Nano-sized particles are widely regarded as a tool to study biologic events at the cellular and molecular levels. However, only some imaging modalities can visualize interaction between nanoparticles and living cells. We present a new technique, pulsed magneto-motive ultrasound imaging, which is capable of in vivo imaging of magnetic nanoparticles in real time and at sufficient depth. In pulsed magneto-motive ultrasound imaging, an external high-strength pulsed magnetic field is applied to induce the motion within the magnetically labeled tissue and ultrasound is used to detect the induced internal tissue motion. Our experiments demonstrated a sufficient contrast between normal and iron-laden cells labeled with ultrasmall magnetic nanoparticles. Therefore, pulsed magneto-motive ultrasound imaging could become an imaging tool capable of detecting magnetic nanoparticles and characterizing the cellular and molecular composition of deep-lying structures.
Background and Objectives
We report on a study to investigate feasibility of utilizing ultrasound imaging to guide laser removal of subcutaneous fat. Ultrasound imaging can be used to identify the tissue composition and to monitor the temperature increase in response to laser irradiation.
Study Design/Materials and Methods
Laser heating was performed on ex vivo porcine subcutaneous fat through the overlying skin using a continuous wave laser operating at 1,210 nm optical wavelength. Ultrasound images were recorded using a 10 MHz linear array-based ultrasound imaging system.
Ultrasound imaging was utilized to differentiate between water-based and lipid-based regions within the porcine tissue and to identify the dermis-fat junction. Temperature maps during the laser exposure in the skin and fatty tissue layers were computed.
Results of our study demonstrate the potential of using ultrasound imaging to guide laser fat removal.
laser therapy; ultrasound imaging; thermal imaging; treatment monitoring; fat removal; body reshaping
Evaluating the regenerative capacity of a tissue-engineered device in a noninvasive and synchronous manner is critical to determining the mechanisms for success in clinical applications. In particular, directly tracking implanted cells in a three-dimensional (3D) scaffold is desirable in that it enables the monitoring of cellular activity in a specific and localized manner. The authors’ group has previously demonstrated that the PEGylation of fibrin results in a 3D scaffold that supports morphologic and phenotypic changes in mesenchymal stem cells that may be advantageous in wound healing applications. Recently, the authors have evaluated adipose-derived stem cells (ASCs) as a mesenchymal cell source to regenerate skin and blood vessels due to their potential for proliferation, differentiation, and production of growth factors. However, tracking and monitoring ASCs in a 3D scaffold, such as a PEGylated fibrin gel, have not yet been fully investigated. In the current paper, nanoscale gold spheres (20 nm) as cell tracers for ASCs cultured in a PEGylated fibrin gel were evaluated. An advanced dual-imaging modality combining ultrasound and photoacoustic imaging was utilized to monitor rat ASCs over time. The ASCs took up gold nanotracers and could be detected up to day 16 with high sensitivity using photoacoustic imaging. There were no detrimental effects on ASC morphology, network formation, proliferation, and protein expression/secretion (ie, smooth muscle α-actin, vascular endothelial growth factor, matrix metalloproteinase-2, and matrix metalloproteinase-9) associated with gold nanotracers. Therefore, utilization of gold nanotracers can be an effective strategy to monitor the regenerative process of a stem cell source in a 3D gel for vascular and dermal tissue engineering applications.
gold nanoparticles; adipose-derived stem cells; fibrin; ultrasound and photoacoustic imaging; angiogenesis; tissue engineering
As applications of nanoparticles in medical imaging and biomedicine rapidly expand, the interactions of nanoparticles with living cells have become an area of active interest. For example, intracellular trafficking of nanoparticles – an important part of cell-nanoparticle interaction, has been well studied using plasmonic nanoparticles and optical or optics-based techniques due to the change in optical properties of the nanoparticle aggregates. However, magnetic nanoparticles, despite their wide range of clinical applications, do not exhibit plasmonic-resonant properties and therefore their intracellular aggregation cannot be detected by optics-based imaging techniques. In this study, we investigated the feasibility of a novel imaging technique – pulsed magneto-motive ultrasound (pMMUS), to identify intracellular trafficking of endocytosed magnetic nanoparticles. In pulsed magneto-motive ultrasound imaging a focused, high intensity, pulsed magnetic field is used to excite the cells labeled with magnetic nanoparticles, and ultrasound imaging is then used to monitor the mechanical response of the tissue. We demonstrated previously that clusters of magnetic nanoparticles amplify the pMMUS signal in comparison to signal from individual nanoparticles. Here we further demonstrate that pMMUS imaging can identify interaction between magnetic nanoparticles and living cells, i.e. intracellular aggregation of nanoparticles within the cells. The results of our study suggest that pMMUS imaging can not only detect the presence of magnetic nanoparticles but also provides information about their intracellular trafficking non-invasively and in real-time.
Pulsed magneto-motive ultrasound imaging; superparamagnetic iron-oxide nanoparticles; macrophage; endocytosis; intracellular trafficking
Longitudinal monitoring of cells is required in order to understand the role of delivered stem cells in therapeutic neovascularization. However, there is not an imaging technique that is capable of quantitative, longitudinal assessment of stem cell behaviors with high spatial resolution and sufficient penetration depth. In this study, in vivo and in vitro experiments were performed to demonstrate the efficacy of ultrasound-guided photoacoustic (US/PA) imaging to monitor mesenchymal stem cells (MSCs) labeled with gold nanotracers (Au NTs). The Au NT labeled MSCs, injected intramuscularly in the lower limb of the Lewis rat, were detected and spatially resolved. Furthermore, our quantitative in vitro cell studies indicate that US/PA imaging is capable of high detection sensitivity (1×104 cells/mL) of the Au NT labeled MSCs. Finally, Au NT labeled MSCs captured in the PEGylated fibrin gel system were imaged in vivo, as well as in vitro, over a one week time period, suggesting that longitudinal cell tracking using US/PA imaging is possible. Overall, Au NT labeling of MSCs and US/PA imaging can be an alternative approach in stem cell imaging capable of noninvasive, sensitive, quantitative, longitudinal assessment of stem cell behaviors with high spatial and temporal resolutions at sufficient depths.
Importance of the field
Photoacoustic imaging is an imaging modality that derives image contrast from the optical absorption coefficient of the tissue being imaged. The imaging technique is able to differentiate between healthy and diseased tissue with either deeper penetration or higher resolution than other functional imaging modalities currently available. From a clinical standpoint, photoacoustic imaging has demonstrated safety and effectiveness in diagnosing diseased tissue regions using either endogenous tissue contrast or exogenous contrast agents. Furthermore, the potential of photoacoustic imaging has been demonstrated in various therapeutic interventions ranging from drug delivery and release to image-guided therapy and monitoring.
Areas covered in this review
This article reviews the current state of photoacoustic imaging in biomedicine from a technological perspective, highlights various biomedical and clinical applications of photoacoustic imaging, and gives insights on future directions.
What the reader will gain
Readers will learn about the various applications of photoacoustic imaging, as well as the various contrast agents that can be used to assist photoacoustic imaging. This review will highlight both pre-clinical and clinical uses for photoacoustic imaging, as well as discuss some of the challenges that must be addressed to move photoacoustic imaging into the clinical realm.
Take home message
Photoacoustic imaging offers unique advantages over existing imaging modalities. The imaging field is broad with many exciting applications for detecting and diagnosing diseased tissue or processes. Photoacoustics is also used in therapeutic applications to identify and characterize the pathology and then to monitor the treatment. Although the technology is still in its infancy, much work has been done in the pre-clinical arena, and photoacoustic imaging is fast approaching the clinical setting.
Atherosclerosis; cancer; contrast agents; drug delivery; molecular imaging; nanoparticles; optoacoustics; photoacoustics; therapy; ultrasound
In both photoacoustic (PA) and ultrasonic (US) imaging, overall image quality is influenced by the optical and acoustical properties of the medium. Consequently, with the increased use of combined PA and US (PAUS) imaging in preclinical and clinical applications, the ability to provide phantoms that are capable of mimicking desired properties of soft tissues is critical. To this end, gelatin-based phantoms were constructed with various additives to provide realistic acoustic and optical properties. Forty-micron, spherical silica particles were used to induce acoustic scattering, Intralipid® 20% IV fat emulsion was employed to enhance optical scattering and ultrasonic attenuation, while India Ink, Direct Red 81, and Evans blue dyes were utilized to achieve optical absorption typical of soft tissues. The following parameters were then measured in each phantom formulation: speed of sound, acoustic attenuation (from 6 to 22 MHz), acoustic backscatter coefficient (from 6 to 22 MHz), optical absorption (from 400 nm to 1300 nm), and optical scattering (from 400 nm to 1300 nm). Results from these measurements were then compared to similar measurements, which are offered by the literature, for various soft tissue types. Based on these comparisons, it was shown that a reasonably accurate tissue-mimicking phantom could be constructed using a gelatin base with the aforementioned additives. Thus, it is possible to construct a phantom that mimics specific tissue acoustical and/or optical properties for the purpose of PAUS imaging studies.
(110.3000) Image quality assessment; (160.4760) Optical properties; (170.5120) Photoacoustic imaging; (170.7170) Ultrasound; (290.5820) Scattering measurements
In vivo monitoring of nanoparticle delivery is essential to better understand cellular and molecular interactions of nanoparticles with tissue and to better plan nanoparticle-mediated therapies. We developed a three-dimensional ultrasound and photoacoustic (PA) imaging system and a spectroscopic PA imaging algorithm to identify and quantify the presence of nanoparticles and other tissue constituents. Using the developed system and approach, three-dimensional in vivo imaging of a mouse with tumor was performed before and after intravenous injection of gold nanorods. The developed spectroscopic PA imaging algorithm estimated distribution of nanoparticle as well as oxygen saturation of blood. Moreover, silver staining of excised tumor tissue confirmed nanoparticle deposition, and showed good correlation with spectroscopic PA images. The results of our study suggest that three-dimensional ultrasound-guided spectroscopic PA imaging can monitor nanoparticle delivery in vivo.
(170.5120) Photoacoustic imaging; (110.7170) Ultrasound; (170.3880) Medical and biological imaging
Brachytherapy seed therapy is an increasingly common way to treat prostate cancer through localized radiation. The current standard of care relies on transrectal ultrasound (TRUS) for imaging guidance during the seed placement procedure. As visualization of individual metallic seeds tends to be difficult or inaccurate under TRUS guidance, guide needles are generally tracked to infer seed placement. In an effort to improve seed visualization and placement accuracy, the use of photoacoustic (PA) imaging, which is highly sensitive to metallic objects in soft tissue, was investigated for this clinical application. The PA imaging properties of bare (i.e., embedded in pure gelatin) and tissue-embedded (at depths of up to 13 mm) seeds were investigated with a multi-wavelength (750 to 1090 nm) PA imaging technique. Results indicate that, much like ultrasonic (US) imaging, an angular dependence (i.e., seed orientation relative to imaging transducer) of the PA signal exists. Despite this shortcoming, however, PA imaging offers improved contrast, over US imaging, of a seed in prostate tissue if sufficient local fluence is achieved. Additionally, although the PA signal of a bare seed is greatest for lower laser wavelengths (e.g., 750 nm), the scattering that results from tissue tends to favor the use of higher wavelengths (e.g., 1064 nm, which is the primary wavelength of Nd:YAG lasers) when the seed is located in tissue. A combined PA and US imaging approach (i.e., PAUS imaging) shows strong potential to visualize both the seed and the surrounding anatomical environment of the prostate during brachytherapy seed placement procedures.
(170.5120) Photoacoustic imaging; (170.7170) Ultrasound; (170.1610) Clinical applications; (170.7230) Urology; (170.3660) Light propagation in tissues; (110.0113) Imaging through turbid media
In ultrasound strain and elasticity imaging, an accurate and cost-effective sub-pixel displacement estimator is required because strain/elasticity imaging quality relies on the displacement SNR, which can often be higher if more computational resources are provided. In this paper, we introduce an autocorrelation-based method to cost-effectively improve sub-pixel displacement estimation quality. To quantitatively evaluate the performance of the autocorrelation method, simulated and tissue-mimicking phantom experiments were performed. The computational cost of the autocorrelation method is also discussed. The results of our study suggest the autocorrelation method can be used for a real-time elasticity imaging system.
Plasmonic metal nanoparticles are used in photoacoustic imaging as contrast agents because of their resonant optical absorption properties in the visible and near-IR regions. However, the nanoparticles could accumulate and result in long-term toxicity in vivo, because they are generally not biodegradable. Recently, biodegradable plasmonic gold nanoclusters, consisting of sub-5 nm primary gold nanoparticles and biodegradable polymer stabilizer, were introduced. In this Letter, we demonstrate the feasibility of biodegradable nanoclusters as a photoacoustic contrast agent. We performed photoacoustic and ultrasound imaging of a tissue-mimicking phantom with inclusions containing nanoclusters at various concentrations. The results indicate that the biodegradable gold nanoclusters can be used as effective contrast agents in photoacoustic imaging.
An ultrasound-based method to locally assess the shear modulus of a medium is reported. The proposed approach is based on the application of an impulse acoustic radiation force to an inhomogeneity in the medium and subsequent monitoring of the spatio-temporal response. In our experimental studies, a short pulse produced by a 1.5-MHz highly focused ultrasound transducer was used to initiate the motion of a rigid sphere embedded into an elastic medium. Another 25 MHz focused ultrasound transducer operating in pulse-echo mode was used to track the displacement of the sphere. The experiments were performed in gel phantoms with varying shear modulus to demonstrate the relationship between the displacement of the sphere and shear modulus of the surrounding medium. Because the magnitude of acoustic force applied to sphere depends on the acoustic material properties and, therefore, cannot be used to assess the absolute value of shear modulus, the temporal behavior of the displacement of the sphere was analyzed. The results of this study indicate that there is a strong correlation between the shear modulus of a medium and spatio-temporal characteristics of the motion of the rigid sphere embedded in this medium.
Stem cells can differentiate into multiple cell types, and therefore can be used for cellular therapies, including tissue repair. However, the participation of stem cells in tissue repair and neovascularization is not well understood. Therefore, implementing a noninvasive, long-term imaging technique to track stem cells in vivo is needed to obtain a better understanding of the wound healing response. Generally, we are interested in developing an imaging approach to track mesenchymal stem cells (MSCs) in vivo after delivery via a polyethylene glycol modified fibrin matrix (PEGylated fibrin matrix) using MSCs loaded with gold nanoparticles as nanotracers. The objective of the current study was to assess the effects of loading MSCs with gold nanoparticles on cellular function.
In this study, we utilized various gold nanoparticle formulations by varying size and surface coatings and assessed the efficiency of cell labeling using darkfield microscopy. We hypothesized that loading cells with gold nanotracers would not significantly alter cell function due to the inert and biocompatible characteristics of gold. The effect of nanoparticle loading on cell viability and cytotoxicity was analyzed using a LIVE/DEAD stain and an MTT assay. The ability of MSCs to differentiate into adipocytes and osteocytes after nanoparticle loading was also examined. In addition, nanoparticle loading and retention over time was assessed using inductively coupled plasma mass spectrometry (ICP-MS).
Our results demonstrate that loading MSCs with gold nanotracers does not alter cell function and, based on the ICP-MS results, long-term imaging and tracking of MSCs is feasible. These findings strengthen the possibility of imaging MSCs in vivo, such as with optical or photoacoustic imaging, to understand better the participation and role of MSCs in neovascularization.
neovascularization; gold nanoparticles; cell tracking; stem cells; fibrin gel; optical imaging
Sound waves generated by light are the basis of a sensitive medical imaging technique with applications to cancer diagnosis and treatment.
An adaptive photoacoustic image reconstruction technique that combines coherence factor (CF) weighting and the minimum variance (MV) method is introduced. The backprojection method is widely used to reconstruct photoacoustic tomography images. Owing to the scattering of light, the quality of the photoacoustic imaging can be degraded. CF, an adaptive weighting technique, is known to improve the lateral resolution of photoacoustic images. In addition, an MV adaptive beamforming method can further improve the image quality by suppressing signals from off-axis directions. Experimental studies are performed to quantify the spatial resolution and contrast of the adaptive photoacoustic beamforming methods.
A recently developed ultrasound technique is evaluated by measuring the behavior of a cavitation bubble that is induced in water by a femtosecond laser pulse. The passive acoustic emission during optical breakdown is used to estimate the location of the cavitation bubble’s origin. In turn, the position of the bubble wall is defined based on the active ultrasonic pulse-echo signal. The results suggest that the developed ultrasound technique can be used for quantitative measurements of femtosecond laser-induced microbubbles.
An ultrasound technique to measure the spatial and temporal behavior of the laser-induced cavitation bubble is introduced. The cavitation bubbles were formed in water and in gels using a nanosecond pulsed Nd:YAG laser operating at 532 nm. A focused, single-element, 25-MHz ultrasound transducer was employed both to detect the acoustic emission generated by plasma expansion and to acoustically probe the bubble at different stages of its evolution. The arrival time of the passive acoustic emission was used to estimate the location of the cavitation bubble’s origin and the time of flight of the ultrasound pulse-echo signal was used to define its spatial extent. The results of ultrasound estimations of the bubble size were compared and found to be in agreement with both the direct optical measurements of the stationary bubble and the theoretical estimates of bubble dynamics derived from the well-known Rayleigh model of a cavity collapse. The results of this study indicate that the proposed quantitative ultrasound technique, capable of detecting and accurately measuring laser-induced cavitation bubbles in water and in a tissue-like medium, could be used in various biomedical and clinical applications.
ultrasonics; shock; emission; laser-induced damage; laser application; lasers in medicine
Intravascular photoacoustic (IVPA) imaging is based on the detection of laser-induced acoustic waves generated within the arterial tissue under pulsed laser irradiation. Generally, laser radiant energy levels are kept low (20 mJ/cm2) during photoacoustic imaging to conform to general standards for safe use of lasers on biological tissues. However, safety standards in intravascular photoacoustic imaging are not yet fully established. Consequently, monitoring spatio-temporal temperature changes associated with laser-tissue interaction is important to address thermal safety of IVPA imaging. In this study we utilize the IVUS based strain measurements to estimate the laser induced temperature increase. Temporal changes in temperature were estimated in a phantom modeling a vessel with an inclusion. A cross-correlation based time delay estimator was used to assess temperature induced strains produced by different laser radiant energies. The IVUS based remote measurements revealed temperature increases of 0.7±0.3°C, 2.9±0.2 °C and 5.0±0.2 °C, for the laser radiant energies of 30 mJ/cm2, 60 mJ/cm2 and 85 mJ/cm2 respectively. The technique was then used in imaging of ex vivo samples of a normal rabbit aorta. For arterial tissues, a temperature elevation of 1.1°C was observed for a laser fluence of 60 mJ/cm2 and lesser than 1°C for lower energy levels normally associated with IVPA imaging. Therefore, the developed ultrasound technique can be used to monitor temperature during IVPA imaging. Furthermore, the analysis based on the Arrhenius thermal damage model indicates no thermal injury in the arterial tissue; suggesting the safety of IVPA imaging
Atherosclerosis; photoacoustic imaging; laser; vulnerable plaque; coronary artery; intravascular ultrasound; optoacoustic imaging; thermoacoustic imaging; temperature; Arrhenius thermal damage model
Previous ultrasound elasticity imaging experiments supported a generally accepted concept that the hardness of deep venous thrombi increases with thrombus aging. Results also showed that this non-invasive imaging technique can accurately predict thrombus age through strain estimates in a well controlled animal study. In the present study, as an alternative means to characterize elastic properties of thrombi, we employed a direct mechanical measurement system to estimate Young’s modulus of ex vivo thrombi. Unlike conventional indentation tests, the device utilizes a specific compression geometry for cylindrical tissue specimens. We also proposed an approximation scheme to retrieve Young’s modulus from force-displacement measurements made using the device. Finite element simulations and calibrations on tissue-mimicking phantoms validated the system. Then, using two groups of rats with surgically induced thrombi, we further investigated the correlation between Young’s modulus measured ex vivo and elasticity images reconstructed in vivo. This comparison was accomplished by converting the intra-thrombus strains measured in the in vivo studies into Young’s modulus estimates using a model based approach. Good agreement between time-dependent Young’s modulus estimates observed in vivo and direct measurements of Young’s modulus using the mechanical device helps to confirm the ability of elasticity imaging to age DVT for efficient treatment.
Deep venous thrombosis; Mechanical measurement; Ultrasound elasticity imaging; Elasticity reconstruction; Young’s modulus
Intravascular photoacoustic (IVPA) imaging is a catheter-based, minimally invasive, imaging modality capable of providing high-resolution optical absorption map of the arterial wall. Integrated with intravascular ultrasound (IVUS) imaging, combined IVPA and IVUS imaging can be used to detect and characterize atherosclerotic plaques building up in the inner lining of an artery. In this paper, we present and discuss various representative applications of combined IVPA/IVUS imaging of atherosclerosis, including assessment of the composition of atherosclerotic plaques, imaging of macrophages within the plaques, and molecular imaging of biomarkers associated with formation and development of plaques. In addition, imaging of coronary artery stents using IVPA and IVUS imaging is demonstrated. Furthermore, the design of an integrated IVUS/IVPA imaging catheter needed for in vivo clinical applications is discussed.
Atherosclerosis; contrast agent; imaging catheter; intravascular photoacoustic (IVPA) imaging; intravascular ultrasound (IVUS) imaging; molecular imaging; stent; vulnerable plaque
Photothermal therapy is a noninvasive, targeted, laser-based technique for cancer treatment. During photothermal therapy, light energy is converted to heat by tumor-specific photoabsorbers. The corresponding temperature rise causes localized cancer destruction. For effective treatment, however, the presence of photoabsorbers in the tumor must be ascertained before therapy and thermal imaging must be performed during therapy. This study investigates the feasibility of guiding photothermal therapy by using photoacoustic imaging to detect photoabsorbers and to monitor temperature elevation. Photothermal therapy is carried out by utilizing a continuous wave laser and metal nanocomposites broadly absorbing in the near-infrared optical range. A linear array-based ultrasound imaging system is interfaced with a nanosecond pulsed laser to image tissue-mimicking phantoms and ex-vivo animal tissue before and during photothermal therapy. Before commencing therapy, photoacoustic imaging identifies the presence and spatial location of nanoparticles. Thermal maps are computed by monitoring temperature-induced changes in the photoacoustic signal during the therapeutic procedure and are compared with temperature estimates obtained from ultrasound imaging. The results of our study suggest that photoacoustic imaging, augmented by ultrasound imaging, is a viable candidate to guide photoabsorber-enhanced photothermal therapy.
photoacoustics; optoacoustics; thermal imaging; ultrasound; photothermal therapy; treatment monitoring
The motion of a rigid sphere in a viscoelastic medium in response to an acoustic radiation force of short duration was investigated. Theoretical and numerical studies were carried out first. To verify the developed model, experiments were performed using rigid spheres of various diameters and densities embedded into tissue-like, gel-based phantoms of varying mechanical properties. A 1.5 MHz, single-element, focused transducer was used to apply the desired radiation force. Another single-element, focused transducer operating at 25 MHz was used to track the displacements of the sphere. The results of this study demonstrate good agreement between theoretical predictions and experimental measurements. The developed theoretical model accurately describes the displacement of the solid spheres in a viscoelastic medium in response to the acoustic radiation force.