Christophoros Mannaris

Dose-Dependent Artifact in the Far Wall of the Carotid Artery at Dynamic Contrast-enhanced US

PURPOSE To quantify a pseudoenhancement phenomenon observed during dynamic contrast material-enhanced ultrasonography (US) of the carotid artery, both in vitro and in vivo. MATERIALS AND METHODS Ethical approval was obtained prior to commencing this prospective case series, and each patient gave written informed consent. Thirty-one patients with 50%-99% internal carotid artery stenosis underwent dynamic contrast-enhanced US of the carotid bifurcation with use of 2 mL of microbubbles. In the final 10 patients, an additional 1 mL bolus was administered after 15 minutes. Raw linear digital imaging and communications in medicine data were analyzed offline. Regions of interest were drawn within the common carotid artery lumen and immediately adjacent to the lumen in the near and far wall adventitia. Peak intensity was measured. An in vitro experiment with a single-channel flow phantom was also performed. This apparatus consisted of an 8-mm-diameter latex tube placed in a tissue-mimicking fluid. Microbubble concentrations of 0.02%, 0.1%, 0.5%, 1%, and 2% were pumped into the tube. Regions of interest were drawn in a similar fashion to the in vivo experiments, and peak intensity was measured. The Wilcoxon signed rank and paired t tests were used to compare the difference between the near and far wall signal intensities at each dose; a multiplication factor comparing near and far wall signal intensity was derived. RESULTS The far wall of the common carotid artery was significantly more echogenic than the near wall at 2 mL contrast agent doses (P<.0001, n=31), and the far wall signal intensity increased synchronously with that of the lumen. The difference in signal intensity between near and far wall regions was significantly greater at 2 mL than at 1 mL (P=.012, n=10). In vitro, the phantom tubing demonstrated a similar pattern and magnitude of enhancement to that seen in vivo. CONCLUSION A dose-dependent, nonlinear propagation artifact known as pseudoenhancement occurs in the far wall adventitia of the carotid artery and should not be mistaken as a marker of plaque vulnerability.

Doxorubicin liposome-loaded microbubbles for contrast imaging and ultrasound-triggered drug delivery

Targeted drug delivery under image guidance is gaining more interest in the drug-delivery field. The use of microbubbles as contrast agents in diagnostic ultrasound provides new opportunities in noninvasive image-guided drug delivery. In the present study, the imaging and therapeutic properties of novel doxorubicin liposome-loaded microbubbles are evaluated. The results showed that at scanning settings (1.7 MHz and mechanical index 0.2), these microbubbles scatter sufficient signal for nonlinear ultrasound imaging and can thus be imaged in real time and be tracked in vivo. In vitro therapeutic evaluation showed that ultrasound at 1 MHz and pressures up to 600 kPa in combination with the doxorubicin liposomeloaded microbubbles induced 4-fold decrease of cell viability compared with treatment with free doxorubicin or doxorubicin liposome-loaded microbubbles alone. The therapeutic effectiveness is correlated to an ultrasound-triggered release of doxorubicin from the liposomes and an enhanced uptake of the free doxorubicin by glioblastoma cells. The results obtained demonstrate that the combination of ultrasound and the doxorubicin liposome-loaded microbubbles can provide a new method of noninvasive image-guided drug delivery.

Nonlinear pulsing schemes for the detection of ultrasound contrast agents

Ultrasound contrast agents are used in cardiology for the assessment of myocardial perfusion and in radiology for the detection and characterization of tumors. One widely used approach of imaging contrast agents is to use a low Mechanical Index (MI) nonlinear imaging technique to avoid bubble destruction and image both the macro‐ and micro‐circulation in real‐time. Various pulsing schemes are employed for the detection of nonlinear echoes from contrast microbubbles. The objective of this paper is to evaluate the various pulsing schemes for low MI imaging of contrast microbubbles and better understand their similarities and differences. The pulsing schemes considered are pulse inversion, power modulation, and their combination. Emphasis is placed on identifying whether nonlinearity due to propagation in tissue may be discriminated from nonlinearity due to scattering from bubbles. Bubble destruction (use of high MI) and tissue motion were not considered in this work. The evaluation of the different pulsing ...

Role of thermal and mechanical effects on drug release from thermosensitive nanocarriers

The combination of focused ultrasound (FUS) with thermosensitive liposomes (TSL) is a promising method for drug delivery since it allows a localized release upon moderate heating with ultrasound. Besides thermal effects, FUS also induces mechanical stresses on drug nanocarriers. We propose in this study to examine the influence of both effects (thermal and mechanical) on drug release. For this, an in-vitro setup allowing liposomal drug delivery using FUS was first evaluated. Calcein was used as a model drug. FUS experiments were performed in water at 37°C using a 1 MHz transducer focused at 48 mm, at 1 kHz PRF and 40% duty cycle. The driving pressure and the insonation duration were varied from 1 to 2 MPa and from 0 to 30 min, respectively. Thermal heating using a water-bath was also performed with temperatures from 37 to 49°C. For TSL, the release reaches a plateau above 42°C (45%) after 10 min heating while no release is observed for non-thermosensitive liposomes (NTSL). Using FUS, a rapid calcein release is observed for pressures from 1 to 1.5 MPa (from 0% to 49%) for TSL. Above 1.5 MPa, the release increases slightly (59% at 2 MPa). For NTSL, a weak calcein release is measured for acoustic pressures higher than 1.5 MPa. This release is attributed to the mechanical stress generated by FUS which is sufficient to destabilize the liposomal membrane. Mechanical stress alone can enhance the calcein release by up to 17% for pressures higher than 1.75 MPa.

Accurate measurement of microbubble response to ultrasound with a diagnostic ultrasound scanner

Ultrasound and microbubbles are often used to enhance drug delivery and the suggested mechanisms are extravasation and sonoporation. Drug delivery schemes with ultrasound and microbubbles at both low and high acoustic amplitudes have been suggested. A diagnostic ultrasound scanner may play a double role as both an imaging and a therapy device. It was not possible to accurately measure microbubble response with an ultrasound scanner for a large range of acoustic pressures and microbubble concentrations until now, mainly because of signal saturation issues. A method for continuously adjusting the receive gain of a scanner and limiting signal saturation was developed to accurately measure backscattered echoes from microbubbles for mechanical indexes (MIs) up to 2.1. The intensity of backscattered echoes from microbubbles increased quarticly with MI without reaching any limit. The signal intensity from microbubbles was found to be linear with concentration at both low and high MIs. However, at very high concentrations, acoustic shadowing occurs which limits the delivered acoustic pressure in deeper areas. The contrastto- tissue ratio was also measured and found to stay constant with MI. These results can be used to better guide drug delivery approaches and to also develop imaging techniques for therapy procedures.

Ultrasound-induced temperature elevation for in-vitro controlled release of temperature-sensitive liposomes

Drug loaded temperature-sensitive liposomes (TSLs) release their payload with mild hyperthermia near their phase transition temperature (Tm = 43–45 °C). Such a release may improve therapeutic efficacy and reduce toxic side effects in cancer treatment. In the present work, two different approaches are considered where focused ultrasound is used to induce the required temperature elevation for the release of doxorubicin from TSLs: (a) primary heating due to thermo-viscous absorption of ultrasound in absorptive media (oil, glycerol) and (b) secondary heating in non-absorptive media (blood, cell medium) due to heat transfer from the surroundings. Fine-wire thermocouple readings where in close agreement with theoretical predictions of temperature elevation with the Bioheat equation. Pulsing schemes to elevate and maintain the temperature at the desired value were designed with the Bioheat equation and validated with experiments. Fluorescence spectroscopy was used to assess the release of free doxorubicin that ...

Use of high intensity focused ultrasound for localized activation of thermosensitive liposomes for drug delivery.

Localized drug delivery holds great promise in improving drug efficacy in cancer treatment. Newly developed temperature‐sensitive liposomes (TSLs) loaded with doxorubicin have been shown to release their payload with mild hyperthermia near their phase transition temperature (Tm = 43–45 °C). In the present work, high intensity focused ultrasound is used to induce the required temperature elevation for the release of the drug from TSLs. A theoretical model based on Pennes’ bioheat equation was initially used to calculate the conditions for temperature elevation in fluids and tissue phantoms under conditions for drug activation. Acoustic pressures of 1–2 MPa at the focus with varying duty cycle (typically 50%) at 1 MHz frequency were calculated. Measurements of temperature rise were found in good agreement with our theoretical predictions. Fluorescence measurements were used to assess the release of free doxorubicin that exhibits higher fluorescence intensity than the liposomal formulation. In vitro experime...

Enhanced delivery of a density-modified therapeutic using ultrasound: Comparing the influence of micro- and nano-scale cavitation nuclei

New strategies are required to enhance the penetration and tumor-wide distribution of cancer therapeutics, including viruses, antibodies, and oligonucleotides. We have shown previously that increasing the density of a nanoparticle can enhance its ultrasound-mediated transport, particularly when exposed to microstreaming. The current study investigates how the physical characteristics and dynamics of different cavitation nucleation agents affect the cavitation-mediated transport of therapeutics not directly bound to the gas nuclei. SonoVue (Bracco, Milan, Italy) microbubble contrast agent (with 3 μm mean initial bubble diameter) and polymeric gas-entrapping nanoparticles (with 260 nm mean initial bubble diameter) were first compared in terms of inertial cavitation threshold and cavitation persistence. Under the same ultrasound exposure conditions (centre frequencies of either 0.5 MHz or 1.6 MHz, and peak negative pressures from 0.2 MPa to 3.5 MPa) their respective cavitation emissions were found to differ ...

Optimisation of ultrasound exposure parameters for extravasation and drug delivery using submicron cavitation nuclei

Sub-micron cavitation nuclei with the ability to passively extravasate through the leaky tumor vasculature have been shown to enhance both extravasation and penetration of pharmaceuticals in tumors. The aim of the present work is to optimize the acoustic parameters and maximize drug delivery mediated by cavitational microstreaming. Either lipid-shell microbubbles, or gas-stabilizing polymeric submicron cups of mean diameter 480 nm are co-administered with a model drug through a flow channel formed in agarose gel. The effect of ultrasound frequency, pressure amplitude, pulse duration, duty cycle, and pulse repetition frequency on the delivery of the drug is first evaluated, using both a single-layer and a dual-layer flow phantom that better represents the leaky tumor vasculature and dense extracellular matrix. The optimal parameters at 0.5 MHz and 1.6 MHz are then selected and used to deliver an oncolytic virus, genetically modified to express the RFP reporter gene, thus allowing the quantification of the ...

Image-guided sonoporation in an ex vivo machine perfused porcine liver

Sonoporation is the transient and reversible cell membrane permeability change induced with ultrasound and microbubbles. It allows for the uptake of normally impermeable macromolecules and has been suggested for improving drug delivery. The exact sonoporation mechanism and the optimal ultrasound parameters are still under investigation. Ex vivo machine perfused porcine livers are an excellent platform for investigating the sonoporation parameters and specifically the interaction of ultrasound driven microbubbles with the capillaries. Our objective was to identify the ultrasound parameters that are capable of causing detectable perfusion changes in the sonoporation area. Three types of perfusion changes were considered: large mechanical damage void of perfusion, reduced perfusion due to capillary destruction, and unaltered perfusion.