Acoustics at the nanoscale (nanoacoustics): A comprehensive literature review.

Abstract

In the past decade, acoustics at the nanoscale (i.e., nanoacoustics) has evolved rapidly with continuous and substantial expansion of capabilities and refinement of techniques. Motivated by research innovations in the last decade, for the first time, recent advancements of acoustics-associated nanomaterials/nanostructures and nanodevices for different applications are outlined in this comprehensive review, which is written in two parts. As part II of this two-part review, this paper concentrates on nanoacoustics in biomedical imaging and therapy applications, including molecular ultrasound imaging, photoacoustic imaging, ultrasound-mediated drug delivery and therapy, and photoacoustic drug delivery and therapy. Firstly, the recent developments of nanosized ultrasound and photoacoustic contrast agents as well as their various imaging applications are examined. Secondly, different types of nanomaterials/nanostructures as nanocarriers for ultrasound and photoacoustic therapies are discussed. Finally, a discussion of challenges and future research directions are provided for nanoacoustics in medical imaging and therapy.

1. Introduction

The breakthroughs and advances in nanotechnology have provided various nanoscaled materials and devices with extraordinary characteristics for ultrasound investigations [1–5]. In many ways, the combination of nanotechnology with ultrasonics has revolutionized conventional approaches to use ultrasound. The emergence of different nanomaterials for the support of diagnosis and treatment of various types of diseases has attracted increasingly more attention in past decades and has now become an important field of medical ultrasound. Motivated by the research innovations over the last decade, this paper, as part II of the two-part review of nanoacoustics, will concentrate on the nanoacoustics in biomedical imaging and therapy fields, including molecular ultrasound imaging, photoacoustic imaging, ultrasound-mediated drug delivery and therapy, and photoacoustic drug delivery and therapy. In the following sections, the review proceeds as follows: in Section 2, different kinds of nanomaterials and nanostructures to be adopted as contrast agents for biomolecular ultrasound imaging and photoacoustic imaging applications will be examined in detail. In Section 3, various types of nanomaterials and nanostructures to be utilized as nanocarriers for ultrasound-mediated drug delivery and therapy as well as photoacoustic drug delivery and therapy applications will be reviewed. The last section (Section 4) provides a summary and outlook for the nanoacoustic research in these fields.

2. Nanoacoustics for biomedical imaging

2.1. Molecular ultrasound imaging

Ultrasound imaging has been a popular medical imaging technique for decades, and is now the second most implemented imaging modality in clinical practice after X-ray radiography [6]. It has been widely utilized by physicians in diverse fields, including obstetrics and gynecology (OB-GYN), cardiology, radiology, as well as gastroenterology [7–11]. Compared with other imaging modalities, such as computed tomography (CT), magnetic resonance imaging (MRI), and photoacoustic (PA) imaging, ultrasound imaging shows the advantages, such as lack of ionizing radiation, high resolution, and low cost [12].

Due to the different acoustic properties of tissues and organs, ultrasound images can be generated while ultrasonic waves propagates through them [13–15]. The generation of images by ultrasound is based on the pulse-echo principle. A short pulse of ultrasonic waves generated by an ultrasonic transducer are transmitted into issues. A short time later, the echo pulses that are reflected from the tissue and organ interface are received by the same transducer or by a different transducer and are converted into electrical signals which are then utilized to generate a grayscale image displayed on the monitor, indicating the structure information. The depth of ultrasound image depends on the time between the transmitted pulse and the received echo. The echo amplitude is digitized and illustrated in the ultrasound image.

2.1.1. Contrast enhanced ultrasound imaging

Over the last few years, molecular ultrasound imaging, combining ultrasound technique with novel molecularly targeted contrast agents, has turned out to be a promising imaging technique for studying biological processes at the molecular level [16]. Microbubbles are by far the most commonly utilized ultrasound contrast agents (UCAs) owing to their widespread approval. They are composed of a gas core surrounded by a shell and usually 1–5 μm in size [17]. The gas core can generate a strong echogenic response which leads to a high contrast ratio on ultrasound images. The gas compositions usually applied in the gas core include air, nitrogen, and biologically-inert gases such as perfluoropropane, perfluorobutane, perfluorohexane, or sulfur hexafluoride (Figure 1a) [18]. The shell is utilized to prevent both gas leakage and stabilize microbubbles. The shell materials mainly consist of phospholipid, albumin and biocompatible polymers.

Figure 1.

(a) Structure of a microbubble. Reproduced with permission [16]. Copyright 2015, Elsevier. (b) Composition of a nanosized ultrasound contrast agent. Reproduced with permission [19]. Copyright 2013, Dove Medical Press Ltd.

The bubble resonant frequency is the main parameter indicating the contrast efficiency. According to the small amplitude expansion of the Raleigh-Plesset equation, a nonlinear equation for describing oscillation of a spherical gas bubble in a liquid medium, the bubble resonant frequency is given by Equation 1 [20]:

$$f_0=\frac{1}{2\pi R_0}\sqrt{\frac{1}{{\rho }_L}[3\gamma P_0+\frac{2(3\gamma -1)\sigma }{R_0}+\frac{4\chi }{R_0}]}$$ (1)

Where f₀ denotes the resonant frequency of a bubble, R₀ denotes the bubble radius, ρ_L denotes the density of surrounding liquid, γ denotes ratio of the specific heat of the gas inside the bubble, P₀ denotes the liquid pressure, σ denotes the surface tension at the gas-liquid interface, and χ denotes the shell elasticity.

While there has been numerous diagnostic applications in clinical radiology and cardiology using microbubbles combined with ultrasound, microbubbles cannot extravasate into the tissue and therefore cannot be utilized as extravascular contrast agents due to their relatively large size. In order to exploring the extravascular space, smaller, submicron UCAs have showed widespread interest, especially in the field of oncology [12]. Submicron UCAs are so small that can extravasate through the leaky cancerous vasculature into interstitium, which is promising for the clinical applications such as highlighting lesion locations and delivering a payload of drugs [21–23]. Nevertheless, according to Equation 1, the bubble size is inversely related with the bubble resonant frequency. The higher ultrasound frequency is required for acoustic signal detection in case of the smaller bubble size. Likewise, improving shell stiffness will also lead to a higher bubble resonant frequency.

In addition, reducing the bubble size into nanoscale will not only lower its echogenicity in ultrasound examination, but decrease its stability in solution as well. The bubble stability is closely related with the Laplace pressure [24], as given by Equation 2:

$$\Delta P=\left(P_b-P_a\right)=\frac{2\sigma }{R}$$ (2)

Where ΔP is the Laplace pressure, P_b is the total pressure in bubble, P_a is the ambient pressure outside obubble. Reducing the bubble radius (R) will result in an increase of the Laplace pressure (ΔP), which will decrease the bubble stability. In order to increase the nanobubble stability, surfactants or lipids are usually bonded to the surface of nanobubbles to serve as elastic shells counteracting the effect of surface tension. Moreover, the choice of the gas core in a nanobubble is crucial to its stability as well. The application of a perfluorocarbon gas core rather than gas compositions commonly used in microbubbles, such as air, nitrogen and sulfur hexafluoride, can largely reduce the bubble dissolution time owing to its low solubility in the blood (Figure 1b).

2.1.2. Organic and inorganic nanosized UCAs

A variety of nanosized UCAs have been developed and reported by many researchers, which offers a paradigm shift of contrast enhanced ultrasound in both medical diagnostics and therapeutics [25–28]. Such agents are chemical-based agents that are produced by organic (Figure 2a–c) or inorganic materials (Figure 2d) [29]. The applications of these agents as ultrasound molecular imaging contrast agents will be examined, while many of them can also be utilized theranostic agents that provide the combination of diagnostics and therapeutics in a single nanoscale agent. The reported organic and inorganic nanosized UCAs and their applications are presented in Table 1 and Table 2, respectively [29].

Figure 2.

Schematic representation of organic and inorganic nanosized UCAs for ultrasound imaging. Organic UCAs: (a) nanobubbles, (b) phase-change droplets, and (c) gas-generating nanoparticles. (a)-(c) Reproduced with permission [29]. Copyright 2018, Elsevier. (d) The summary of inorganic nanoparticles. Reproduced with permission [30]. Copyright 2017, Elsevier.

Table 1.

Summary of organic nanosized UCAs for ultrasound imaging applications.

Organic nano sized UCAs	Composition	Diameter	Cytotoxicity	Applications
Nanobubbles (NBs)	NB-affibody conjugates [31]	478.2 ± 29.7 nm	No distinct cytotoxicity	Molecular ultrasound imaging of tumor
	Anti-PSMA nanobody coated nanobubbles [32]	487.60 ± 33.55 nm	N/A	Prostate cancer-targeted imaging
	Herceptin-targeted nanobubbles [33]	613.0 ± 25.4 nm	Low cytotoxicity	Diagnosis and treatment of Her-2-positive breast cancers
	CA-125 antibody-conjugated nanobubbles [34]	74.6 ± 16.7 nm	No recognizable cytotoxicity	Diagnosis of ovarian cancer
	Crosslinked pluronic-lipid-perfluorocarbon nanobubbles [35]	95.2 ± 25.2 nm	N/A	Tumor detection
Phase-change nanodroplets	Liquid core of perfluorooctyl bromide liposome conjugated with folic acid and polyethylene glycol [36]	301 ± 10.8 nm	N/A	Diagnosis of folate receptor (FR)-overexpressing tumors
	Phase changeable perfluoropentane nanodroplets [37]	321 ± 67 nm	N/A	tumor-targeted ultrasound imaging
	Diatrizoic acid conjugated glycol chitosan nanoparticles [38]	454.9 ± 3.33 nm	Having potential toxicity	Tumor diagnosis
	Oleic-acid-coated Fe3O4 nanoparticles [39]	294 nm	No appreciable toxicity	Tumor imaging and therapy
	Liquid perfluorohexane core encapsulated by a fluorosurfactant polymer shell [40]	180 nm	N/A	Molecular ultrasound imaging of tumor
	Perfluorohexane and carbon nanoparticles encapsulated by PLGA [41]	435.9 ± 41.31 nm	Low cytotoxicity	Sentinel lymph nodes detection and therapy of lymph nodes
	Photosensitizer and perfluorocarbon coencapsulated by lipids [42]	200 nm	No significant cytotoxicity	Photodynamic therapy
	Perfluorohexane-loaded magnetic hollow iron oxide nanoparticles [43]	537.3 nm	No distinct cytotoxicity	Stimuli-responsive cancer theranostics
	Poly lactide-glycolide acid (PLGA) nanocapsules [44]	450 nm	Negligible cytotoxicity	Cancer theranostics
	Perfluorocarbons core and lipid shell [45]	132 ± 78 nm	Low cytotoxicity	Tumor therapy
Gas-generating nanoparticles	CO2-generating carbonate copolymer nanoparticles [46]	290 ± 18 nm	No significant cytotoxicity	Tumor-targeted US imaging and US-triggered drug delivery
	H2O2-triggered CO2-generating antioxidant poly(vanillin oxalate) [47]	~550 nm	Negligible cytotoxicity	Ultrasound imaging and therapy for hepatic ischemia/reperfusion treatment
	CO2-generating nano-lipid carriers [48]	152.4 ± 0.6	No significant cytotoxicity	Temperature-controlled drug release and multimodal imaging
	Calcium carbonate loaded alginate nanocarrier [49]	~160 nm	No appreciable toxicity	In vivo tumor imaging
	CO2-generating mesoporous calcium carbonate (MCC) nanoparticles [50]	~250 nm	No significant cytotoxicity	Ultrasound imaging-guided cancer therapy
	Calcium carbonate encapsulated by poly(d,l-lactide-co-glycolide) [51]	220 nm	No significant cytotoxicity	Ultrasound imaging and treating neuroblastoma
	Doxorubicin (DOX)-loaded calcium carbonate nanoparticles [52]	240.8 ± 8.1 nm	No appreciable toxicity	Theranostic agent for cancer treatment
	Superparamagnetic iron oxide particles @ glycol chitosan nanogel [53]	218 nm	No appreciable toxicity	Dual-modality US/MR imaging
	O2-generating hybrid ICG-HANP/MnO2 nanocomplex [54]	239 ± 5.8 nm	No significant cytotoxicity	Ultrasound imaging-guided tumor photodynamic therapy

Table 2.

Summary of inorganic nanosized UCAs for ultrasound imaging applications.

Inorganic nanoparticles	Composition	Size	Cytotoxicity	Applications
Silica NPs [55]	Solid spheres	330 nm in diameter	No acute toxicity	Targetable contrast agent for ultrasound imaging
Hollow silica NPs [56]	Surface PEGylated hollow silica microspheres	~1250 nm in diameter	Low cytotoxicity	Ultrasound contrast agents for imaging
Hollow hard porous silica NPs [57]	Hollow hard porous silica shell; filled with perfluorocarbon gas	2 μm in diameter; 10 nm thick porous silica shell	Low cytotoxicity	Tumor imaging
Mesoporous silica NPs [58]	Mesoporous structure	Pore diameter 2.4 nm; pore volume 0.98 cm2.	Tumor-specific cytotoxicity	US-based breast cancer imaging
Mesoporous silica NPs [59]	Superhydrophobicity	Pore diameter < 1 nm	N/A	Targeting ultrasound imaging
Rattle-type mesoporous silica NPs [60]	Nanorattle hollow structure	NP diameter 420 nm; core diameter 260 nm	No evident toxicities	Intracellular ultrasound molecular imaging
Exosome-like silica NPs [61]	Concave mesoporous structure	140 nm in diameter	No detectable toxicity	Stem cells imaging and drug delivery
Fe-doped hollow silica nanoshells [62]	Ultrathin nanoshells with varying shell thicknesses and fused nanoflakes	2.4–22.8 nm in diameter; nanoflakes thicknesses 1.4–3.8 nm;	Minimal toxicity	Long-imaging capability for tumor
Au-PLGA NPs [63]	PLGA shell and perfluorohexane liquid core	200 nm in diameter	No detectable toxicity	Contrast-enhanced US imaging
Au-nanoshelled PLA nanocapsules [64]	Perfluorooctylbromide and superparamagnetic iron oxide nanoparticles coloading	300.4 nm in diameter	No evident toxicity	Contrast imaging-guided photothermal therapy
MnO2 NPs [54]	Catalytic property	180 nm in diameter	No significant cytotoxicity	Tumor photodynamic therapy
Albumin-MnO2 NPs [65]	Polyelectrolyte-albumin complex and MnO2 nanoparticles	~50 nm in diameter	Low cytotoxicity	Treatment of cancer
CO2-generating CaCO3 NPs [52]	Doxorubicin-loaded calcium carbonate hybrid nanoparticles	Mean diameter 145.2 to 240.8 nm	No appreciable toxicity	US imaging and simultaneous therapy of tumors
Prussian Blue NPs [66]	Catalytic property	Mean diameter 126.7 nm	No acute toxicity	Contrast-enhanced US imaging
Hollow mesoporous Prussian Blue nanocubes [67]	Hollow mesoporous Prussian Blue shell and perfluoropentane core	500 nm	No observable toxicity	US imaging-guided tumor detection and therapy
Superparamagnetic hollow Fe3O4 NPs [43]	Perfluorohexane-encapsulated hollow nanoparticles	Average particle size 537.3 nm	No distinct cytotoxicity	Stimuli-responsive cancer theranostics
Multiwalled carbon nanotubes [68]	Oxidized and functionalized to make them biocompatible (ox-MWCNT-NH3+)	Diameter 20–30 nm; Mean length 400 nm	No detectable toxicity	Contrast-enhanced US imaging
Multiwalled carbon nanotubes [69]	Antibody modification	Mean length ~150 nm	Negligible cytotoxicity	Targeted US imaging of tumor

2.1.3. Biogenic gas vesicles

Besides chemical-based agents, genetically engineered protein-shelled nanoparticles, gas vesicles (GVs), have also illustrated remarkable imaging performance as molecular ultrasound contrast agents. GVs are hollow proteinaceous nanostructures with cylinder or spindle shapes, which are composed of a 2 nm-thick protein shell that encapsulates a hollow interior normally filled with air (Figure 3a–b) [70]. The shell of GVs has a highly hydrophobic inner surface, thereby water cannot pass through it and water vapor cannot condense inside [71]. However, ambient gases are able to freely diffuse into and out of GVs. In 2014, Shapiro et al. [72] first found that purified GVs from a kind of photosynthetic bacteria which were used to regulate buoyancy could scatter ultrasound waves and thus produce ultrasound contrast.

Figure 3.

(a) Transmission electron microscopy (TEM) image of a gas vesicle (scale bars, 150 nm). (b) Composition of a gas vesicle. (c) Engineered gene, ARG1, comprising genes from A. flos-aquae (green) and B. megaterium (gray) to make the gas vesicles detectable by ultrasound in heterologous host. (d) TEM image of an E. coli Nissle 1917 cell expressing ARG1 (scale bar, 50 nm). (e) Ultrasound image of a live mouse with ARG1-expressing E. coli arranged in the colon (scale bar, 2.5 mm). (f) Ultrasound images of ARG1 and ARG2 before and after the application of two different collapse pressures. Reproduced with permission [83]. Copyright 2018, Elsevier.

Because GVs are filled with gas, it was assumed that GVs could act as nanoscale contrast agents for ultrasound imaging. This hypothesis was then verified by using GVs purified from Anabaena flos-aquae and halobacteria NRC-1 and ultrasound imaging in a gel phantom at 4.8, 8.6 and 17 MHz [72]. Following this pioneering finding, many studies about using GVs as biomolecular contrast agents have been conducted, such as exploring their acoustic properties [73,74], engineering them through genetic and biochemical modulations [75,76], investigating the in vivo biodistribution as molecular contrast agents [77], using them as biomolecular tool for deep tissue imaging [78–80], imaging of microorganisms inside living animals (Figure 3c–f) [81], as well as more dedicated imaging of generic expression in mammalian cells [82].

2.2. Photoacoustic (PA) imaging

2.2.1. Principle of PA imaging

Photoacoustic (PA) imaging (also referred to as optoacoustic imaging) is an emerging multiscale functional, anatomical, and molecular imaging modality, having demonstrated great promise in preclinical researches and clinical applications [84]. PA imaging is based on photoacoustic effect that light absorbed by tissues is used to excite ultrasonic waves via thermoelastic expansion. In PA imaging, nanosecond duration laser pulse is delivered into the animal tissues (Figure 4). The absorption of short laser pulses by endogenous chromophores (such as hemoglobin and melanin) or exogenous contrast agents (such as small molecule dyes and carbon nanoparticles) is converted into thermal energy that results in transient thermoelastic expansion to generate acoustic waves in 3D space. The ultrasound waves can then be detected with an ultrasonic transducer or a transducer array surrounding the tissue or animal body. The detected ultrasonic waves are analyzed to reconstruct a photoacoustic image of tissue optical absorption [85].

Figure 4.

Schematic of working principle of PA imaging. Reproduced with permission [85]. Copyright 2016, Ivyspring International Publisher.

A PA imaging system mainly consists of an optical source for short laser pulse generation, an ultrasonic transducer array for acoustic wave detection, and a data acquisition (DAQ) system [86]. The pulse repetition frequency of the laser pulse and the time for ultrasonic signals detection are two major factors determining the PA imaging frame rate. Combining advantages from both optical and ultrasound imaging, PA imaging offers unique advantages over existing imaging modalities. First of all, PA imaging is free of ionizing radiation and noninvasive, thus it can be safely used in vivo. Secondly, it has higher resolution and deeper penetration depth (~ 4 cm in vivo and ~ 12 cm in vitro) compared to optical imaging techniques (~ 1 mm). Moreover, compared with other imaging modalities such as MRI and CT, PA imaging is much faster and less expensive. Other benefits include speckle free and label free [85,87–90]. With all these benefits, PA imaging has been found applications in diverse fields, including oncology, vascular biology, dermatology, neurology, ophthalmology, and cardiology [91–94].

2.2.2. Organic and inorganic nanosized contrast agents

Although PA imaging is able to penetrate deeper into tissue than other optical imaging methods, owing to the laser light scattering, the photoacoustic signal-to-noise ratio and light intensity attenuate exponentially with the increase of penetration depth, affecting the imaging contrast and resolution [85,95]. In order to enhance the PA imaging sensitivity and contrast, various oral or injectable chemicals, i.e., exogenous contrast agents, have been adopted to change the optical and acoustic properties of structures within the body [96]. Thus, the PA imaging contrast and resolution are reinforced with the assistance of exogenous contrast agents. In this section, the recent development of exogenous contrast agents that address biomedical applications and clinical practice will be examined. For the characteristics of endogenous chromophores and biomedical applications of PA imaging assisted with these chromophores, readers are referred to the references [97].

The exogenous PA contrast agents can be classified into organic and inorganic contrast agents according to their structures. A summary of recent reported PA organic and inorganic nanosized contrast agents is shown in Table 3. The highlights of different types of organic and inorganic nanosized contrast agents are presented in the following section. Various biomedical applications of PA imaging including bioimaging of tumors and biosensing of tumor microenvironments are summarized in Table 4.

Table 3.

Organic and inorganic nanosized contrast agents reported in the literatures for PA imaging applications.

Organic nanomaterials	Small organic molecules	Cyanine-based dyes, croconine, naphthalocyanines, perylene-diimide (PDI), phthalocyanine, porphyrin, porphysome, squaraine
	Semiconducting polymer NPs	semiconducting polymers, PDI-based NPs, organic NPs, oligomers, chlorin dimers, polypyrrole, polylysine, natural humic-acid-based nanoagents
Inorganic nanomaterials	Metallic NPs	Gold NPs, gold NP assemblies, Prussian blue, copper neodecanoate NPs, Ag NPs, Pd nanosheets
	Transition metal chalcogenides/MXene-based NPs	CuS, Ag2S, CulnS/ZnS, MoS2, TiS2, WS2, CoS2, ReS2, Bi2S3, MoSe2, Nb2C, TaC, Ti3C2, S2V
	Carbon-based NPs	Au coated SWCNTs, single-walled carbon nanotubes (SWCNTs)-Arg-Gly-Asp (RGD) conjugation, rGO loaded AuNR, rGO loaded AuNRVe, CNTR@AuNPs
	Others	Te nanosheets, B nanosheets, Cu-Ag2S NPs, black-phosphorus quantum dots, black-phosphorus nanosheets, Fe@γ-Fe2O3@H-TiO2 nanocomposites, MnOx loaded Ti3C2

Table 4.

Summary of PA imaging applications using different nanosized contrast agents.

Application	Nanosized UCA	Features	Performance
Deep tumor imaging [98]	Micelle enveloped PDI NPs	High PA imaging sensitivity, high photostability and biocompatibility	The NPs provided high PA imaging contrast for detection of deep brain tumor in living mice.
Brain tumor imaging [99]	Second NIR (NIR II) conjugated polymer NPs	High imaging contrast, high photostability and good biocompatibility	The NPs showed much weakened light attenuation, strong signal/background ratio, and large penetration depth for brain tumor imaging.
Brain vascular imaging [100]	Conjugated polymer nanoparticles (CPNs)	High photostability, exhibiting no toxicities both in vitro and in vivo	The NPs can greatly enhance PA signals for brain vascular imaging.
Deep brain glioma imaging [101]	MoS2-ICG hybrid	High PA imaging sensitivity, enhanced photoacoustic conversion efficiency compared with ICG	Brain tumor at twofold deeper site beneath a mouse scalp was clearly identified by using the hybrid as contrast agent.
Mouse brain imaging [102]	ICG - containing porous Si NPs	ICG sealed within a rigid NP that can protect the ICG payload from thermal and photolytic degradation	The PA efficiency was increased 17-fold compared with free ICG.
Vessel atherosclerosis imaging [103]	ICG@PEG-Ag2S nanoprobe	Long blood circulation, hemocompatibility, and no organ toxicity	Atherosclerotic plaques were imaged in a high contrast-enhanced manner.
Venous thrombosis imaging and antithrombotic therapy [104]	Thrombus-specific theranostic NPs	H2O2-triggered photoacoustic signal amplification	Photoacoustic contrast was greatly enhanced by using the NPs as contrast agent in a thrombosed vessel.
Venous thrombosis imaging [105]	cRGD peptide modified PDI NPs	High PA intensity, high photostability and biocompatibility, and excellent binding ability with GPIIb/IIIa	The PA contrast enhanced by the NPs provided effective information about the thrombus including the profile, size, and spatial distribution.
Sentinel lymph node (SLN) imaging [106]	Silica-coated gold nanoplates	High photothermal stability, and low cytotoxicity	Strong and sustained PA signals were obtained for SLN imaging in a mouse model.
Lymph node imaging in living mice [107]	Semiconducting polymer nanoparticles	High structural flexibility, high photostability, and narrow photoacoustic spectral profiles	They can not only be utilized for LN mapping but also for PA imaging of ROS.
Sentinel lymph node imaging [108]	Encapsulated conjugated oligomer NPs	Excellent photostability and biocompatibility, strong absorption in the NIR range	The NPs showed great potential for SLN imaging and photothermal therapy.
Detection of cancer-related matrix metalloproteinases [109]	Copper sulfide NPs	Strong absorbance in the NIR range, good photostability	The NPs could dramatically improve the tissue penetration depth for PA imaging.
Tumor hypoxia imaging [110]	Ratiometric hypoxia probe 1 (rHyP-1)	Strong NIR absorption, and deep penetration depth	The probe enabled high resolution hypoxia detection using PA imaging at centimeter depths.
Tumor pH imaging [111]	pH - responsive albumin - based nanoprobe	High biocompatibility, easy to operate, and depth independent accuracy	The nanoprobe was clinically adoptable for real-time pH imaging of the whole tumor.
pH imaging [112]	Semiconducting oligomer NPs	Ultrasmall size, high stability, and high pH sensitivity	The NPs allowed real-time PA imaging of pH in tumors in living mice.
Lithium detection [113]	Lithium ionophore	Monitoring therapeutic drug concentrations in vivo based on PA imaging	To track the concentrations of therapeutic drugs and improve disease management.
Methylmercury (MeHg+) detection [114]	Liposome (LP) and cyanine dye (hCy7) nanoprobe	The hydrophobic hCy7 was encapsulated in the lipid layer of the liposome. High biocompatibility and negligible cytotoxicity	The nanoprobe can detect MeHg+ in vivo by ratiometric photoacoustic bioimaging.

(1). Organic contrast agents.

A typical representative of small organic molecules is cyanine-based dyes that are commonly utilized as PA imaging contrast agents as a result of their good photostability, high photoconversion efficiencies and negligible toxicity [115]. Kang et al. [102] reported a method to improve the PA response of indocyanine green (ICG), one of the commonly applied PA imaging agents, by encapsulating it in a porous silicon nanoparticle host. Because of the low thermal conductivity of the host, the PA generation efficiency of the composite structure was increased 17-fold compared with free ICG (Figure 5a). Liu et al. [114] developed a ratiometric photoacoustic nanoprobe consisting of liposome and MeHg⁺-responsive NIR cyanine dye for MeHg⁺ detection in living subjects (Figure 5b). It was found that in the presence of MeHg⁺, the ratiometric photoacoustic signal was increased remarkably. Chen et al. [111] fabricated a nanoprobe composed of human serum albumin (HSA) and NIR dye (BPO_x and IR825) nanocomplexes for sensing tumor microenvironment pH using ratiometric photoacoustic imaging (Figure 5c). The tumor microenvironment pH could be detected by using the nanoprobe in combination of ratiometric photoacoustic imaging, which was very useful for regulating the proliferation of cancer cells.

Figure 5.

(a) Procedure used to simultaneously load and seal CG into the pores of porous silicon nanoparticles (pSiNPs). Reproduced with permission [102]. Copyright 2018, Wiley-VCH. (b) Mechanism of ratiometric PA imaging of MeHg⁺. Reproduced with permission [114]. Copyright 2017, Wiley-VCH. (c) The preparation process of C-HSA-BPOx-IR825. Reproduced with permission [111]. Copyright 2015, Wiley-VCH.

Besides, owing to their strong optical absorption and high photostability, semiconducting polymer nanoparticles (SPNs) have also been commonly adopted [115]. By encapsulating PDI into micelle, a kind of amphiphilic molecule, Fan et al. [98] developed a highly efficient PA imaging contrast agent, micelle-enveloped PDI, for imaging of deep brain tumors (Figure 6a). Their studies found that the organic NPs illustrated strong PA absorption in the NIR range, and they could be utilized to detect brain tumors successfully in living mice. Following that, to diagnosis of early thrombus and monitoring thrombolysis, Cui et al. [105] developed cyclic Arg-Gly-Asp (cRGD) peptide modified PDI (cRGD-PDI) NPs as PA agents. The cRGD-PDI NPs showed strong PA imaging contrast and provided 4-folder greater PA intensity of early thrombus than the control group in a living mouse (Figure 6b). In another study, in order to achieve the combined PA and US imaging, Tang et al. [116] fabricated an organic semiconducting nanodroplet consisting of a PDI shell, a liquid perfluoropentane core and photosensitizers of ZnF₁₆P_c molecules at the interface of the core-shell as PA and US dual-contrast agents (Figure 6c). The nanodroplets illustrated a high tumor accumulation in a living mouse due to their nanoscale size. Under a 671 nm laser irradiation, the nanodroplets could be effectively applied for dual-modal PA and US imaging-guided photothermal and photodynamic therapy.

Figure 6.

(a) PA imaging process of brain tumor in vivo by PDI NPs. Reproduced with permission [98]. Copyright 2015, Wiley-VCH. (b) The preparation of cRGD-PDI NPs. Reproduced with permission [105]. Copyright 2017, American Chemical Society. (c) IRDye800CW-labeled photosensitizer ZnF₁₆P_c-loaded PDI photoacoustic nanodroplet (PS-PDI-PAnD). Reproduced with permission [116]. Copyright 2018, American Chemical Society.

(2). Inorganic contrast agents.

Metallic nanomaterials, especially gold nanoparticles (AuNPs), can achieve strong optical absorption via the localized surface plasmon resonance (LSPR) [85]. By synthesizing AuNPs with different morphologies, their optical absorption properties can be maximized in the NIR region. Other notable advantages of AuNPs include their biocompatibility, high photothermal conversion efficiency, and good chemical inertness [117,118]. To date, different types of AuNPs have been reported including nanospheres [119–121], nanorods [122–125], nanoprisms [126–128], nanocages [129–131], nanostars [132–134], nanoplates [106,135,136], nanodisks [137–139], nanoshells [64,140–142], nanovesicles [123], nanotripods [143], nanowreaths [144], nanoflowers [145,146], and bipyramids and rhombic dodecahedra [147], as shown in Figure 7.

Figure 7.

Microscopy photographs of different types of gold nanoparticles: (a) nanospheres, (b) nanorods, (c) nanoprisms, (d) nanocages, (e) nanostars, (f) nanoplates, (g) nanodisks, and (h) nanoshells. (a) Reproduced with permission [148]. Copyright 2009, Elsevier. (b) Reproduced with permission [149]. Copyright 2012, American Chemical Society. (c) Reproduced with permission [126]. Copyright 2015, American Chemical Society. (d) Reproduced with permission [131]. Copyright 2017, Elsevier. (e) Reproduced with permission [150]. Copyright 2018, Wiley-VCH. (f) Reproduced with permission [135]. Copyright 2014, Wiley-VCH. (g) Reproduced with permission [139]. Copyright 2017, American Chemical Society. (h) Reproduced with permission [142]. Copyright 2017, American Chemical Society.

Besides the gold nanocrystals, gold nanocrystal assemblies also show outstanding optical absorption properties in the NIR region. Bai et al. [151] fabricated a multifunctional nanocomposite combining imaging (PA imaging and MRI) and therapeutic functions for cancer cells imaging and photothermal therapy, which was composed of hollow gold nanospheres as cores and superparamagnetic iron oxide NPs as satellites. Song et al. [152] developed a plasmonic vesicular nanostructure comprising of amphiphilic gold nanocrystals with polymer brush coatings. The nanocrystals imparted their remarkable properties to the assembly, and the strong interparticle plasmonic coupling caused collective properties different from those of the constitutive units. In another work, Deng et al. [153] developed a theranostic agent composed of comb-like amphipathic polymer template and DOX@gold nanomicelles for dual-modal imaging (CT and PA) and the combination of chemotherapy and photothermal therapy. Their studies in a mouse model demonstrated that the nanomicelles could be effectively used to detect tumors for CT and PA imaging simultaneously. Carbon-based nanomaterials mainly consisting of CNTs [154–156] and graphene-based nanomaterials [157–161], have been widely adopted as contrast agents in PA imaging as well (Figure 8a). The major advantages of carbon-based nanomaterials are their broad absorbance band ranging from UV to NIR region as well as ease to integrate with various materials or compounds for multiple functions. De La Zerda et al. [162] synthesized a PA contrast agent based on SWNT conjugated with RGD peptides for PA imaging of tumors. According to the tumor detection in a living mouse, the PA signal of the tumor in the mouse injected with the contrast agents was eight times stronger than that of the control group. Kim et al. [163] reported gold-plated carbon nanotubes as contrast agents for PA imaging, which were composed of shortened SWNT core coated by a thin layer of gold. This novel hybrid plasmonic contrast agent, combined the advantages of gold and CNTs, presented a unique set of features. Song et al. [164] proposed a PA contrast agent fabricated by carbon nanotube ring (CNTR) template and AuNPs coating for imaging-guided cancer therapy (Figure 8b). Compared with CNTR and CNTR@AuNS, the CNTR@AuNP illustrated much stronger Raman and optical signals. Moon et al. [165] developed a PA imaging contrast agent rGO-AuNRs using an optically absorbing material rGO coated with gold nanorods (Figure 8c). The rGO-AuNRs demonstrated greatly enhanced PA signal intensity and photothermal efficiency, which was a promising nanomaterial for sensitive disease diagnosis and photothermal therapy. In addition, Song et al. [166] fabricated a carbon-metal hybrid rGO-loaded plasmonic gold nanorod vesicle for dual chemo-photothermal therapy. The rGO was conjugated with DOX and the rGO-DOX conjugation was encapsulated in the AuNRVe to avoid direct interaction with the physiological environment.

Figure 8.

(a) Structures of 0D fullerene, 1D carbon nanotube, 2D graphene and 3D graphite formed by the carbon atoms. (b) Preparation of CNT ring Au nanoparticles (CNTR @ AuNPs). (c) Preparation and application of reduced graphene oxide Au nanorods (rGO-AuNRs). (a) Reproduced with permission [167]. Copyright 2011, Wiley-VCH. (b) Reproduced with permission [164]. Copyright 2016, American Chemical Society. (c) Reproduced with permission [165]. Copyright 2015, American Chemical Society.

3. Nanoacoustics for medical therapy

3.1. Ultrasonic drug delivery

Apart from diagnostic applications of ultrasound, therapeutic applications of ultrasound, such as ultrasound-mediated drug/gene delivery, has become an emerging topic of expanding interests. Starting from the mid-90s, many studies have reported that the exposure to ultrasound can temporarily enhance the cell membrane permeability (i.e., sonoporation), thus creating a physical channel for delivery of cell-impermeable drugs (Figure 9) [168]. The main benefit of delivery of therapeutics using ultrasound is that the drug delivery and release is spatially and temporally controlled in the target region, thus lowering the dosage use and the adverse side effects [169,170]. Different therapeutics including small-molecule drugs, protein drugs, peptide drugs, and genetic drugs have been reported to be successfully delivered with the assistance of ultrasound waves [171–176].

Figure 9.

Schematic illustration of the mechanism of ultrasound-mediated nanoparticle delivery. Reproduced with permission [177]. Copyright 2017, Korean Society of Ultrasound in Medicine.

While microbubbles have been proved to be reliable tools for ultrasound-mediated therapeutic delivery, the key limitation of microbubbles is that the size of microbubbles is too large to penetrate into tumor tissues, thereby only confined their applications in blood circulation and cannot be effectively utilized for cancer therapy [23]. Various techniques have been devised for the use of nanosized bubbles/particles for delivery of therapeutics to targeted tissues [178]. Nanobubbles/particles, acted as delivery carriers, are able to deliver drugs precisely to the target tissues while maintaining efficiency. Moreover, combined with the visual and noninvasive features of ultrasound, the motion of drug nanocarriers can be tracked and the release of drugs from nanocarriers can be controlled by the ultrasound irradiation [179,180]. For all these reasons, the development of safe and efficient novel ultrasound-responsive therapeutic nanocarriers could greatly impact therapy strategies.

3.1.1. Nanocarriers for ultrasonic drug delivery

Recently, there have been various nanomaterials served as nanocarriers for delivery of anticancer drugs to tumor tissues [181–183], delivery of thrombolytics to break up or dissolve blood clots [184–186], delivery gene to target tissues [187–190], as well as delivery insulin to regulate blood glucose level [191,192] (Figure 10). The recent development of different types of nanocarriers for ultrasonic drug delivery are highlighted in this section.

Figure 10.

Schematic illustration of the 8 most reported nanocarriers: (I) liposomes, (II) micelles, (III) dendrimers, (IV) meso-porous silica nanoparticles (MSNs), (V) gold nanoparticles (AuNPs), (VI) super paramagnetic iron oxide nanoparticles (SPIONs), (VII) carbon nanotubes (CNTs), and (VIII) quantum dots (QDs). Reproduced with permission [193]. Copyright 2019, Elsevier.

(1). Liposomes.

Liposomes, typically with a diameter of 100–200 nm, are composed of an aqueous core surrounded by a self-assembled lipid bilayer membrane. Due to their excellent biocompatibility and versatility, liposomes has become one of the most widely utilized nanocarriers for ultrasound-mediated drug delivery [194]. Marxer et al. [195] developed a long-circulating drug nanocarrier with three different lipid formulations for ultrasound-enhanced drug delivery. The physico-chemical properties of the developed lipid dispersions were compared with a commercially available contrast agent, which demonstrated adjustable properties including circulation stability, biocompatibility, and ultrasound reflectivity. In another study, Becker et al. [196] compared the sonothrombolytic efficacy of the three nanoscaled lipid formulations with the commercially available contrast agent for human blood clot under diagnostic ultrasound. Their results illustrated that the nanoscaled formulation of DSPC/PEG40S had the most significant clot weight reduction under the irradiation of ultrasound at 1.4 MHz, even without thrombolytic drugs. Lattin et al. [197] reported an echogenic liposome that can release drugs from liposomes under ultrasound excitation. The developed liposomal drug nanocarrier was called an eLiposome (liposomes containing emulsion droplets), formed by using a lipid sheet folding technique. The eLiposome had an overall diameter of 800 nm and contained emulsion droplets 100–150 nm and 470 nm, respectively. Under ultrasound excitation, vapor phase was formed and expanded from the droplets, which would disrupt the liposomal membrane and lead to local drug release.

(2). Micelle.

Micelle is typically spherical in shape, which is formed by a series of amphiphilic molecules that have a natural tendency to clump together. The hydrophobic drugs can be segregated in the hydrophobic core of micelle until they are released. Diaz de la Rosa et al. [198] investigated anticancer drug DOX release behavior from Pluronic P105 unstabilized and stabilized micelles. The in vitro experiments found that the drug release from unstabilized as well as stabilized micelles at ultrasound frequency < 90 kHz, while no drug was released from the micelles at ~500 kHz. This was mainly attributed to the different dynamics of oscillating bubbles under the different ultrasound excitations. Husseini et al. [199] fabricated DOX encapsulated Pluronic P105 micelles with a folic acid moiety to enhance the tumor cells targeting. They found that the percentage of drug release was improved with the rise of ultrasound power intensity under 70 kHz excitation. The maximum amount of DOX release was 14%, which was measured at an ultrasound intensity of 5.4 W/cm². Wu et al. [200] developed anticancer drug curcumin (Cur) encapsulated pluronic P123/F127 mixed micelles for site-specific sonochemotherapy. Under 1 MHz focused ultrasound excitation, the Cur-loaded micelles demonstrated tumor-targeting deposition, resulting from the ultrasound sonoporation. Moreover, the ultrasound-triggered Cur release from the Cur-loaded micelles was highly dependent on the ultrasound intensity.

(3). Polymeric nanoparticles.

Polymeric nanoparticles mainly consist of nanocapsules, nanospheres, and polymersomes, which demonstrate the properties of enhanced encapsulation and controlled release of therapeutics [201]. Nestor et al. [202] fabricated air-filled nanocapsules with a biodegradable polymeric shell for tumor detection. The nanocapsules had a mean diameter of 370 nm and a spherical shape with smooth surfaces as well as a capsular morphology. The polymeric shell could potentially entrap drugs inside it, which made this kind of structure promising for theranostics agents. Yang et al. [203] reported a biodegradable nanocapsule that was filled with perfluorohexane and had the DOX-loaded PMAA as wall for achieving the ultrasound imaging and drug delivery functions. The developed nanocapsules could easily enter the tumor tissues due to their small and uniform size. Moreover, the filled perfluorohexane could be easily vaporized via ultrasound excitation, thus enhancing ultrasound imaging signals and ensuring imaging-guided drug delivery. Chen and Du [204] developed polymer vesicles that responded to both ultrasound and pH stimuli for drug entrapment and release. The size of the vesicles would become smaller under either ultrasound excitation or pH decrease. The encapsulated anticancer drugs could reach controllable release when exposed to ultrasound irradiation or pH variation.

(4). Nanobubbles.

Nanobubbles are commonly adopted as theranostic agents due to their great imaging ability. Cavalli et al. [205] developed DNA-loaded chitosan nanobubbles which were composed of a perfluoropentane core and a chitosan shell as ultrasound-triggered gene delivery system. The plasmid DNA was loaded onto the chitosan shell, which was expected to have long circulation time and accumulate into tumor tissues easily. Based on the in vitro experiments, while in the absence of ultrasound, the nanobubble-bound plasmid DNA was not released at all, after one minute of 2.5 MHz ultrasound excitation, the release of plasmid DNA occurred. Yin et al. [206] developed siRNA-loaded nanobubbles which were fabricated by the negatively charged gas-cored liposomes and positively charged siRNA micelles for tumor treatment. Under 1 MHz ultrasound irradiation, the siRNA micelles were effectively released from the nanobubbles in tumor tissue and delivered into cancer cells.

(5). Perfluorocarbons (PFCs).

Rapoport et al. [207] reported paclitaxel (PTX)-loaded perfluoropentane (PFP) nanoemulsions that could effectively accumulate in tumor tissue and transformed into microbubbles in sonication. The ultrasound imaging properties and therapeutic efficacy of the nanoemulsions were experimentally verified in a mouse model. After injection of the drug-loaded nanoemulsions, tumors were significantly regressed after1 MHz ultrasound irradiation. In another study, Thakkar et al. [208] explored the effect of ultrasound irradiation on the penetration of drug-loaded nanoemulsions through vascular walls. Mouse carotid arteries were utilized as a blood vessel model. They found that under 1 MHz irradiation, the permeability of the arteries to the nanoemulsions was much more pronounced by using continuous ultrasound wave than that of pulsed ultrasound.

(6). Quantum dot.

Based on the fact that ultrasound irradiation can enhance cell membrane permeability (sonoporation), Thein et al. [187] developed an ultrasonic micro-transducer array for cellular level controllable sonoporation that accelerates delivery of drugs or gene therapeutics to cells (Figure 11). The developed transducer array was composed of 3 × 3 arrays of PMN-PT transducers (each area 25 μm × 25 μm) with a central frequency of 30 MHz and lateral resolution of 18 μm for site-specific sonoporation. CdSe/ZnS quantum dots were utilized as nanocarriers to quantitatively evaluate the sonoporation degree of the human melanoma cells seeded on the surface of the transducer array. It was found that the cell membrane wound size and cellular uptake of quantum dots were increased linearly with the rise of ultrasound irradiation pressure.

Figure 11.

(a) SEM mage of a 3 × 3 micro-transducer array (each area 25 μm × 25 μm). (b) Electrode above each micro-transducer. (c) Human melanoma cells above a micro-transducer. (d) Schematic illustration of ultrasound-mediated quantum dots (QDs) delivery at cellular level using the micro-transducer array. (a)-(d) Reproduced with permission [187]. Copyright 2011, Elsevier.

Besides the nanocarriers reviewed above, other inorganic materials including mesoporous silica nanoparticles [209–214], Au nanocages [215–217], magnetic nanoparticles [218–220], and carbon-based nanovehicles [69,221–224] have also been reported by researchers to be utilized as nanocarriers for ultrasound-mediated drug delivery.

3.2. Ultrasound therapy

3.2.1. Ultrasound-triggered therapy

Ultrasound has long been employed in medicine for the treatment of many different pathologies, such as ultrasound-assisted liposuction, lithotripsy for kidney stone, and accelerating wound healing, mainly attributed to its advantages of safety to human tissues, low cost and deep tissue penetration capability [225]. Recently, high-intensity focused ultrasound (HIFU) has been served as a new noninvasive treatment technique for thermal ablation of local tumors [226]. Sonodynamic therapy (SDT) that is based on low-intensity ultrasound and chemotherapeutic agents (known as sonosensitizer) has been reported by researchers to noninvasively eradiate solid tumors due to the ultrasound-induced cytotoxic effects [227]. Compared with the current major therapeutic techniques including chemotherapy and gene and cell therapy, ultrasound-triggered therapy is able to focalize at target tissues and reduce biotoxicity. In addition, better treatment could be achieved while combining the ultrasound method with conventional therapies [228].

Over the past few decades, the rapid development of nanomaterials and nanostructures not only advances ultrasound imaging by providing nanosized contrast agents with outstanding properties, but also accelerates ultrasound therapy with greatly enhanced therapeutic efficiency owing to their unique physiochemical properties, such as surface features, geometry, porosity, and composition [229]. Different nanoparticles can also potentially combine several imaging modalities and therapeutic functions into one nanoplatform to be utilized as multifunctional theranostic agents for imaging-guided synergistic therapy. Miscellaneous nanoparticles including organic nanomaterials and inorganic nanomaterials have been adopted in studies for treatment various diseases such as tumors, cardiovascular diseases, and central nervous system diseases [228].

While this section is focused on nanomaterials and nanostructures for ultrasound-triggered therapy applications, many recent published review articles have already covered this topic, especially the review about nanoparticles for ultrasound-triggered cancer therapy. Considering that several comprehensive reviews have summarized and discussed the emerging development of exploring organic and inorganic nanomaterials for ultrasound-triggered therapies, the same topic will not be reviewed again in this paper and the readers could refer to very recent reviews. Yang et al. [228] focused on the current applications of ultrasound-triggered therapy (2019). Zhou et al. [230] summarized the development of organic and inorganic nanoparticles for ultrasound-based cancer therapy (2020). Yang et al. [231] reviewed the recent advance of 2D nanomaterials for ultrasound-triggered cancer therapy (2020). Fan et al. [232] comprehensively examined nanotechnology-mediated synergistic cancer therapy (2017). Xu et al. [233] focused on different nanoparticles in sonodynamic therapy (2016). Unga et al. [234] reviewed nanosized agents for ultrasound-induced cancer immunotherapy (2014).

3.2.2. Nanoparticle-mediated sonothrombolysis

While blood clot, or thrombus, helps human body stop bleeding when a person gets hurt, blood clots forming in veins or arteries are dangerous to human health. When blood clots form inside a vein or artery (i.e., thrombosis), they will reduce the blood flow past the clot or even totally block the blood flow. If the thrombosis forms in the legs, it causes deep vein thrombosis; if it is formed in the lung, life-threatening pulmonary embolism will be occurred; if the blood clots block the blood supply to the brain, ischemic strokes will happen [235]. Thrombolysis, known as thrombolytic therapy, is a treatment to dissolve blood clots that have blocked veins or arteries and pose potentially serious or life-threatening complications. Sonothrombolysis is an emerging technique that employs ultrasound waves to break down blood clots in vessels. The widely accepted mechanisms of sonothrombolysis are ultrasound-induced stable and inertial cavitation, microstreaming, microjetting and increased diffusion of therapeutics into the clots due to acoustic radiation force [236]. Conventionally, sonothrombolysis is usually adopted in combination with thrombolytic agents, such as tissue plasminogen activator (tPA) or microbubbles. However, the application of tPA will carry the risk of systemic hemorrhage in patients, and the use of microbubble is showed to be slow for treatment (usually several hours) and less effective for aged clots [237,238].

Nanodroplets, a generally applied ultrasound contrast agent that has a diameter of 100–300 nm, has been shown to be effective for sonothrombolysis [239]. Compared with microbubbles, the advantages of nanodroplets include their longer circulation time in blood and deeper penetration into clots due to their smaller size. Recently, low-boiling-point (−2°C) lipid-shell perfluorocarbon phase-change nanodroplets were adopted by our group for sonothrombolysis based on the assumption that the smaller nanodroplets could permeate into a clot easier compared to microbubbles, thereby greater cavitation effects would be generated inside the clot and lysis rate would be improved [240]. The efficiency of the nanodroplets for sonothrombolysis was compared with that of microbubble-mediated sonothrombolysis. Under the same ultrasound excitation conditions (I_SPTA 0.384 W/cm², peak negative pressure 8.0 MPa), the nanodroplets-mediated treatment demonstrated a thrombolysis rate of 140% over the microbubble-mediated treatment. In another study, we conducted in-vitro study to compare the sonothrombolysis efficacy of nanodroplets on unretracted (< 24 hours old) and retracted (> 3 days old) blood blots with that of microbubbles [241]. The results illustrated that while the unretracted clot mass reduction was similar for both treatment conditions, the retracted clot mass reduction for the nanodroplet-mediated treatment was 1.6-fold over that of microbubble-mediated treatment. Under the treatment condition that tPA was combined with the nanodroplet-mediated sonothrombolysis for retracted clots, compared with control group (tPA + ultrasound), the nanodroplet-mediated sonothrombolysis with tPA illustrated greatly larger mass reduction (38% vs. 4%) [242].

Besides nanodroplets, magnetic microbubbles are also proved to be promising for sonothrombolysis. Magnetic microbubble (MMB), composed of a gas core and a shell of Fe₃O₄ nanoparticles, not only possesses the acoustic properties of a microbubble but also is sensitive to magnetic field excitation (Figure 12c) [243]. In our recent studies [244,245], MMBs were utilized in conjunction with an intravascular forward-looking ultrasound transducer (620 kHz) for MMBs-mediated sonothrombolysis (Figure 12a). We assumed that under a rotational magnetic field, the MMBs would be trapped and oscillated close to the target blood clot; the induced vortex-like microstreaming would enhance the cavitation effect generated by sonication. Based on our in vitro studies, the mass reduction of MMBs-mediated sonothrombolysis for unretracted and retracted clot was 1.4-fold over that of microbubble-mediated sonothrombolysis. To achieve precise and controllable delivery of thrombolytic drugs, Wang et al. [186] synthesized a nanoparticle-shelled magnetic microbubble (MMB-SiO₂-tPA) for realizing the magnetic targeting under a magnetic field and ultrasound-triggered tPA release due to the microstreaming. The microbubble was composed of an air core and a shell of nanoparticles consisting of Fe₃O₄ nanoparticles and tPA-containing mesoporous silica nanoparticles (Figure 12b). Under a low-intensity ultrasound excitation, both the penetration depth of tPA into clot and thrombolysis efficacy were improved. Compared with conventional injection of tPA (0.03 mg/kg), the thrombus mass in a mouse model was reduced by 67.5% using the MMB-SiO₂-tPA treatment.

Figure 12.

(a) Illustration of magnetic microbubbles (MMBs)-mediated sonothrombolysis. (b) Schematic view of SiO₂-tPA shelled MMBs for targeted tPA delivery and controlled release. (c) Composition of MMBs. (a) Reproduced with permission [245]. Copyright 2019, Elsevier. (b) Reproduced with permission [186]. Copyright 2020, American Association for the Advancement of Science. (c) Reproduced with permission [243]. Copyright 2016, Springer Nature.

3.3. Photoacoustic drug delivery and therapy

The PA imaging nanosized contrast agents have also been adopted by researchers as drug delivery vehicles since studies have found that the ultrasound irradiation generated by the PA effect could effectively accelerate the delivery and release of drugs in target tissues [246–248]. Furthermore, the local temperature rise of nanoparticles induced by the laser irradiation can cause thermal ablation and subsequent cancer cell death, i.e., photothermal therapy (PTT) [249]. The encapsulated drug release from nanocarriers can be promoted as well by the local heating generated in tumor tissues exposed to NIR laser irradiation [250,251].

3.3.1. Organic nanocarriers

Small molecular dyes and semiconducting polymer nanoparticles have also been commonly adopted for theranostic applications [252–258]. Gong et al. [259] reported a multifunctional nanomicelle fabricated by polyethylene glycol (PEG) coating and C18PMH conjugation for imaging-guided cancer therapy. Photosensitizer chlorin e6 (Ce6) was conjugated with C18PMH and photothermal agent IR825 was encapsulated into the nanomicelles. The resulted IR825@C18PMH-PEG-Ce6 nanomicelles could realize photoacoustic, fluorescence, and magnetic resonance triple modal imaging of tumors as well as combined photothermal and photodynamic therapy. Wang et al. [260] developed acid-switchable multifunctional nanomicelles for photothermal and chemotherapy of drug-resistant tumors. The nanomicelles were fabricated with PDPA-based polymeric matrix, which could cause dissociation of the nanostructures under acid condition. Photosensitizer Ce6 was conjugated with PDPA nanomicelles and DOX was loaded into the nanomicelles. The developed nanomicelles illustrated strong light absorption in the NIR region. Under NIR laser irradiation, the nanomicelles could effectively convert optical energy into local heat, which promoted PA imaging, drug penetration into tumor tissues as well as photothermal therapy. Cai et al. [261] synthesized a diketopyrrolopyrrole (DPP)-triphenylamine (TPA) organic nanoagent for PA imaging-guided photodynamic/photothermal cancer therapy (Figure 13a). TPA, a commonly used donor, was conjugated with the DPP core to form a donor-acceptor- donor (D-A-D) structure. The nanoagent illustrated excellent NIR absorption, high photothermal conversion efficiency, and outstanding synergetic therapeutic effect due to the enhanced D-A-D structure of the DPP-TPA conjugation. Recently, Zhang et al. [262] developed electron donor-acceptor (D-A) conjugated semiconducting polymer nanoparticles (PPor-PEG NPs) for PA imaging-guided photothermal cancer therapy. The developed NPs could induce strong PA signals in tumor site for a long time, indicating the potential to be a long-term contrast agent. Additionally, the NPs showed a remarkedly high photothermal conversion efficiency 62.3%, the highest reported value among the reported polymer NPs.

Figure 13.

(a) D-A-D structured DPP-TPA NPs as theranostic agents for PA imaging-guided PDT/PTT. (b) Multimodality molecular imaging of MNPs. (c) Laser-generated-focused ultrasound-mediated drug delivery. (a) Reproduced with permission [261]. Copyright 2017, American Chemical Society. (b) Reproduced with permission [263]. Copyright 2014, American Chemical Society. (c) Reproduced with permission [248]. Copyright 2015, Elsevier.

Fan et al. [263] developed a novel multimodal imaging nanoplatform using ultra-small (~ 7.5 nm) water-soluble melanin nanoparticles (MNPs) (Figure 13b). The NPs not only demonstrated good photoacoustic properties for PA imaging but also illustrated natural binding ability with metal ions (Fe³⁺ and ⁶⁴Cu²⁺) for MRI and PET. Following that, Zhang et al. [264] developed a melanin nanoparticle-based nanosystem for PA imaging-guided chemotherapy. Anticancer drug, sorafenib (SRF), was bonded with melanin NPs to form a water-soluble nanoplatform, demonstrating great biocompatibility and PA property for imaging-guided therapy. The developed SRF-MNPs exhibited excellent tumor therapeutic effect after injection into the tail-vein of a living mice. Di et al. [248] recently developed an alginate sphere microgel encapsulated with DOX and CIF-loaded PLGA nanoparticles as a platform for laser ultrasound controlled drug delivery (Figure 13c). The drug release from the microgels was triggered by the laser generated pulsed ultrasound excitation that was transmitted from a laser-generated focused ultrasound transducer, made with a carbon black/PDMS-photoacoustic lens. Once the microgels were excited by the ultrasound waves, cavitation effects were induced at the microgels, which accelerated the release of drugs from the nanoparticles.

3.3.2. Inorganic nanocarriers

(1). Metallic Nanomaterials.

Numerous investigations have been demonstrated that metallic nanomaterials, especially AuNPs, can act as drug carriers for PA imaging-guided chemotherapy and PTT [215,265–267]. For instance, Wilson et al. [268] developed a photoacoustic nanodroplet composed of a liquid PFC core containing AuNRs and a bovine serum albumin shell for combinational PA/ultrasound imaging and potential drug delivery. The nanodroplets could provide PA and ultrasound contrast improvement via the laser-induced vaporization of the liquid PFC nanodroplet and long duration thermal expansion of the AuNRs. Following that, Zhong et al. [269] designed a paclitaxel (PTX)-containing nanoparticle which was based on loading PTX and AuNRs into PFH nanodroplets for PA imaging-guided photothermal/chemo synergistic cancer therapy. Upon pulsed laser irradiation, the nanoparticle could be rapidly decomposed owing to PFH vaporization, resulting in fast drug release, which induced efficient apoptosis of cancer cells. Duan et al. [270] developed organic/inorganic nanohybrids consisting of AuNRs, quantum dots (QDs), mesoporous silica layer, and CD-PGEA for multimodal imaging-guided cancer therapy (Figure 14a). The AuNRs were integrated with QDs for combined PA, CT and fluorescent imaging. In addition, the high photothermal conversion efficiency of AuNRs facilitated them to be utilized for photothermal therapy. Antitumor drug, DOX, was loaded into the mesoporous structure of the silica layer and sealed by the CD-PGEA gatekeeper for both the drug and gene delivery.

Figure 14.

(a) Preparation processes of multimodal imaging-guided therapeutic platforms. (b) Preparation of GNR @ IOs-DOX nanocapsules. (c) Loading the hollow interior of an AuNC with a dye doped PCM and then releasing t from the AuNC. (a) Reproduced with permission [270]. Copyright 2016, Wiley-VCH. (b) Reproduced with permission [271]. Copyright 2016, American Chemical Society. (c) Reproduced with permission [130]. Copyright 2011, American Chemical Society.

Besides coating AuNRs with mesoporous silica, Huang et al. [271] synthesized a hollow-mesoporous nanocapsule consisting of AuNRs and iron oxide nanoparticles (IOs) for dual-modal PA imaging and MRI-guided synergistic photothermal/chemo tumor treatment (Figure 14b). The AuNRs was combined with the magnetic IOs via a yolk-shell structure, in which movable AuNRs were confined in the permeable IO nanoshells using silica templates. DOX was loaded into the mesoporous nanocapsule after the formation of the yolk-shell structure. Moon et al. [130] designed a new theranostic system composed of hollow nanostructured Au nanocages (AuNCs) and a phase-change material (PCM) (Figure 14c). Drugs were encapsulated into the PCM, and the PCM was functionalized as a gatekeeper to control the drug release corresponding to temperature rise. Two dyes used as drug models were loaded into the PCM. Under HIFU irradiation, the PCM was melted and escaped from the interior of AuNCs, simultaneously releasing the encapsulated drugs.

Prussian blue, a clinical medicine approved by FDA, shows strong optical absorbance in the NIR region (650–900 nm) due to the charge transition between iron ions Fe²⁺ and Fe³⁺.

Recently, it has attracted interest for application in PA imaging and PTT [272–275]. Cai et al. [276] utilized hollow mesoporous Prussian blue NPs to develop a theranostic nanoplatform for ultrasound and PA dual-modal imaging-guided chemo-thermal cancer therapy. Anticancer drug, DOX, was loaded into the nanoparticles with relatively high loading capacity due to the mesoporous outer shell and big cavity. Since the drug loaded nanoparticles exhibited excellent photothermal conversion efficiency and high loading capacity, the Prussian blue-based nanoplatform opened a new avenue for chemo-thermal tumor therapy. Chen et al. [277] developed a dual-modal therapy platform fabricated by red blood cell membrane coated hollow mesoporous Prussian blue NPs for the combination of photothermal and chemotherapy. DOX was loaded into the NPs with a loading capacity up to 130%, resulting from the hollow mesoporous structure. Due to the photothermal effect, the DOX release from the NPs was promoted by a NIR laser irradiation. In addition, the acidic tumor microenvironments could also accelerate the DOX release from the NPs.

Apart from metallic nanoparticles, transition metal-based nanomaterials have been reported as theranostic nanoagents for PA imaging-guided therapy as well. Bao et al. [278] synthesized hollow structure PEG-MoO_3-x nanospheres for PA imaging-guided chemo-photothermal cancer therapy. The hollow nanospheres had strong LSPR absorption in the NIR region, good biocompatibility, as well as high photothermal conversion efficiency. In addition, due to their large surface area and porous structure, anticancer drug could be loaded into them. Song et al. [279] synthesized polyacrylic acid functionalized Co₉Se₈ nanoplates for PA imaging and MRI-guided chemo-photothermal combinational cancer therapy. The nanoplates demonstrated strong NIR absorption spectrum, T₂ shortening effect, and good photothermal conversion efficiency. Moreover, due to the large surface area to mass ratio, the nanoplates could also act as drug nanocarriers for pH-responsive chemotherapy.

(2). Carbon-based nanomaterials.

CNTs are also explored as drug delivery vehicles for cancer diagnostics and chemotherapies due to their unique properties. For example, Liu et al. [156] developed mesoporous silica (MS) coated SWNTs with polyethylene glycol (PEG) coating for imaging-guided cancer therapy. SWNTs provided strong contrast in both PA imaging and MRI. The PEG coating could improve the stability and solubility of the nanoparticles in a physiological environment. Anticancer drug, DOX, was loaded into the mesoporous structure of the synthesized SWNT@MS-PEG nanoparticle for chemotherapy. Using a low power of laser stimulation and a low dose of DOX, combined photothermal and chemotherapy of cancer tissues was achieved due to the accelerated nanoparticles penetration into cells and the light-triggered drug release.

rGO has an extremely high surface area and strong NIR absorption, and has very high potential to be utilized as a drug or gene delivery vehicle and as a photothermal agent for cancer therapy [280]. For example, Song et al. [166] conjugated DOX with rGO first and then encapsulated of rGO-DOX conjugation in a gold nanorod vesicle (rGO-AuNRVe-DOX) for dual chemo-photothermal therapy. With a relatively low power density of laser irradiation (0.25 W/cm²), the hybrid vesicle was disrupted and rGO-DOX and DOX was released from the vesicle due to the mild temperature rise in the tumor region with the vesicle injected. Moreover, the increased temperature also resulted in the improved intracellular uptake of DOX for chemotherapy.

3.3.3. Laser-generated ultrasound-accelerated thrombolysis

Due to its ability to generate high pressure and high frequency shock waves with tight focal spot, laser-generated ultrasound has recently been reported to be utilized for thrombolysis. Compared with existing thrombolysis techniques, the other advantages of laser-generated ultrasound includes its biocompatibility, low cost as well as high clot lysis rate. Kim et al. [281] firstly reported a laser-generated focused ultrasound (LGFU) transducer fabricated by a plano-concave glass lens coated with carbon black/PDMS thermoelastic layer. The transducer could generate shock waves with a central frequency of 14 MHz and peak negative pressure of ~ 12 MPa under a laser energy input of 20 mJ. Combined with 10⁸/mL microbubble injection (100 μL/min for 30 min), 29% reduction of blood clot mass was achieved using the LGFU transducer for treatment. Following that, Wu et al. [282] developed a laser-generated ultrasound transducer consisting of candle soot nanoparticles-PDMS composite on an optical fiber for intravascular thrombolysis. It was showed that the fiber-optic laser ultrasound transducer had a central frequency of 6.8 MHz and peak negative pressure of 1.6 MPa under a laser fluence of 1.5 mJ/cm². Under the microbubble-mediated thrombolysis test, 32% reduction of blood clot mass was achieved. Most recently, Wu et al. [283] fabricated another fiber-optic laser ultrasound transducer, having the same structure as reported in [282]. The transducer had a central frequency and peak negative pressure of 5.8 MHz and 3.7 MPa, respectively. At the same time of thrombolysis using the developed laser ultrasound transducer, the blood clot status was monitored using a 1064 nm laser source and a commercial linear array transducer based on PA imaging.

4. Conclusions and outlook

In this review, for the first time, the recent progress of nanoacoustics in biomedical imaging and therapy fields was comprehensively examined. The recent advancement made in exploring nanomaterials and nanostructures for biomedical ultrasound and photoacoustic imaging as well as ultrasound and photoacoustic therapies was reviewed in terms of nanomaterials, nanostructures and nanodevice implementations. Based on the literature survey, while many progresses have been made for translating basic scientific research outcomes into routine clinical applications, several obstacles hamper the advancement. Some recommendations for the development of nanoacoustics in biomedical imaging and therapy are outlined here.

Over the past decade, there’s an increasing demand for miniaturized acoustic devices with the growing market shares of mobile devices and internet of things. While conventional piezoelectric acoustic devices currently still dominate the market, there has been extensive studies exploring capacitive micromachined ultrasonic transducers (CMUTs) and piezoelectric micromachined ultrasonic transducers (PMUTs) for various applications, especially for biomedical imaging and therapy. Compared with conventional piezoelectric transducers, one of the key benefits of CMUTs and PMUTs is their easy integration with integrated circuits. Other advantages include their batch production with submicron accuracy and uniformity, low power consumption and compact size.

Many reports focusing on nanosized contrast agents for ultrasound as well as photoacoustic imaging have appeared during the past few years, but the investigations about contrast agents are still largely confined to the research laboratory. While many different kinds of nanosized contrast agent illustrate substantial advantages for imaging applications, the nanomaterials adopted usually possess shortcomings that need to be addressed, including relatively poor biocompatibility, requirements for large doses, low targeting efficiency, and difficulties in accurately quantifying agents. Clearly, many issues still exist and there remains a long road ahead for implementing nanosized contrast agents for clinical applications.

Ultrasound and photoacoustic-mediated drug delivery and therapy using various nanoparticles as drug delivery nanocarriers have demonstrated their great potentials for simultaneous biomedical imaging and synergistic chemotherapy and photoacoustic therapy. While the drug delivery systems fabricated by different nanoparticles are believed to be very promising for future therapeutics, especially oncotherapy, there still exist multiple challenges in the clinical translation of these nanoparticles. For example, although metallic NPs have been reported to have remarkable and tunable acoustic and optical properties, their long-term systemic cytotoxicity and biocompatibility have attracted much concerns from many researchers. In addition, in order to further advance ultrasound and photoacoustic-mediated therapeutic delivery strategies, developing a platform that can monitor the acoustic effect and quantify the drugs delivered into the target tissue in a real-time manner is considered to be very necessary and important. This is particularly crucial for tumor treatment since the tissue-related heterogeneity during drug delivery may bring about incomplete treatment and cancer recurrence.