CNN-Based Image Reconstruction Method for Ultrafast Ultrasound Imaging.

Ultrafast ultrasound (US) revolutionized biomedical imaging with its capability of acquiring full-view frames at over 1 kHz, unlocking breakthrough modalities such as shear-wave elastography and functional US neuroimaging. Yet, it suffers from strong diffraction artifacts, mainly caused by grating lobes, sidelobes, or edge waves. Multiple acquisitions are typically required to obtain a sufficient image quality, at the cost of a reduced frame rate. To answer the increasing demand for high-quality imaging from single unfocused acquisitions, we propose a two-step convolutional neural network (CNN)-based image reconstruction method, compatible with real-time imaging. A low-quality estimate is obtained by means of a backprojection-based operation, akin to conventional delay-and-sum beamforming, from which a high-quality image is restored using a residual CNN with multiscale and multichannel filtering properties, trained specifically to remove the diffraction artifacts inherent to ultrafast US imaging. To account for both the high dynamic range and the oscillating properties of radio frequency US images, we introduce the mean signed logarithmic absolute error (MSLAE) as a training loss function. Experiments were conducted with a linear transducer array, in single plane-wave (PW) imaging. Trainings were performed on a simulated dataset, crafted to contain a wide diversity of structures and echogenicities. Extensive numerical evaluations demonstrate that the proposed approach can reconstruct images from single PWs with a quality similar to that of gold-standard synthetic aperture imaging, on a dynamic range in excess of 60 dB. In vitro and in vivo experiments show that trainings carried out on simulated data perform well in experimental settings.

CNN-Based Ultrasound Image Reconstruction for Ultrafast Displacement Tracking.

Thanks to its capability of acquiring full-view frames at multiple kilohertz, ultrafast ultrasound imaging unlocked the analysis of rapidly changing physical phenomena in the human body, with pioneering applications such as ultrasensitive flow imaging in the cardiovascular system or shear-wave elastography. The accuracy achievable with these motion estimation techniques is strongly contingent upon two contradictory requirements: a high quality of consecutive frames and a high frame rate. Indeed, the image quality can usually be improved by increasing the number of steered ultrafast acquisitions, but at the expense of a reduced frame rate and possible motion artifacts. To achieve accurate motion estimation at uncompromised frame rates and immune to motion artifacts, the proposed approach relies on single ultrafast acquisitions to reconstruct high-quality frames and on only two consecutive frames to obtain 2-D displacement estimates. To this end, we deployed a convolutional neural network-based image reconstruction method combined with a speckle tracking algorithm based on cross-correlation. Numerical and in vivo experiments, conducted in the context of plane-wave imaging, demonstrate that the proposed approach is capable of estimating displacements in regions where the presence of side lobe and grating lobe artifacts prevents any displacement estimation with a state-of-the-art technique that relies on conventional delay-and-sum beamforming. The proposed approach may therefore unlock the full potential of ultrafast ultrasound, in applications such as ultrasensitive cardiovascular motion and flow analysis or shear-wave elastography.

Contrast-Enhanced Ultrasound Quantification: From Kinetic Modeling to Machine Learning.

Ultrasound contrast agents (UCAs) have opened up immense diagnostic possibilities by combined use of indicator dilution principles and dynamic contrast-enhanced ultrasound (DCE-US) imaging. UCAs are microbubbles encapsulated in a biocompatible shell. With a rheology comparable to that of red blood cells, UCAs provide an intravascular indicator for functional imaging of the (micro)vasculature by quantitative DCE-US. Several models of the UCA intravascular kinetics have been proposed to provide functional quantitative maps, aiding diagnosis of different pathological conditions. This article is a comprehensive review of the available methods for quantitative DCE-US imaging based on temporal, spatial and spatiotemporal analysis of the UCA kinetics. The recent introduction of novel UCAs that are targeted to specific vascular receptors has advanced DCE-US to a molecular imaging modality. In parallel, new kinetic models of increased complexity have been developed. The extraction of multiple quantitative maps, reflecting complementary variables of the underlying physiological processes, requires an integrative approach to their interpretation. A probabilistic framework based on emerging machine-learning methods represents nowadays the ultimate approach, improving the diagnostic accuracy of DCE-US imaging by optimal combination of the extracted complementary information. The current value and future perspective of all these advances are critically discussed.

Towards Ultrasound Everywhere: A Portable 3D Digital Back-End Capable of Zone and Compound Imaging.

Ultrasound imaging is a ubiquitous diagnostic technique, but does not fit the requirements of the telemedicine approach, because it relies on the real-time manipulation and image recognition skills of a trained expert, called sonographer. Sonographers are only available in hospitals and clinics, negating or at least delaying access to ultrasound scans in many locales-rural areas, developing countries-as well as in medical rescue operations. Telesonography would require an advanced imager that supports three-dimensional (3-D) acquisition; this would allow untrained operators to acquire broad scans and upload them remotely for diagnosis. Such advanced imagers do exist, but do not meet several other requirements for telesonography, such as being portable, inexpensive, and sufficiently low power to enable battery operation. In this work, we present our prototype of the first portable 3-D digital ultrasound back-end system. The prototype is implemented in a single midrange Xilinx field programmable gate array (FPGA), for an estimated power consumption of 5 W. The device supports up to 1024 input channels, which is state of the art and could be scaled further, and supports multiple image reconstruction modes. We evaluate the resource utilization of the FPGA and provide various quality metrics to ascertain the output image quality.

Ultrafast Ultrasound Imaging as an Inverse Problem: Matrix-Free Sparse Image Reconstruction.

Conventional ultrasound (US) image reconstruction methods rely on delay-and-sum (DAS) beamforming, which is a relatively poor solution to the image reconstruction problem. An alternative to DAS consists in using iterative techniques, which require both an accurate measurement model and a strong prior on the image under scrutiny. Toward this goal, much effort has been deployed in formulating models for US imaging, which usually require a large amount of memory to store the matrix coefficients. We present two different techniques, which take advantage of fast and matrix-free formulations derived for the measurement model and its adjoint, and rely on sparsity of US images in well-chosen models. Sparse regularization is used for enhanced image reconstruction. Compressed beamforming exploits the compressed sensing framework to restore high-quality images from fewer raw data than state-of-the-art approaches. Using simulated data and in vivo experimental acquisitions, we show that the proposed approach is three orders of magnitude faster than non-DAS state-of-the-art methods, with comparable or better image quality.

Efficient Sample Delay Calculation for 2-D and 3-D Ultrasound Imaging.

Ultrasound imaging is a reference medical diagnostic technique, thanks to its blend of versatility, effectiveness, and moderate cost. The core computation of all ultrasound imaging methods is based on simple formulae, except for those required to calculate acoustic propagation delays with high precision and throughput. Unfortunately, advanced three-dimensional (3-D) systems require the calculation or storage of billions of such delay values per frame, which is a challenge. In 2-D systems, this requirement can be four orders of magnitude lower, but efficient computation is still crucial in view of low-power implementations that can be battery-operated, enabling usage in numerous additional scenarios. In this paper, we explore two smart designs of the delay generation function. To quantify their hardware cost, we implement them on FPGA and study their footprint and performance. We evaluate how these architectures scale to different ultrasound applications, from a low-power 2-D system to a next-generation 3-D machine. When using numerical approximations, we demonstrate the ability to generate delay values with sufficient throughput to support 10 000-channel 3-D imaging at up to 30 fps while using 63% of a Virtex 7 FPGA, requiring 24 MB of external memory accessed at about 32 GB/s bandwidth. Alternatively, with similar FPGA occupation, we show an exact calculation method that reaches 24 fps on 1225-channel 3-D imaging and does not require external memory at all. Both designs can be scaled to use a negligible amount of resources for 2-D imaging in low-power applications and for ultrafast 2-D imaging at hundreds of frames per second.

Echo-power estimation from log-compressed video data in dynamic contrast-enhanced ultrasound imaging.

Ultrasound (US) scanners typically apply lossy, non-linear modifications to the US data for visualization purposes. The resulting images are then stored as compressed video data. Some system manufacturers provide dedicated software for quantification purposes to eliminate such processing distortions, at least partially. This is currently the recommended approach for quantitatively assessing changes in contrast-agent concentration from clinical data. However, the machine-specific access to US data and the limited set of analysis functionalities offered by each dedicated-software package make it difficult to perform comparable analyses with different US systems. The objective of this work was to establish if linearization of compressed video images obtained with an arbitrary US system can provide an alternative to dedicated-software analysis of machine-specific files for the estimation of echo-power. For this purpose, an Aplio 50 system (Toshiba Medical Systems, Tochigi, Japan), coupled with dedicated CHI-Q (Contrast Harmonic Imaging Quantification) software by Toshiba Medical Systems, was used. Results were compared with two approaches that apply algorithms to estimate relative echo-power from compressed video images: commercially available VueBox software by Bracco Suisse SA (Geneva, Switzerland) and in-laboratory software called PixPower. The echo-power estimated by CHI-Q analysis indicated a strong linear relationship versus agent concentration in vitro (R(2) ≥ 0.9996) for dynamic range (DR) settings of DR60 and DR80, with slopes between 9.22 and 9.57 dB/decade (p = 0.05). These values approach the theoretically predicted dependence of 10.0 dB/decade (equivalent to 3 dB for each concentration doubling). Echo-power estimations obtained from compressed video images with VueBox and PixPower also exhibited strong linear proportionality with concentration (R(2) ≥ 0.9996), with slopes between 9.30 and 9.68 dB/decade (p = 0.05). On an independent in vivo data set (N = 24), the difference in echo-power estimation between CHI-Q and each of the other two approaches was calculated after excluding regions that contain pixels affected by saturated or thresholded pixel values. The mean difference in estimates (expressed in decibels) was -0.25 dB between VueBox and CHI-Q (95% confidence interval: -0.75 to 0.26 dB) and -0.17 dB between PixPower and CHI-Q (95% confidence interval: -0.67 to 0.13 dB). To achieve linearization of data, one of the approaches (VueBox) requires calibration files provided by the software manufacturer for each machine type and setting. The other (PixPower) requires empirical correction of the imaging dynamic range based on ground truth data. These requirements could potentially be removed if US system manufacturers were willing to make relevant information on the applied processing publically available. Reliable echo-power estimation from linearized data would facilitate inclusion of different US systems in multicentric studies and more widespread implementation of emerging techniques for quantitative analysis of contrast ultrasound.

In vitro sonothrombolysis of human blood clots with BR38 microbubbles.

Microbubble-mediated sonothrombolysis is a promising approach for ischemic stroke treatment. The aim of this in vitro study was to evaluate a new microbubble (MB) formulation (BR38) for sonothrombolysis and to investigate the involved mechanisms. Human whole-blood clots were exposed to different combinations of recombinant tissue plasminogen activator (rtPA), ultrasound (US) and MB. Ultrasound at 1.6 MHz was used at 150, 300, 600 and 1000 kPa (peak-negative pressure). Thrombolysis efficacy was assessed by measuring clot diameter changes during 60-min US exposure. The rate of clot diameter loss (RDL) in µm/min was determined and clot lysis profiles were analyzed. The most efficient clot lysis (5.9 µm/min) was obtained at acoustic pressures of 600 and 1000 kPa in combination with MB and a low concentration of rtPA (0.3 µg/mL). This is comparable with the rate obtained with rtPA at 3 µg/mL alone (6.6 µm/min, p > 0.05). Clot lysis profiles were shown to be related to US beam profiles and microbubble cavitation.

Effects of acoustic radiation force on the binding efficiency of BR55, a VEGFR2-specific ultrasound contrast agent.

This work describes an in vivo study analyzing the effect of acoustic radiation force (ARF) on the binding of BR55 VEGFR2-specific contrast-agent microbubbles in a model of prostatic adenocarcinoma in rat. A commercial ultrasound system was modified by implementing high duty-cycle 3.5-MHz center frequency ARF bursts in a scanning configuration. This enabled comparing the effects of ARF on binding in tumor and healthy tissue effectively in the same field of view. Bubble binding was established by measuring late-phase enhancement in amplitude modulation (AM) contrast-specific imaging mode (4 MHz, 150 kPa) 10 min after agent injection when the unbound bubbles were cleared from the circulation. Optimal experimental conditions, such as agent concentration (0.4 × 10(8)-1.6 × 10(8) bubbles/kg), acoustic pressure amplitude (26-51 kPa) and duty-cycle (20%-95%) of the ARF bursts, were evaluated in their ability to enhance binding in tumor without significantly increasing binding in healthy tissue. Using the optimal conditions (38 kPa peak-negative pressure, 95% duty cycle), ARF-assisted binding of BR55 improved significantly in tumor (by a factor of 7) at a lower agent dose compared with binding without ARF, and it had an insignificant effect on binding in healthy tissue. Thus, the high binding specificity of BR55 microbubbles for targeting VEGFR2 present at sites of active angiogenesis was confirmed by this study. Therefore, it is believed that based on the results obtained in this work, ultrasound molecular imaging using target-specific contrast-agent microbubbles should preferably be performed in combination with ARF.

Dynamic contrast enhanced ultrasound assessment of the vascular effects of novel therapeutics in early stage trials.

Imaging is key in the accurate monitoring of response to cancer therapies targeting tumour vascularity to inhibit its growth and dissemination. Dynamic contrast enhanced ultrasound (DCE ultrasound) is a quantitative method with the advantage of being non-invasive, widely available, portable, cost effective, highly sensitive and reproducible using agents that are truly intravascular. Under the auspices of the initiative of the Experimental Cancer Medicine Centre Imaging Network, bringing together experts from the UK, Europe and North America for a 2-day workshop in May 2010, this consensus paper aims to provide guidance on the use of DCE ultrasound in the measurement of tumour vascular support in clinical trials. Key Points • DCE ultrasound can quantify and extract specific blood flow parameters, such as flow velocity, relative vascular volume and relative blood flow rate. • DCE ultrasound can be performed repeatedly and is therefore ideally suited for pharmacokinetic and pharmacodynamic studies evaluating vascular-targeted drugs. • DCE ultrasound provides a reproducible method of assessing the vascular effects of therapy in pre-clinical and early clinical trials, which is easily translatable into routine clinical practice.

Differentiation of focal liver lesions: usefulness of parametric imaging with contrast-enhanced US.

PURPOSE: To evaluate whether parametric imaging with contrast material-enhanced ultrasonography (US) is superior to visual assessment for the differential diagnosis of focal liver lesions (FLLs). MATERIALS AND METHODS: This study had institutional review board approval, and verbal patient informed consent was obtained. Between August 2005 and October 2008, 146 FLLs in 145 patients (63 women, 82 men; mean age, 62.5 years; age range, 22-89 years) were imaged with real-time low-mechanical-index contrast-enhanced US after a bolus injection of 2.4 mL of a second-generation contrast agent. Clips showing contrast agent uptake kinetics (including arterial, portal, and late phases) were recorded and subsequently analyzed off-line with dedicated image processing software. Analysis of the dynamic vascular patterns (DVPs) of lesions with respect to adjacent parenchyma allowed mapping DVP signatures on a single parametric image. Cine loops of contrast-enhanced US and results from parametric imaging of DVP were assessed separately by three independent off-site readers who classified each lesion as benign, malignant, or indeterminate. Sensitivity, specificity, accuracy, and positive and negative predictive values were calculated for both techniques. Interobserver agreement (κ statistics) was determined. RESULTS: Sensitivities for visual interpretation of cine loops for the three readers were 85.0%, 77.9%, and 87.6%, which improved significantly to 96.5%, 97.3%, and 96.5% for parametric imaging, respectively (P < .05, McNemar test), while retaining high specificity (90.9% for all three readers). Accuracy scores of parametric imaging were higher than those of conventional contrast-enhanced US for all three readers (P < .001, McNemar test). Interobserver agreement increased with DVP parametric imaging compared with conventional contrast-enhanced US (change of κ from 0.54 to 0.99). CONCLUSION: Parametric imaging of DVP improves diagnostic performance of contrast-enhanced US in the differentiation between malignant and benign FLLs; it also provides excellent interobserver agreement.

BR38, a new ultrasound blood pool agent.

OBJECTIVE: : To evaluate BR38, a new microbubble-based blood pool agent for contrast-enhanced ultrasound imaging. MATERIALS AND METHODS: : The size characteristics of BR38 microbubbles were measured by Coulter counting. The backscatter and attenuation coefficients were determined as a function of frequency. Additional measurements included the surface charge, osmolality, viscosity, and resistance to hydrostatic pressure. Extensive pharmacological and toxicological studies were conducted on the final formulation in rats and dogs. The blood levels and elimination of the gaseous component C4F10 were determined in the rabbit. Contrast-enhanced echographic examinations were performed in pigs focusing on the myocardium and the liver. Finally, safety testing and preliminary imaging experiments were performed in a Phase I clinical study in human volunteers. RESULTS: : BR38 suspensions are isotonic, nonviscous, and show a high resistance to hydrostatic pressure. Their backscatter coefficient is high at ≥ 2 MHz and attenuation shows a maximum at 4 MHz, slowly decreasing at higher frequencies. The no adverse effect levels of 1 µL/kg (rats) and 5 µL/kg (dogs) expressed as microbubble gas volume, observed in repeated toxicology studies, correspond to 50 and 250 times the expected imaging dose in human beings (0.02 µL/kg), respectively. No effects on cardiovascular and respiratory parameters were observed in rats and dogs. C4F10 is eliminated within minutes from blood and excreted in expired air. Imaging experiments showed strong and persistent enhancement of the myocardium and the liver. A late phase was observed in the liver, in animals and in human volunteers. No serious adverse events and no significant changes in vital signs, electrocardiographs, and laboratory tests were observed in Phase I human volunteers. CONCLUSIONS: : BR38 shows a very good safety profile. It is characterized by a long persistence and low shadowing. BR38 is a promising ultrasound blood pool agent for noncardiac and cardiac applications including myocardial perfusion imaging.

Parametric imaging for characterizing focal liver lesions in contrast-enhanced ultrasound.

The differentiation between benign and malignant focal liver lesions plays an important role in diagnosis of liver disease and therapeutic planning of local or general disease. This differentiation, based on characterization, relies on the observation of the dynamic vascular patterns (DVP) of lesions with respect to adjacent parenchyma, and may be assessed during contrast-enhanced ultrasound imaging after a bolus injection. For instance, hemangiomas (i.e., benign lesions) exhibit hyper-enhanced signatures over time, whereas metastases (i.e., malignant lesions) frequently present hyperenhanced foci during the arterial phase and always become hypo-enhanced afterwards. The objective of this work was to develop a new parametric imaging technique, aimed at mapping the DVP signatures into a single image called a DVP parametric image, conceived as a diagnostic aid tool for characterizing lesion types. The methodology consisted in processing a time sequence of images (DICOM video data) using four consecutive steps: (1) pre-processing combining image motion correction and linearization to derive an echo-power signal, in each pixel, proportional to local contrast agent concentration over time; (2) signal modeling, by means of a curve-fitting optimization, to compute a difference signal in each pixel, as the subtraction of adjacent parenchyma kinetic from the echopower signal; (3) classification of difference signals; and (4) parametric image rendering to represent classified pixels as a support for diagnosis. DVP parametric imaging was the object of a clinical assessment on a total of 146 lesions, imaged using different medical ultrasound systems. The resulting sensitivity and specificity were 97% and 91%, respectively, which compare favorably with scores of 81 to 95% and 80 to 95% reported in medical literature for sensitivity and specificity, respectively.

Subharmonic scattering of phospholipid-shell microbubbles at low acoustic pressure amplitudes.

Subharmonic scattering of phospholipid-shell microbubbles excited at relatively low acoustic pressure amplitudes (<30 kPa) has been associated with echo responses from compression-only bubbles having initial surface tension values close to zero. In this work, the relation between sbharmonics and compression-only behavior of phospholipid-shell microbubbles was investigated, experimentally and by simulation, as a function of the initial surface tension by applying ambient overpressures of 0 and 180 mmHg. The microbubbles were excited using a 64-cycle transmit burst with a center frequency of 4 MHz and peak-negative pressure amplitudes ranging from 20 of 150 kPa. In these conditions, an increase in subharmonic response of 28.9 dB (P < 0.05) was measured at 50 kPa after applying an overpressure of 180 mmHg. Simulations using the Marmottant model, taking into account the effect of ambient overpressure on bubble size and initial surface tension, confirmed the relation between subharmonics observed in the pressure-time curves and compression-only behavior observed in the radius-time curves. The trend of an increase in subharmonic response as a function of ambient overpressure, i.e., as a function of the initial surface tension, was predicted by the model. Subharmonics present in the echo responses of phospholipid-shell microbubbles excited at low acoustic pressure amplitudes are indeed related to the echo responses from compression-only bubbles. The increase in subharmonics as a function of ambient overpressure may be exploited for improving methods for noninvasive pressure measurement in heart cavities or big vessels in the human body.

Acoustic characterization of single ultrasound contrast agent microbubbles.

Individual ultrasound contrast agent microbubbles (BR14) were characterized acoustically. The bubbles were excited at a frequency of 2 MHz and at peak-negative pressure amplitudes of 60 and 100 kPa. By measuring the transmit and receive transfer functions of both the transmit and receive transducers, echoes of individual bubbles were recorded quantitatively and compared to simulated data. At 100 kPa driving pressure, a second harmonic response was observed for bubbles with a size close to their resonance size. Power spectra were derived from the echo waveforms of bubbles of different sizes. These spectra were in good agreement with those calculated from a Rayleigh-Plesset-type model, incorporating the viscoelastic properties of the phospholipid shell. Small bubbles excited below their resonance frequency have a response dominated by the characteristics of their phospholipid shell, whereas larger bubbles, excited above resonance, have a response identical to those of uncoated bubbles of similar size.

Effects of simultaneous application of ultrasound and microbubbles on intracerebral hemorrhage in an animal model.

Microbubble-enhanced sonothrombolysis (MEST) may be an alternative therapeutic option in ischemic stroke. Clinical study of the efficacy of MEST as an adjunct stroke therapy, before imaging with CT or MRI, requires experimental data on the safety of this approach in the presence of hemorrhagic stroke. We, therefore, investigated the effect of diagnostic transcranial ultrasound combined with microbubbles (US + MB) in an experimental animal model of intracerebral hemorrhage (ICH). ICH was induced in anesthetized rats by intracerebral collagenase injection. Transcranial ultrasound (2 MHz, mechanical index 1.3, 1051 kPa) was applied 3 h after ICH induction to rat brains for 30 min during a continuous IV infusion of sulfur hexafluoride microbubbles (SonoVue). The size of cerebral hemorrhage, the extent of brain edema, and the amount of apoptosis were compared with those from control rats with ICH but without US + MB. Results showed no significant effect of US + MB on hemorrhage size (control 23.3 +/- 10.7 mm(3), US + MB 20.3 +/- 5.8 mm(3)), on the extent of brain edema (control 3.3 +/- 2.0%, US +MB 3.5 +/- 1.9%), or on the rate of apoptosis (control 5.2 +/- 1.5%, US + MB 5.2 +/- 1.0%). We conclude that diagnostic ultrasound in combination with microbubbles does not cause additional damage to the rat brain during ICH in our experimental set-up. This finding provides support for the use of MEST as an early stroke therapy.

A new formalism for the quantification of tissue perfusion by the destruction-replenishment method in contrast ultrasound imaging.

A new formalism is presented for the destruction-replenishment perfusion quantification approach at low mechanical index. On the basis of physical considerations, best-fit methods should be applied using perfusion functions with S-shape characteristics. These functions are first described for the case of a geometry with a single flow velocity, then extended to the case of vascular beds with blood vessels having multiple flow velocity values and directions. The principles guiding the analysis are, on one hand, a linearization of video echo signals to overcome the log-compression of the imaging instrument, and, on the other hand, the spatial distribution of the transmit-receive ultrasound beam in the elevation direction. An in vitro model also is described; it was used to confirm experimentally the validity of the approach using a commercial contrast agent. The approach was implemented in the form of a computer program, taking as input a sequence of contrast-specific images, as well as parameters related to the ultrasound imaging equipment used. The generated output is either flow-parameter values computed in regions-of-interest, or parametric flow-images (e.g., mean velocity, mean transit time, mean flow, flow variance, or skewness). This approach thus establishes a base for extracting information about the morphology of vascular beds in vivo, and could allow absolute quantification provided that appropriate instrument calibration is implemented.

Different effects of microbubble destruction and translation in Doppler measurements.

In flow measurements in which microbubbles are involved, the amplitude and phase of the received echo signal are noticeably influenced by the transmitted ultrasound intensity. Previous studies have shown that, when such intensity is progressively increased, the Doppler spectrum is accordingly distorted, i.e., it is asymmetrically broadened toward the negative frequency side. Such deformation has been attributed to radiation force, which pushes the microbubbles into the sound propagation direction, thus yielding additional phase delays in the received echoes. However, the possible contribution of microbubble destruction to this spectral deformation has not been considered yet. In this paper, this issue is investigated by analyzing the experimental spectra produced by two different types of microbubbles suspended in a moving fluid and insonified in pulsed wave (PW) mode at programmable pulse repetition frequency (PRF) and pressure. Conditions are created in which either the radiation force or the destruction mechanism is expected to be dominant. Effects produced by the two phenomena on the Doppler spectrum are shown to be different. When the PRF is low (2 kHz), so that, according to theoretical simulations, the radiation force effect is negligible, a 26 dB noise floor increase is observed for a 13 dB pressure increment. For a higher PRF (16 kHz), the same pressure increase not only affects the noise floor, but also causes the bubbles to deviate from their original streamlines, yielding a Doppler bandwidth increase by a factor of 5. It is concluded that asymmetrical spectral broadening is mainly due to radiation force, and microbubble destruction mainly results in an increased noise floor without affecting the spectral shape.