Ultrafast Background-Free Ultrasound Imaging Using Blinking Nanoparticles.

Localization-based ultrasound imaging methods that use microbubbles or nanodroplets offer high-resolution imaging with improved sensitivity and reduced background signal. However, these methods require long acquisition times (typically seconds to minutes), preventing their use for real-time imaging and, thus, limiting their clinical translational potential. Here, we present a new ultrafast localization method using blinking ultrasound-responsive nanoparticles (BNPs). When activated with high frame rate (1 kHz) plane wave ultrasound pulses with a mechanical index of 1.5, the BNPs incept growth of micrometer-sized bubbles, which in turn collapse and generate a blinking ultrasound signal. We showed that background-free ultrasound images could be obtained by localizing these blinking events using acquisition times as low as 11 ms. In addition, we demonstrated that BNPs enable in vivo background-free ultrasound imaging in mice. We envision that BNPs will facilitate the clinical translation of localization-based ultrasound imaging for more sensitive detection of cancer and other diseases.

In Silico Screening and Testing of FDA-Approved Small Molecules to Block SARS-CoV-2 Entry to the Host Cell by Inhibiting Spike Protein Cleavage.

The COVID-19 pandemic began in 2019, but it is still active. The development of an effective vaccine reduced the number of deaths; however, a treatment is still needed. Here, we aimed to inhibit viral entry to the host cell by inhibiting spike (S) protein cleavage by several proteases. We developed a computational pipeline to repurpose FDA-approved drugs to inhibit protease activity and thus prevent S protein cleavage. We tested some of our drug candidates and demonstrated a decrease in protease activity. We believe our pipeline will be beneficial in identifying a drug regimen for COVID-19 patients.

Affinity Proteomics Identifies Interaction Partners and Defines Novel Insights into the Function of the Adhesion GPCR VLGR1/ADGRV1.

The very large G-protein-coupled receptor 1 (VLGR1/ADGRV1) is the largest member of the adhesion G-protein-coupled receptor (ADGR) family. Mutations in VLGR1/ADGRV1 cause human Usher syndrome (USH), a form of hereditary deaf-blindness, and have been additionally linked to epilepsy. In the absence of tangible knowledge of the molecular function and signaling of VLGR1, the pathomechanisms underlying the development of these diseases are still unknown. Our study aimed to identify novel, previously unknown protein networks associated with VLGR1 in order to describe new functional cellular modules of this receptor. Using affinity proteomics, we have identified numerous new potential binding partners and ligands of VLGR1. Tandem affinity purification hits were functionally grouped based on their Gene Ontology terms and associated with functional cellular modules indicative of functions of VLGR1 in transcriptional regulation, splicing, cell cycle regulation, ciliogenesis, cell adhesion, neuronal development, and retinal maintenance. In addition, we validated the identified protein interactions and pathways in vitro and in situ. Our data provided new insights into possible functions of VLGR1, related to the development of USH and epilepsy, and also suggest a possible role in the development of other neuronal diseases such as Alzheimer's disease.

Ultrasonographic evaluation of distal femoral and talar cartilage thicknesses in patients with early rheumatoid arthritis and their relationship with disease activity.

INTRODUCTION: The aims of this study were to compare the distal femoral and talar cartilage thicknesses by ultrasonography (US) between patients with early rheumatoid arthritis (RA) and healthy controls and to investigate the potential relationship of these parameters with disease activity and clinical parameters. METHODS: Fifty-five patients with RA with a disease duration of up to 12 months (RA group), and 55 healthy volunteers (control group) were included in the study. The bilateral distal femur and talus cartilage thicknesses of all participants were measured by US and compared between the groups. The relationship of the results with the clinical data of the patients with early RA was investigated. RESULTS: The distal femur (medial, central, and lateral) and talus cartilage thicknesses of both extremities were significantly lower in the RA group. The tender joint count was found to be slightly to moderately negatively correlated with cartilage thickness. However, no correlation was found between the swollen joint count and cartilage thickness. In addition, the disease activity scores were moderately and inversely correlated with cartilage thickness in certain areas. CONCLUSION: The results of this preliminary study using US indicate that the distal femoral and talus cartilage of patients with RA can be affected in the early period, and the cartilage thickness of patients seems to be associated with disease activity. These results may inspire new studies for the evaluation of femoral and talar cartilage with US. Key Points • The distal femoral and talus cartilage thicknesses seem to be affected in the early period of rheumatoid arthritis. • There may be a relationship between the distal femur and talus cartilage thicknesses and disease activity in patients with early rheumatoid arthritis. • The tender joint count appears to be more associated with cartilage thickness than the swollen joint count in patients with early rheumatoid arthritis.

Gas-stabilizing nanoparticles for ultrasound imaging and therapy of cancer.

The use of ultrasound in the clinic has been long established for cancer detection and image-guided tissue biopsies. In addition, ultrasound-based methods have been widely explored to develop more effective cancer therapies such as localized drug delivery, sonodynamic therapy, and focused ultrasound surgery. Stabilized fluorocarbon microbubbles have been in use as contrast agents for ultrasound imaging in the clinic for several decades. It is also known that microbubble cavitation could generate thermal, mechanical, and chemical effects in the tissue to improve ultrasound-based therapies. However, the large size, poor stability, and short-term cavitation activity of microbubbles limit their applications in cancer imaging and therapy. This review will focus on an alternative type of ultrasound responsive material; gas-stabilizing nanoparticles, which can address the limitations of microbubbles with their nanoscale size, robustness, and high cavitation activity. This review will be of interest to researchers who wish to explore new agents to develop improved methods for molecular ultrasound imaging and therapy of cancer.

YouTube videos as a source of information concerning Behçet's disease: a reliability and quality analysis.

In this study, we aimed to investigate the reliability and quality of YouTube videos concerning Behçet disease, for the first time in the literature. The most viewed 50 YouTube videos about Behçet disease were included. Study data used in the analysis included video length, image type, video content, uploaders, time since the date of upload, the number of total views, daily views, and the numbers of like, dislike and comments. The Video Power Index was used to determine popularity of the videos. Reliability and quality of the videos were evaluated using DISCERN and Global Quality Scale (GQS) instruments. The examined YouTube videos were divided into two groups as uploaded by professionals (physicians, health and hospital channels, institutions and societies) and non-professionals (patients, social organizations, others). Twenty-three (46%) videos were uploaded by professionals. The mean DISCERN score was found as 3.98 ± 0.77 in the videos uploaded by healthcare professionals and 2.83 ± 0.64 in those uploaded by non-professionals (p < 0.01). The mean GQS score was found as 4.09 ± 0.72 in the videos uploaded by healthcare professionals and 3.41 ± 0.69 in those uploaded by non-professionals (p < 0.01). 46% of the videos were of low-to-moderate reliability and 56% of low-to-moderate quality. Nearly half of the YouTube videos concerning Behçet disease were uploaded by non-physician sources with poor reliability and quality. Physicians should be encouraged to upload more professional videos to help and guide patients appropriately. YouTube videos should be subjected to an objective evaluation for content quality before they can be published.

SANS (USH1G) regulates pre-mRNA splicing by mediating the intra-nuclear transfer of tri-snRNP complexes.

Splicing is catalyzed by the spliceosome, a compositionally dynamic complex assembled stepwise on pre-mRNA. We reveal links between splicing machinery components and the intrinsically disordered ciliopathy protein SANS. Pathogenic mutations in SANS/USH1G lead to Usher syndrome-the most common cause of deaf-blindness. Previously, SANS was shown to function only in the cytosol and primary cilia. Here, we have uncovered molecular links between SANS and pre-mRNA splicing catalyzed by the spliceosome in the nucleus. We show that SANS is found in Cajal bodies and nuclear speckles, where it interacts with components of spliceosomal sub-complexes such as SF3B1 and the large splicing cofactor SON but also with PRPFs and snRNAs related to the tri-snRNP complex. SANS is required for the transfer of tri-snRNPs between Cajal bodies and nuclear speckles for spliceosome assembly and may also participate in snRNP recycling back to Cajal bodies. SANS depletion alters the kinetics of spliceosome assembly, leading to accumulation of complex A. SANS deficiency and USH1G pathogenic mutations affects splicing of genes related to cell proliferation and human Usher syndrome. Thus, we provide the first evidence that splicing dysregulation may participate in the pathophysiology of Usher syndrome.

Determination of the antioxidant, antimicrobial and anticancer properties of the honey phenolic extract of five different regions of Bingöl province.

Bingol is a famous region for honey in Turkey. The amount of phenolic substance is also considered important for the anticancer, antioxidant and antimicrobial properties of honey. Anticancer activity of honey extract was determined as the most effective dose of 1 mg/mL using the WST1 anti-proliferation kit in the PC-3 cell line. Total phenol content were found between 476.09 ± 1.67 and 865.22 ± 3.57 mg GAE/100 g honey, total flavonoid content 41.67 ± 0.25 and 1249.74 ± 0.85 mg QE/100 g honey, total phenolic acid content 0.74 ± 0.21 and 58.35 ± 1.56 mg SA/100 g honey, ß-carotene 1.71 ± 0.06-3.61 ± 0.08 mg/kg honey, lycopene content 0.89 ± 0.03 and 3.41 ± 0.08 mg/kg honey, respectively. Percent removal of H2O2 was determined in the range of 69.79 ± 1.24 and 75.37 ± 1.72 at 10 mg/mL. DPPH percentage removal for 200 mg/mL was between 69.79 ± 1.24 and 75.37 ± 1.72. Phenolic compounds in honey extract were determined as gallic acid, caffeic acid, syringe acid, chlorogenic acid, p-coumaric acid, ferulic acid, catechin, quercetin, chrysin using HPLC analysis. The honey extracts were tested on Gram(+) and Gram(-) bacteria and yeast and their antimicrobial effects were determined. As a result, phenolic honey extraction, performed from five loci from the region of interest, showed anticancer, antioxidant and antimicrobial properties, and can be used as a functional food additive to replace synthetic counterparts.

Drug repurposing using transcriptome sequencing and virtual drug screening in a patient with glioblastoma.

Background Precision medicine and drug repurposing are attractive strategies, especially for tumors with worse prognosis. Glioblastoma is a highly malignant brain tumor with limited treatment options and short survival times. We identified novel BRAF (47-438del) and PIK3R1 (G376R) mutations in a glioblastoma patient by RNA-sequencing. Methods The protein expression of BRAF and PIK3R1 as well as the lack of EGFR expression as analyzed by immunohistochemistry corroborated RNA-sequencing data. The expression of additional markers (AKT, SRC, mTOR, NF-κB, Ki-67) emphasized the aggressiveness of the tumor. Then, we screened a chemical library of > 1500 FDA-approved drugs and > 25,000 novel compounds in the ZINC database to find established drugs targeting BRAF47-438del and PIK3R1-G376R mutated proteins. Results Several compounds (including anthracyclines) bound with higher affinities than the control drugs (sorafenib and vemurafenib for BRAF and PI-103 and LY-294,002 for PIK3R1). Subsequent cytotoxicity analyses showed that anthracyclines might be suitable drug candidates. Aclarubicin revealed higher cytotoxicity than both sorafenib and vemurafenib, whereas idarubicin and daunorubicin revealed higher cytotoxicity than LY-294,002. Liposomal formulations of anthracyclines may be suitable to cross the blood brain barrier. Conclusions In conclusion, we identified novel small molecules via a drug repurposing approach that could be effectively used for personalized glioblastoma therapy especially for patients carrying BRAF47-438del and PIK3R1-G376R mutations.

Gas-Stabilizing Sub-100 nm Mesoporous Silica Nanoparticles for Ultrasound Theranostics.

Recent studies have demonstrated that gas-stabilizing particles can generate cavitating micron-sized bubbles when exposed to ultrasound, offering excellent application potential, including ultrasound imaging, drug delivery, and tumor ablation. However, the majority of the reported gas-stabilizing particles are relatively large (>200 nm), and smaller particles require high acoustic pressures to promote cavitation. Here, this paper reports the preparation of sub-100 nm gas-stabilizing nanoparticles (GSNs) that can initiate cavitation at low acoustic intensities, which can be delivered using a conventional medical ultrasound imaging system. The highly echogenic GSNs (F127-hMSN) were prepared by carefully engineering the surfaces of ∼50 nm mesoporous silica nanoparticles. It was demonstrated that the F127-hMSNs could be continuously imaged with ultrasound in buffer or biological solutions or agarose phantoms for up to 20 min. Also, the F127-hMSN can be stored in phosphate-buffered saline for at least a month with no loss in ultrasound responsiveness. The particles significantly degraded when diluted in simulated body fluids, indicating possible biodegradation of the F127-hMSNs in vivo. Furthermore, at ultrasound imaging conditions, F127-hMSNs did not cause detectable cell death, supporting the potential safety of these particles. Finally, strong cavitation activity generation by the F127-hMSNs under high-intensity focused ultrasound insonation was demonstrated and applied to effectively ablate cancer cells.

PCARE and WASF3 regulate ciliary F-actin assembly that is required for the initiation of photoreceptor outer segment disk formation.

The outer segments (OS) of rod and cone photoreceptor cells are specialized sensory cilia that contain hundreds of opsin-loaded stacked membrane disks that enable phototransduction. The biogenesis of these disks is initiated at the OS base, but the driving force has been debated. Here, we studied the function of the protein encoded by the photoreceptor-specific gene C2orf71, which is mutated in inherited retinal dystrophy (RP54). We demonstrate that C2orf71/PCARE (photoreceptor cilium actin regulator) can interact with the Arp2/3 complex activator WASF3, and efficiently recruits it to the primary cilium. Ectopic coexpression of PCARE and WASF3 in ciliated cells results in the remarkable expansion of the ciliary tip. This process was disrupted by small interfering RNA (siRNA)-based down-regulation of an actin regulator, by pharmacological inhibition of actin polymerization, and by the expression of PCARE harboring a retinal dystrophy-associated missense mutation. Using human retinal organoids and mouse retina, we observed that a similar actin dynamics-driven process is operational at the base of the photoreceptor OS where the PCARE module and actin colocalize, but which is abrogated in Pcare-/- mice. The observation that several proteins involved in retinal ciliopathies are translocated to these expansions renders it a potential common denominator in the pathomechanisms of these hereditary disorders. Together, our work suggests that PCARE is an actin-associated protein that interacts with WASF3 to regulate the actin-driven expansion of the ciliary membrane at the initiation of new outer segment disk formation.

SANS (USH1G) Molecularly Links the Human Usher Syndrome Protein Network to the Intraflagellar Transport Module by Direct Binding to IFT-B Proteins.

The human Usher syndrome (USH) is a retinal ciliopathy, characterized by profound congenital deafness, variable vestibular dysfunction and pre-pubertal onset of retinitis pigmentosa. In the effected sensory cells, USH protein networks are assumed to function in ciliary transport processes. The USH1G protein SANS is a scaffold of the ciliary/periciliary USH protein network of photoreceptor cells. Moreover, SANS is associated with microtubules, the transport routes for protein delivery toward the cilium. To enlighten the role of SANS in ciliary transport processes, we aimed to identify transport related proteins associated with SANS. The intraflagellar transport (IFT) system is a conserved mechanism for bi-directional transport toward and through primary cilia. Thus, we tested the direct binding of SANS to IFT molecules, namely IFT20, IFT57, and IFT74 in 1:1 yeast-two-hybrid assay. The identified SANS-IFT interactions were validated in vitro via independent complementary interaction assays and in cells by applying membrane targeting assays. Quantitative immunofluorescence microscopy revealed the co-localization of SANS with IFT20, IFT52, and IFT57 particularly at ciliary base of wild type mouse photoreceptor cells. Analysis of photoreceptor cells of SANS knock out mice revealed the decrease of IFTs in the ciliary compartment indicating a role of SANS in the proper positioning of IFT-B molecules in primary cilia. Our study demonstrated direct binding of IFT complex B proteins IFT52 and IFT57 to the N-terminal ankyrin repeats and the central domain of SANS. Our data also indicate that pathologic mutations in the N-terminus of SANS lead to the loos of SANS binding to IFT-B molecules. Our findings provide direct evidence for a molecular link between the ciliary USH protein network and the IFT transport module in primary cilia.

Colloids, nanoparticles, and materials for imaging, delivery, ablation, and theranostics by focused ultrasound (FUS).

This review focuses on different materials and contrast agents that sensitize imaging and therapy with Focused Ultrasound (FUS). At high intensities, FUS is capable of selectively ablating tissue with focus on the millimeter scale, presenting an alternative to surgical intervention or management of malignant growth. At low intensities, FUS can be also used for other medical applications such as local delivery of drugs and blood brain barrier opening (BBBO). Contrast agents offer an opportunity to increase selective acoustic absorption or facilitate destructive cavitation processes by converting incident acoustic energy into thermal and mechanical energy. First, we review the history of FUS and its effects on living tissue. Next, we present different colloidal or nanoparticulate approaches to sensitizing FUS, for example using microbubbles, phase-shift emulsions, hollow-shelled nanoparticles, or hydrophobic silica surfaces. Exploring the science behind these interactions, we also discuss ways to make stimulus-responsive, or "turn-on" contrast agents for improved selectivity. Finally, we discuss acoustically-active hydrogels and membranes. This review will be of interest to those working in materials who wish to explore new applications in acoustics and those in acoustics who are seeking new agents to improve the efficacy of their approaches.

Temperature-Responsive Hydrophobic Silica Nanoparticle Ultrasound Contrast Agents Directed by Phospholipid Phase Behavior.

In this paper, we report ultrasonically active nanoscale contrast agents that behave as thermometric sensors through phase change in their stabilizing phospholipid monolayer. Phospholipid-stabilized, hydrophobic mesoporous silica nanoparticles (P@hMSNs) are known to interact with high-intensity focused ultrasound (HIFU) to promote cavitation at their surfaces, which can be used for both imaging and therapy. We show that the lateral lipid phase behavior of the phosphocholine lipid dictates the acoustic contrast of the P@hMSNs. When the lipids are in the gel phase below their melting temperature, the P@hMSNs generate detectable microbubbles when exposed to HIFU. However, if the lipids exhibit a liquid expanded phase, the P@hMSNs cease to generate bubbles in response to HIFU insonation. We verify that the heating and subsequent transition of lipid coating the hMSN are associated with the loss of acoustic response by doping laurdan dye into the lipid monolayer and imaging lipid phase through red shifts in emission spectra. Similarly, cessation of cavitation was also induced by adding a fluidizing surfactant such as Triton X, which could be reversed upon washing away the excess surfactant. Finally, by controlling for the partial fluidization caused by the adsorption of protein, P@hMSNs may be used as thermometric sensors of the bulk fluid temperature. These findings not only impact the utilization of nanoscale agents as stimulus-responsive ultrasound contrast agents but also have broader implications for how cavitation may be initiated at surfaces coated by a surfactant.

Epicardial adipose tissue thickness as a new risk factor for atherosclerosis in patients with ankylosing spondylitis.

PURPOSE: The aim of this study was to compare epicardial adipose tissue thickness (EATT), which is a novel cardiometabolic risk factor in patients with ankylosing spondylitis (AS), and carotid intima media thickness (CIMT) with healthy controls to assess the relationship between these markers and disease activity. MATERIALS AND METHODS: This study involved 38 patients with AS and 38 controls with no history of cardiovascular disease. CIMT was measured by Doppler ultrasound, and EATT by echocardiography, in all participants. Total cholesterol (TC), low density lipoprotein (LDL) cholesterol, high density lipoprotein cholesterol, triglycerides, thyroid stimulating hormone, erythrocyte sedimentation rate, C-reactive protein, and hemogram measurements were performed in all subjects after 8 hours of fasting. In addition, the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and the Bath Ankylosing Spondylitis Functional Index (BASFI) scores of the patients with AS were recorded. RESULTS: EATT was significantly higher in patients with AS (0.45 ± 0.17 mm) compared with healthy (0.37 ± 0.10 mm) controls (p= 0.01). Significant positive correlations were detected between EAT thickness and BASFI, CIMT, and disease duration in patients with AS (p= 0.008, p= 0.024, and p= 0.012, respectively). Significant negative correlations were observed between EATT and TC and LDL cholesterol concentrations (p= 0.016 and p= 0.009, respectively). No significant difference was detected in CIMT between the patients (0.515 ± 0.08 mm) and the controls (0.517 ± 0.094 mm, p= 0.98). No significant correlation was observed between CIMT of the patients with AS and the BASDAI, BASFI, TC, HDL cholesterol, systolic blood pressure, or diastolic blood pressure values. Significant positive correlations were observed between CIMT and waist circumference, weight, body mass index, and LDL cholesterol in patients with AS and the controls. CONCLUSIONS: This study demonstrated increased EATT in patients with AS compared with a healthy population, which was positively correlated with disease duration and BASFI.

Nanoparticle-Mediated Acoustic Cavitation Enables High Intensity Focused Ultrasound Ablation Without Tissue Heating.

While thermal ablation of various solid tumors has been demonstrated using high intensity focused ultrasound (HIFU), the therapeutic outcomes of this technique are still unsatisfactory because of common recurrence of thermally ablated cancers and treatment side effects due to the high ultrasound intensity and acoustic pressure requirements. More precise ablation of tumors can be achieved by generating cavitating bubbles in the tissue using shorter pulses with higher acoustic pressures, which induce mechanical damage rather than thermal. However, it has remained as a challenge to safely deliver the acoustic pressures required for mechanical ablation of solid tumors. Here, we report a method to achieve mechanical ablation at lower acoustic pressures by utilizing phospholipid-stabilized hydrophobic mesoporous silica nanoparticles (PL-hMSN). The PL-hMSNs act as seeds for nucleation of cavitation events and thus significantly reduce the peak negative pressures and spatial-average temporal-average HIFU intensities needed to achieve mechanical ablation. Substantial mechanical damage was observed in the red blood cell or tumor spheroid containing tissue mimicking phantoms at PL-hMSN concentrations as low as 10 µg mL-1, after only 5 s of HIFU treatment with peak negative pressures ∼11 MPa and duty cycles ∼0.01%. Even the application of HIFU (peak negative pressure of 16.8 MPa and duty cycle of 0.017%) for 1 min in the presence of PL-hMSN (200 µg mL-1) did not cause any detectable temperature increase in tissue-mimicking phantoms. In addition, the mechanical effects of cavitation promoted by PL-hMSNs were observed up to 0.5 mm from the center of the cavitation events. This method may thus also improve delivery of therapeutics or nanoparticles to tumor environments with limited macromolecular transport.

Analysis of centrosome and DNA damage response in PLK4 associated Seckel syndrome.

Microcephalic primordial dwarfism (MPD) is a group of autosomal recessive inherited single-gene disorders with intrauterine and postnatal global growth failure. Seckel syndrome is the most common form of the MPD. Ten genes are known with Seckel syndrome. Using genome-wide SNP genotyping and homozygosity mapping we mapped a Seckel syndrome gene to chromosomal region 4q28.1-q28.3 in a Turkish family. Direct sequencing of PLK4 (polo-like kinase 4) revealed a homozygous splicing acceptor site transition (c.31-3 A>G) that results in a premature translation termination (p.[=,Asp11Profs*14]) causing deletion of all known functional domains of the protein. PLK4 is a master regulator of centriole biogenesis and its deficiency has recently been associated with Seckel syndrome. However, the role of PLK4 in genomic stability and the DNA damage response is unclear. Evaluation of the PLK4-Seckel fibroblasts obtained from patient revealed the expected impaired centriole biogenesis, disrupted mitotic morphology, G2/M delay, and extended cell doubling time. Analysis of the PLK4-Seckel cells indicated that PLK4 is also essential for genomic stability and DNA damage response. These findings provide mechanistic insight into the pathogenesis of the severe growth failure associated with PLK4-deficiency.

Phospholipid Capped Mesoporous Nanoparticles for Targeted High Intensity Focused Ultrasound Ablation.

The mechanical effects of cavitation can be effective for therapy but difficult to control, thus potentially leading to off-target side effects in patients. While administration of ultrasound active agents such as fluorocarbon microbubbles and nanodroplets can locally enhance the effects of high intensity focused ultrasound (HIFU), it has been challenging to prepare ultrasound active agents that are small and stable enough to accumulate in tumors and internalize into cancer cells. Here, this paper reports the synthesis of 100 nm nanoparticle ultrasound agents based on phospholipid-coated, mesoporous, hydrophobically functionalized silica nanoparticles that can internalize into cancer cells and remain acoustically active. The ultrasound agents produce bubbles when subjected to short HIFU pulses (≈6 µs) with peak negative pressure as low as ≈7 MPa and at particle concentrations down to 12.5 µg mL-1 (7 × 109 particles mL-1 ). Importantly, ultrasound agents are effectively uptaken by cancer cells without cytotoxic effects, but HIFU insonation causes destruction of the cells by the acoustically generated bubbles, as demonstrated by (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) and lactate dehydrogenase assays and flow cytometry. Finally, it is showed that the HIFU dose required to effectively eliminate cancer cells in the presence of ultrasound agents causes only a small temperature increase of ≈3.5 °C.

Nanoparticles Formed by Acoustic Destruction of Microbubbles and Their Utilization for Imaging and Effects on Therapy by High Intensity Focused Ultrasound.

This work reports that when PEG-lipid-shelled microbubbles with fluorocarbon interior (C4F10, C5F12, or C6F14) are subjected to ultrasound pulses, they produce metastable, fluid-filled nanoparticles that can be re-imaged upon administration of HIFU. The nanoparticles produced by destruction of the microbubbles (MBNPs) are of 150 nm average diameter and can be re-imaged for up to an hour after creation for C 4F10, and for at least one day for C5F12. The active species were found to be fluid (gas or liquid) filled nanoparticles rather than lipid debris. The acoustic droplet vaporization threshold of the nanoparticles was found to vary with the vapor pressure of the encapsulated fluorocarbon, and integrated image brightness was found to increase dramatically when the temperature was raised above the normal boiling point of the fluorocarbon. Finally, the vaporization threshold decreases in serum as compared to buffer, and administration of HIFU to the nanoparticles caused breast cancer cells to completely detach from their culture substrate. This work demonstrates a new functionality of microbubbles that could serve as a platform technology for ultrasound-based theranostics.