118 mJ/cm2) at 16 µs, and 71 µJ (142 mJ/cm2) at 20 µs. Subretinal injection over the lasered area resulted in a controlled, shallow bRD rise, whereas control blebs were convex in shape, with less predictable spread. Conclusions: Large-area, laser-based removal of host RPE without visible photoreceptor damage is possible and facilitates surgical retinal detachment. Translational Relevance: Selective microsecond laser-based, large-area RPE removal prior to retinal cell therapy may reduce iatrogenic trauma.

Backscattering polarimetric imaging of the human brain to determine the orientation and degree of alignment of nerve fiber bundles.

The nerve fiber bundles constitutive of the white matter in the brain are organized in such a way that they exhibit a certain degree of structural anisotropy and birefringence. The birefringence exhibited by such aligned fibrous tissue is known to be extremely sensitive to small pathological alterations. Indeed, highly aligned anisotropic fibers exhibit higher birefringence than structures with weaker alignment and anisotropy, such as cancerous tissue. In this study, we performed experiments on thick coronal slices of a healthy human brain to explore the possibility of (i) measuring, with a polarimetric microscope the birefringence exhibited by the white matter and (ii) relating the measured birefringence to the fiber orientation and the degree of alignment. This is done by analyzing the spatial distribution of the degree of polarization of the backscattered light and its variation with the polarization state of the probing beam. We demonstrate that polarimetry can be used to reliably distinguish between white and gray matter, which might help to intraoperatively delineate unstructured tumorous tissue and well organized healthy brain tissue. In addition, we show that our technique is able to sensitively reconstruct the local mean nerve fiber orientation in the brain, which can help to guide tumor resections by identifying vital nerve fiber trajectories thereby improving the outcome of the brain surgery.

Multiple illumination learned spectral decoloring for quantitative optoacoustic oximetry imaging.

SIGNIFICANCE: Quantitative measurement of blood oxygen saturation (sO2) with optoacoustic (OA) imaging is one of the most sought after goals of quantitative OA imaging research due to its wide range of biomedical applications. AIM: A method for accurate and applicable real-time quantification of local sO2 with OA imaging. APPROACH: We combine multiple illumination (MI) sensing with learned spectral decoloring (LSD). We train LSD feedforward neural networks and random forests on Monte Carlo simulations of spectrally colored absorbed energy spectra, to apply the trained models to real OA measurements. We validate our combined MI-LSD method on a highly reliable, reproducible, and easily scalable phantom model, based on copper and nickel sulfate solutions. RESULTS: With this sulfate model, we see a consistently high estimation accuracy using MI-LSD, with median absolute estimation errors of 2.5 to 4.5 percentage points. We further find fewer outliers in MI-LSD estimates compared with LSD. Random forest regressors outperform previously reported neural network approaches. CONCLUSIONS: Random forest-based MI-LSD is a promising method for accurate quantitative OA oximetry imaging.

Bayesian Approach for a Robust Speed-of-Sound Reconstruction Using Pulse-Echo Ultrasound.

Computed ultrasound tomography in echo mode (CUTE) is a promising ultrasound (US) based multi-modal technique that allows to image the spatial distribution of speed of sound (SoS) inside tissue using hand-held pulse-echo US. It is based on measuring the phase shift of echoes when detected under varying steering angles. The SoS is then reconstructed using a regularized inversion of a forward model that describes the relation between the SoS and echo phase shift. Promising results were obtained in phantoms when using a Tikhonov-type regularization of the spatial gradient (SG) of SoS. In-vivo, however, clutter and aberration lead to an increased phase noise. In many subjects, this phase noise causes strong artifacts in the SoS image when using the SG regularization. To solve this shortcoming, we propose to use a Bayesian framework for the inverse calculation, which includes a priori statistical properties of the spatial distribution of the SoS to avoid noise-related artifacts in the SoS images. In this study, the a priori model is based on segmenting the B-Mode image. We show in a simulation and phantom study that this approach leads to SoS images that are much more stable against phase noise compared to the SG regularization. In a preliminary in-vivo study, a reproducibility in the range of 10 ms-1 was achieved when imaging the SoS of a volunteer's liver from different scanning locations. These results demonstrate the diagnostic potential of CUTE for example for the staging of fatty liver disease.

A novel technique for laser-assisted revascularization: an in vitro pilot study.

The common limitation of surgical revascularization procedures for severe tissue ischemia due to cardiovascular diseases is the need to interrupt blood flow during the intervention. We aim to introduce a new technique that allows a sutureless, non-occlusive revascularization. A 3-step technique was developed using rabbit's aorta to simulate a side-to-side anastomosis model. It enables the creation of a bypass circuit for revascularization. The first step was the soldering of 2 vessels in a side-to-side fashion based on the laser-assisted vascular anastomosis (LAVA) principle using a diode laser emitting irradiation at 810 nm with an albumin-based solder patch between them, followed by the creation of a channel within the patch using either a holmium-doped yttrium aluminum garnet laser (Ho:YAG) at λ = 2100 nm or a xenon-chloride excimer laser (XeCl) at λ = 308 nm. Thereby, a bypass circuit was created, thus allowing a non-ischemic revascularization. The system was deemed functional when a flow was observed across the anastomosis. The highest average tensile strength recorded after side-to-side LAVA using a diode laser power of 3.2 W for 60 s was 2278.6 ± 800 mN (n = 20). The Ho:YAG laser created the channels with less tension on the anastomosis than the excimer laser. Histological analysis showed limited thermal damage and good patch-tissue adaptation. The preliminary results of this feasibility study outline the foundations for an entirely sutureless laser-assisted revascularization procedure. The next studies will evaluate the rheological parameters across the bypass circuit to optimize the post-anastomotic flow.

Multi-scale alignment of respiratory cilia and its relation to mucociliary function.

The tracheobronchial tree is lined by a mucociliary epithelium containing millions of multiciliated cells. Their integrated oscillatory activity continuously propels an overlying pollution-protecting mucus layer in cranial direction, leading to mucociliary clearance - the primary defence mechanism of the airways. Mucociliary transport is commonly thought to co-emerge with the collective ciliary motion pattern under appropriate geometrical and rheological conditions. Proper ciliary alignment is therefore considered essential to establish mucociliary clearance in the respiratory system. Here, we used volume electron microscopy in combination with high-speed reflection contrast microscopy in order to examine ciliary orientation and its spatial organization, as well as to measure the propagation direction of metachronal waves and the direction of mucociliary transport on bovine tracheal epithelia with reference to the tracheal long axis (TLA). Ciliary orientation is measured in terms of the basal body orientation (BBO) and the axonemal orientation (AO), which are commonly considered to coincide, both equivalently indicating the effective stroke as well as the mucociliary transport direction. Our results, however, reveal that only the AO is in line with the mucociliary transport, which was found to run along a left-handed helical trajectory, whereas the BBO was found to be aligned with the TLA. Furthermore, we show that even if ciliary orientation remains consistent between adjacent cells, ciliary orientation exhibits a gradual shift within individual cells. Together with the symplectic beating geometry, this intracellular orientational pattern could provide for the propulsion of highly viscous mucus and likely constitutes a compromise between efficiency and robustness.