Full-visibility 3D imaging of oxygenation and blood flow by simultaneous multispectral photoacoustic fluctuation imaging (MS-PAFI) and ultrasound Doppler.

We present a method and setup that provide complementary three-dimensional (3D) images of blood oxygenation (via quantitative photoacoustic imaging) and blood flow dynamics (via ultrasound Doppler). The proposed approach is label-free and exploits blood-induced fluctuations, and is implemented on a sparse array with only 256 elements, driven with a commercially available ultrasound electronics. We first implement 3D photoacoustic fluctuation imaging (PAFI) to image chicken embryo, and obtain full-visibility images of the vascular morphology. We obtain simultaneously 3D ultrasound power Doppler with a comparable image quality. We then introduce multispectral photoacoustic fluctuation imaging (MS-PAFI), and demonstrate that it can provide quantitative measurements of the absorbed optical energy density with full visibility and enhanced contrast, as compared to conventional delay-and-sum multispectral photoacoustic imaging. We finally showcase the synergy and complementarity between MS-PAFI, which provides 3D quantitative oxygenation (SO[Formula: see text]) imaging, and 3D ultrasound Doppler, which provides quantitative information on blood flow dynamics. MS-PAFI represents a promising alternative to model-based inversions with the advantage of resolving all the visibility artefacts without prior and regularization, by use of a straightforward processing scheme.

Compensating for visibility artefacts in photoacoustic imaging with a deep learning approach providing prediction uncertainties.

Conventional photoacoustic imaging may suffer from the limited view and bandwidth of ultrasound transducers. A deep learning approach is proposed to handle these problems and is demonstrated both in simulations and in experiments on a multi-scale model of leaf skeleton. We employed an experimental approach to build the training and the test sets using photographs of the samples as ground truth images. Reconstructions produced by the neural network show a greatly improved image quality as compared to conventional approaches. In addition, this work aimed at quantifying the reliability of the neural network predictions. To achieve this, the dropout Monte-Carlo procedure is applied to estimate a pixel-wise degree of confidence on each predicted picture. Last, we address the possibility to use transfer learning with simulated data in order to drastically limit the size of the experimental dataset.

Super-resolution photoacoustic and ultrasound imaging with sparse arrays.

It has previously been demonstrated that model-based reconstruction methods relying on a priori knowledge of the imaging point spread function (PSF) coupled to sparsity priors on the object to image can provide super-resolution in photoacoustic (PA) or in ultrasound (US) imaging. Here, we experimentally show that such reconstruction also leads to super-resolution in both PA and US imaging with arrays having much less elements than used conventionally (sparse arrays). As a proof of concept, we obtained super-resolution PA and US cross-sectional images of microfluidic channels with only 8 elements of a 128-elements linear array using a reconstruction approach based on a linear propagation forward model and assuming sparsity of the imaged structure. Although the microchannels appear indistinguishable in the conventional delay-and-sum images obtained with all the 128 transducer elements, the applied sparsity-constrained model-based reconstruction provides super-resolution with down to only 8 elements. We also report simulation results showing that the minimal number of transducer elements required to obtain a correct reconstruction is fundamentally limited by the signal-to-noise ratio. The proposed method can be straigthforwardly applied to any transducer geometry, including 2D sparse arrays for 3D super-resolution PA and US imaging.

Adaptive Spatiotemporal SVD Clutter Filtering for Ultrafast Doppler Imaging Using Similarity of Spatial Singular Vectors.

Singular value decomposition of ultrafast imaging ultrasonic data sets has recently been shown to build a vector basis far more adapted to the discrimination of tissue and blood flow than the classical Fourier basis, improving by large factor clutter filtering and blood flow estimation. However, the question of optimally estimating the boundary between the tissue subspace and the blood flow subspace remained unanswered. Here, we introduce an efficient estimator for automatic thresholding of subspaces and compare it to an exhaustive list of thirteen estimators that could achieve this task based on the main characteristics of the singular components, namely the singular values, the temporal singular vectors, and the spatial singular vectors. The performance of those fourteen estimators was tested in vitro in a large set of controlled experimental conditions with different tissue motion and flow speeds on a phantom. The estimator based on the degree of resemblance of spatial singular vectors outperformed all others. Apart from solving the thresholding problem, the additional benefit with this estimator was its denoising capabilities, strongly increasing the contrast to noise ratio and lowering the noise floor by at least 5 dB. This confirms that, contrary to conventional clutter filtering techniques that are almost exclusively based on temporal characteristics, efficient clutter filtering of ultrafast Doppler imaging cannot overlook space. Finally, this estimator was applied in vivo on various organs (human brain, kidney, carotid, and thyroid) and showed efficient clutter filtering and noise suppression, improving largely the dynamic range of the obtained ultrafast power Doppler images.

Overcoming the acoustic diffraction limit in photoacoustic imaging by the localization of flowing absorbers.

The resolution of photoacoustic imaging deep inside scattering media is limited by the acoustic diffraction limit. In this Letter, taking inspiration from super-resolution imaging techniques developed to beat the optical diffraction limit, we demonstrate that the localization of individual optical absorbers can provide super-resolution photoacoustic imaging well beyond the acoustic diffraction limit. As a proof-of-principle experiment, photoacoustic cross-sectional images of microfluidic channels were obtained with a 15 MHz linear capacitive micromachined ultrasonic transducer array, while absorbing beads were flown through the channels. The localization of individual absorbers allowed us to obtain a super-resolved cross-sectional image of the channels by reconstructing both the channel width and position with an accuracy better than λ/10. Given the discrete nature of endogenous absorbers such as red blood cells, or that of exogenous particular contrast agents, localization is a promising approach to push the current resolution limits of photoacoustic imaging.

Pulsed Cavitational Ultrasound Softening: a new non-invasive therapeutic approach of calcified bioprosthetic valve stenosis.

BACKGROUND: The majority of prosthetic heart valves currently implanted are tissue valves that can be expected to calcify with time and eventually fail. Surgical or percutaneous redux valve replacement is associated with higher rate of complications. We propose a novel non-invasive therapeutic approach based on the use of pulsed cavitational ultrasound (PCU) to improve the valvular function of degenerative calcified bioprosthesis. OBJECTIVES: Our study aims to demonstrate in vitro and in vivo on an ovine model that PCU can significantly improve the bioprosthesis opening by softening remotely the calcified stiff cusps. METHODS: All the experiments were performed on calcified bioprosthetic valves explanted from human patients. PCU was performed in vitro on calcified bioprosthesis mounted on a hydraulic bench with pulsatile flow (n=8) and in vivo on an ovine model with implanted calcified bioprosthesis (n=7). We used 3D echocardiography, pressure and flow sensors, quantitative stiffness evaluation using shear wave elastography, micro-CT imaging and histology to evaluate in vitro and in vivo the effect of PCU. RESULTS: The transvalvular gradient was found to decrease by a mean of 50% after PCU in both in vitro (from 21.1±3.9 to 9.6±1.7 mmHg, p<0.001) and in vivo setup (from 16.2±3.2 to 8.2±1.3 mmHg, p<0.001), with a decrease of valve stiffness (in vitro: from 105.8±9 to 46.6±4 kPa, p<0.001; in vivo: from 82.6±10 to 41.7±7 kPa, p<0.001) and an increase of valve area (from 1.10±0.1 to 1.58±0.1 cm2, p<0.001). Histology and micro-CT imaging showed modifications of calcification structure without loss of calcification volume or alteration of the leaflet superficial structures. CONCLUSIONS: We have demonstrated in vitro and in vivo that PCU can decrease a calcified bioprosthesis stenosis by softening the leaflets remotely. This new non-invasive approach has the potential to improve the outcome of patients with severe bioprosthesis stenosis.

Pulsed cavitational ultrasound for non-invasive chordal cutting guided by real-time 3D echocardiography.

AIMS: Basal chordae surgical section has been shown to be effective in reducing ischaemic mitral regurgitation (IMR). Achieving this section by non-invasive mean can considerably decrease the morbidity of this intervention on already infarcted myocardium. We investigated in vitro and in vivo the feasibility and safety of pulsed cavitational focused ultrasound (histotripsy) for non-invasive chordal cutting guided by real-time 3D echocardiography. METHODS AND RESULTS: Experiments were performed on 12 sheep hearts, 5 in vitro on explanted sheep hearts and 7 in vivo on beating sheep hearts. In vitro, the mitral valve (MV) apparatus including basal and marginal chordae was removed and fixed on a holder in a water tank. High-intensity ultrasound pulses were emitted from the therapeutic device (1-MHz focused transducer, pulses of 8 µs duration, peak negative pressure of 17 MPa, repetition frequency of 100 Hz), placed at a distance of 64 mm under 3D echocardiography guidance. In vivo, after sternotomy, the same therapeutic device was applied on the beating heart. We analysed MV coaptation and chordae by real-time 3D echocardiography before and after basal chordal cutting. After sacrifice, the MV apparatus were harvested for anatomical and histological post-mortem explorations to confirm the section of the chordae. In vitro, all chordae were completely cut after a mean procedure duration of 5.5 ± 2.5 min. The procedure duration was found to increase linearly with the chordae diameter. In vivo, the central basal chordae of the anterior leaflet were completely cut. The mean procedure duration was 20 ± 9 min (min = 14, max = 26). The sectioned chordae was visible on echocardiography, and MV coaptation remained normal with no significant mitral regurgitation. Anatomical and histological post-mortem explorations of the hearts confirmed the section of the chordae. CONCLUSIONS: Histotripsy guided by 3D echo achieved successfully to cut MV chordae in vitro and in vivo in beating heart. We hope that this technique will open the door in the near future to the non-invasive treatment of functional IMR.

Sono-photoacoustic imaging of gold nanoemulsions: Part I. Exposure thresholds.

Integrating high contrast bubbles from ultrasound imaging with plasmonic absorbers from photoacoustic imaging is investigated. Nanoemulsion beads coated with gold nanopsheres (NEB-GNS) are excited with simultaneous light (transient heat at the GNS's) and ultrasound (rarefactional pressure) resulting in a phase transition achievable under different scenarios, enhancing laser-induced acoustic signals and enabling specific detection of nanoprobes at lower concentration. An automated platform allowed dual parameter scans of both pressure and laser fluence while recording broadband acoustic signals. Two types of NEB-GNS and individual GNS were investigated and showed the great potential of this technique to enhance photoacoustic/acoustic signals. The NEB-GNS size distribution influences vaporization thresholds which can be reached at both permissible ultrasound and light exposures at deep penetration and at low concentrations of targets. This technique, called sono-photoacoustics, has great potential for targeted molecular imaging and therapy using compact nanoprobes with potentially high-penetrability into tissue.

Sono-photoacoustic imaging of gold nanoemulsions: Part II. Real time imaging.

Photoacoustic (PA) imaging using exogenous agents can be limited by degraded specificity due to strong background signals. This paper introduces a technique called sono-photoacoustics (SPA) applied to perfluorohexane nanodroplets coated with gold nanospheres. Pulsed laser and ultrasound (US) excitations are applied simultaneously to the contrast agent to induce a phase-transition ultimately creating a transient microbubble. The US field present during the phase transition combined with the large thermal expansion of the bubble leads to 20-30 dB signal enhancement. Aqueous solutions and phantoms with very low concentrations of this agent were probed using pulsed laser radiation at diagnostic exposures and a conventional US array used both for excitation and imaging. Contrast specificity of the agent was demonstrated with a coherent differential scheme to suppress US and linear PA background signals. SPA shows great potential for molecular imaging with ultrasensitive detection of targeted gold coated nanoemulsions and cavitation-assisted theranostic approaches.

Real-time integrated photoacoustic and ultrasound (PAUS) imaging system to guide interventional procedures: ex vivo study.

Because of depth-dependent light attenuation, bulky, low-repetition-rate lasers are usually used in most photoacoustic (PA) systems to provide sufficient pulse energies to image at depth within the body. However, integrating these lasers with real-time clinical ultrasound (US) scanners has been problematic because of their size and cost. In this paper, an integrated PA/US (PAUS) imaging system is presented operating at frame rates >30 Hz. By employing a portable, low-cost, low-pulse-energy (~2 mJ/pulse), high-repetition-rate (~1 kHz), 1053-nm laser, and a rotating galvo-mirror system enabling rapid laser beam scanning over the imaging area, the approach is demonstrated for potential applications requiring a few centimeters of penetration. In particular, we demonstrate here real-time (30 Hz frame rate) imaging (by combining multiple single-shot sub-images covering the scan region) of an 18-gauge needle inserted into a piece of chicken breast with subsequent delivery of an absorptive agent at more than 1-cm depth to mimic PAUS guidance of an interventional procedure. A signal-to-noise ratio of more than 35 dB is obtained for the needle in an imaging area 2.8 × 2.8 cm (depth × lateral). Higher frame rate operation is envisioned with an optimized scanning scheme.

Magneto-optical nanoparticles for cyclic magnetomotive photoacoustic imaging.

Photoacoustic imaging has emerged as a highly promising tool to visualize molecular events with deep tissue penetration. Like most other modalities, however, image contrast under in vivo conditions is far from optimal due to background signals from tissue. Using iron oxide-gold core-shell nanoparticles, we have previously demonstrated the concept of magnetomotive photoacoustic (mmPA) imaging, which is capable of dramatically reducing the influence of background signals and producing high-contrast molecular images. Here, we report two significant advances toward clinical translation of this technology. First, we introduce a new class of compact, uniform, magneto-optically coupled core-shell nanoparticles, prepared through localized copolymerization of polypyrrole (PPy) on an iron oxide nanoparticle surface. The resulting iron oxide-PPy nanoparticles feature high colloidal stability and solve the photoinstability and small-scale synthesis problems previously encountered by the gold coating approach. In parallel, we have developed a new generation of mmPA featuring cyclic magnetic motion and ultrasound speckle tracking (USST), whose imaging capture frame rate is several hundred times faster than the photoacoustic speckle tracking (PAST) method we demonstrated previously. These advances enable robust artifact elimination caused by physiologic motions and demonstrate the application of the mmPA technology for in vivo sensitive tumor imaging.

Shear wave elastography using amplitude-modulated acoustic radiation force and phase-sensitive optical coherence tomography.

Investigating the elasticity of ocular tissue (cornea and intraocular lens) could help the understanding and management of pathologies related to biomechanical deficiency. In previous studies, we introduced a setup based on optical coherence tomography for shear wave elastography (SWE) with high resolution and high sensitivity. SWE determines tissue stiffness from the propagation speed of shear waves launched within tissue. We proposed acoustic radiation force to remotely induce shear waves by focusing an ultrasound (US) beam in tissue, similar to several elastography techniques. Minimizing the maximum US pressure is essential in ophthalmology for safety reasons. For this purpose, we propose a pulse compression approach. It utilizes coded US emissions to generate shear waves where the energy is spread over a long emission, and then numerically compressed into a short, localized, and high-energy pulse. We used a 7.5-MHz single-element focused transducer driven by coded excitations where the amplitude is modulated by a linear frequency-swept square wave (1 to 7 kHz). An inverse filter approach was used for compression. We demonstrate the feasibility of performing shear wave elastography measurements in tissue-mimicking phantoms at low US pressures (mechanical index < 0.6)

Optimization of the laser irradiation pattern in a high frame rate integrated photoacoustic / ultrasound (PAUS) imaging system.

To integrate real-time photoacoustics (PA) into ultrasound (US) scanners and accelerate clinical translation of combined PAUS imaging, we previously developed a system using a portable, low-cost, low pulse energy, high-repetition rate laser (~1kHz) with a 1D galvo-mirror for rapid laser beam scanning over the imaging area. However, the frame rate and pulse energy are limited because of regulations on the radiance (1 W/cm2). Therefore, a laser scan scheme needs to be optimized to provide high frame rate within this safety limit. In addition, the laser light should be evenly distributed to minimize any artifacts caused by the scanning approach. In this paper, we calculated the laser light distribution using 3D Monte Carlo simulation and further developed the system to scan the laser beam in elevation as well as laterally using a 2-dimensional galvo-mirror scanner to achieve higher frame rates within the radiance safety limit. Insertion of a needle into chicken breast tissue was used to demonstrate our optimized scan scheme.

Real-time interleaved photoacoustic/ultrasound (PAUS) imaging for interventional procedure guidance.

Ultrasound-guided photoacoustic imaging has shown great potential for many clinical applications including vascular visualization, detection of nanoprobes sensing molecular profiles, and guidance of interventional procedures. However, bulky and costly lasers are usually required to provide sufficient pulse energies for deep imaging. The low pulse repetition rate also limits potential real-time applications of integrated photoacoustic/ultrasound (PAUS) imaging. With a compact and low-cost laser operating at a kHz repetition rate, we aim to integrate photoacoustics (PA) into a commercial ultrasound (US) machine utilizing an interleaved scanning approach for clinical translation, with imaging depth up to a few centimeters and frame rates > 30 Hz. Multiple PA sub-frames are formed by scanning laser firings covering a large scan region with a rotating galvo mirror, and then combined into a final frame. Ultrasound pulse-echo beams are interleaved between laser firings/PA receives. The approach was implemented with a diode-pumped laser, a commercial US scanner, and a linear array transducer. Insertion of an 18-gauge needle into a piece of chicken tissue, with subsequent injection of an absorptive agent into the tissue, was imaged with an integrated PAUS frame rate of 30 Hz, covering a 2.8 cm × 2.8 cm imaging plane. Given this real-time image rate and high contrast (> 40 dB at more than 1-cm depth in the PA image), we have demonstrated that this approach is potentially attractive for clinical procedure guidance.

Cyclic Magnetomotive Photoacoustic/Ultrasound Imaging.

Magnetomotive photoacoustic/ultrasound imaging has shown superior specificity in visualizing targeted objects at cellular and molecular levels. By detecting magnet-induced displacements, magnetic-particle-targeted objects can be differentiated from background signals insensitive to the magnetic field. Unfortunately, background physiologic motion interferes during measurement, such as cardiac-induced motion and respiration, greatly reducing the robustness of the technique. In this paper, we propose cyclic magnetomotive imaging with narrowband magnetic excitation. By synchronizing magnetic motion with the excitations, targeted objects moving coherently can be distinguished from background static signals and signals moving incoherently. HeLa cells targeted with magnetic nanoparticle-polymer core-shell particles were used as the targets for an initial test. A linear ultrasound array was interfaced with a commercial scanner to acquire a photoacoustic/ultrasound image sequence (maximum 1000 frames per second) during multi-cycle magnetic excitation (0.5 - 40 Hz frequency range) with an electromagnet. An image mask defined by a threshold on the displacement-coherence map was applied to the original images for background suppression. The results show that contrast was increased by more than 60 dB in an in-vitro experiment with the tagged cells fixed in a polyvinyl-alcohol gel and sandwiched between porcine liver tissues. Using a single sided system, cells injected subcutaneously on the back of a mouse were successfully differentiated from the background, with less than 20 µm coherent magnetic induced displacements isolated from millimetric background breathing motion. These results demonstrate the technique's motion robustness for highly sensitive and specific diagnosis.

Toric focusing for radiation force applications using a toric lens coupled to a spherically focused transducer.

Dynamic elastography using radiation force requires that an ultrasound field be focused during hundreds of microseconds at a pressure of several megapascals. Here, we address the importance of the focal geometry. Although there is usually no control of the elevational focal width in generating a tissue mechanical response, we propose a tunable approach to adapt the focus geometry that can significantly improve radiation force efficiency. Several thin, in-house-made polydimethylsiloxane lenses were designed to modify the focal spot of a spherical transducer. They exhibited low absorption and the focal spot widths were extended up to 8-fold in the elevation direction. Radiation force experiments demonstrated an 8-fold increase in tissue displacements using the same pressure level in a tissue-mimicking phantom with a similar shear wave spectrum, meaning it does not affect elastography resolution. Our results demonstrate that larger tissue responses can be obtained for a given pressure level, or that similar response can be reached at a much lower mechanical index (MI). We envision that this work will impact 3-D elastography using 2-D phased arrays, where such shaping can be achieved electronically with the potential for adaptive optimization.

Laser-induced cavitation in nanoemulsion with gold nanospheres for blood clot disruption: in vitro results.

Optically activated cavitation in a nanoemulsion contrast agent is proposed for therapeutic applications. With a 56°C boiling point perfluorohexane core and highly absorptive gold nanospheres at the oil-water interface, cavitation nuclei in the core can be efficiently induced with a laser fluence below medical safety limits (70 mJ/cm2 at 1064 nm). This agent is also sensitive to ultrasound (US) exposure and can induce inertial cavitation at a pressure within the medical diagnostic range. Images from a high-speed camera demonstrate bubble formation in these nanoemulsions. The potential of using this contrast agent for blood clot disruption is demonstrated in an in vitro study. The possibility of simultaneous laser and US excitation to reduce the cavitation threshold for therapeutic applications is also discussed.

Visualizing ultrasonically induced shear wave propagation using phase-sensitive optical coherence tomography for dynamic elastography.

We report on the use of phase-sensitive optical coherence tomography (PhS-OCT) to detect and track temporal and spatial shear wave propagation within tissue, induced by ultrasound radiation force. Kilohertz-range shear waves are remotely generated in samples using focused ultrasound emission and their propagation is tracked using PhS-OCT. Cross-sectional maps of the local shear modulus are reconstructed from local estimates of shear wave speed in tissue-mimicking phantoms. We demonstrate the feasibility of combining ultrasound radiation force and PhS-OCT to perform high-resolution mapping of the shear modulus.

Shear wave pulse compression for dynamic elastography using phase-sensitive optical coherence tomography.

Assessing the biomechanical properties of soft tissue provides clinically valuable information to supplement conventional structural imaging. In the previous studies, we introduced a dynamic elastography technique based on phase-sensitive optical coherence tomography (PhS-OCT) to characterize submillimetric structures such as skin layers or ocular tissues. Here, we propose to implement a pulse compression technique for shear wave elastography. We performed shear wave pulse compression in tissue-mimicking phantoms. Using a mechanical actuator to generate broadband frequency-modulated vibrations (1 to 5 kHz), induced displacements were detected at an equivalent frame rate of 47 kHz using a PhS-OCT. The recorded signal was digitally compressed to a broadband pulse. Stiffness maps were then reconstructed from spatially localized estimates of the local shear wave speed. We demonstrate that a simple pulse compression scheme can increase shear wave detection signal-to-noise ratio (>12 dB gain) and reduce artifacts in reconstructing stiffness maps of heterogeneous media.

Shear modulus imaging by direct visualization of propagating shear waves with phase-sensitive optical coherence tomography.

We propose an integrated method combining low-frequency mechanics with optical imaging to map the shear modulus within the biological tissue. Induced shear wave propagating in tissue is tracked in space and time using phase-sensitive optical coherence tomography (PhS-OCT). Local estimates of the shear-wave speed obtained from tracking results can image the local shear modulus. A PhS-OCT system remotely records depth-resolved, dynamic mechanical waves at an equivalent frame rate of ∼47 kHz with the high spatial resolution. The proposed method was validated by examining tissue-mimicking phantoms made of agar and light scattering material. Results demonstrate that the shear wave imaging can accurately map the elastic moduli of these phantoms.