Influence of shell properties on high-frequency ultrasound imaging and drug delivery using polymer-shelled microbubbles

Abstract

This two-part study investigated shell rupture of ultrasound contrast agents (UCAs) under static overpressure conditions and the subharmonic component from UCAs subjected to 20-MHz tonebursts. Five different polylactide-shelled UCAs with shell-thickness-to-radius ratios (STRRs) of 7.5, 30, 40, 65, and 100 nm/¿m were subjected to static overpressure in a glycerol-filled test chamber. A video microscope imaged the UCAs as pressure varied from 2 to 330 kPa over 90 min. Images were postprocessed to obtain the pressure threshold for rupture and the diameter of individual microbubbles. Backscatter from individual UCAs was investigated by flowing a dilute UCA solution through a wall-less flow phantom placed at the geometric focus of a 20-MHz transducer. UCAs were subjected to 10- and 20-cycle tonebursts of acoustic pressures ranging from 0.3 to 2.3 MPa. A method based on singular-value decomposition (SVD) was employed to obtain a cumulative subharmonic score (SHS). Different UCA types exhibited distinctly different rupture thresholds that were linearly related to their STRR, but uncorrelated with UCA size. The rupture threshold for the UCAs with an STRR = 100 nm/μm was more than 4 times greater than the UCAs with an STRR = 7.5 nm/μm. The polymer-shelled UCAs produced substantial subharmonic response but the subharmonic response to 20- MHz excitation did not correlate with STRRs or UCA-rupture pressures. The 20-cycle excitation resulted in an SHS that was 2 to 3 times that of UCAs excited with 10-cycle tonebursts.

Fig. 1

(a) Schematic of the experimental setup for the overpressure study. Ultrasound contrast agents were subjected to overpressure in a glycerol-filled, cylindrical test chamber while being imaged with a video microscope. (b) A sample image and (c)–(e) illustration of the steps involved in the image-processing algorithm.

Fig. 2

Schematic of the subharmonic-study experimental setup. A dilute solution of ultrasound contrast agents (UCAs) was injected into a test tank via a 200-μm needle. The focus of a 20-MHz transducer was aligned 2 mm downstream. Individual UCAs were subjected to 20-MHz tonebursts and backscatter signals were recorded.

Fig. 3

Histograms (12 bins) of ultrasound contrast agent (UCA) (a) diameter and (b) rupture pressure. Each UCA population consisted of at least 300 UCAs. All five Philips UCA populations exhibited a Gaussian size distribution centered around the nominal mean diameter of 2 μm; the Point UCA population exhibited a polydisperse size distribution that was centered around its nominal mean diameter of 3.4 μm. Rupturepressure histograms indicated that different UCA populations exhibited different rupture thresholds. The Point UCAs had the lowest threshold for rupture; the Philips UCAs with a shell-thickness-to-radius ratio (STRR) of 100 nm/μm had the highest rupture threshold. Only 50% of these UCAs ruptured when subjected to up to 330 kPa of overpressure. UCAs that did not rupture at overpressures below 330 kPa are categorized as NR.

Fig. 4

Mean ultrasound contrast agent (UCA)-rupture pressure with respect to shell-thickness-to-radius ratio (STRR). The box plot was obtained by performing one-way analysis of variance (ANOVA). The mean rupture pressure for each UCA population was statistically different from mean rupture pressures of other UCAs (indicated by *). The rupture pressure was linearly related to STRR.

Fig. 5

Representative waveforms of the backscattered ultrasound in response to (a) 10-cycle and (b) 20-cycle excitations. The envelope of the signals highlights the subharmonic component. (c) and (d) Corresponding power spectra. Substantial subharmonic activity was detected in many of the backscatter events.

Fig. 6

(a)–(e) Singular-value decomposition (SVD)-derived subharmonic score (SHS) for each ultrasound contrast agent (UCA) population for (dashed line) 10-cycle excitation and (solid line) 20-cycle excitation. Each data point represents a mean of SHS values obtained from six 500-UCA data sets. The error bars indicate the standard deviation. For all UCAs, the 20-cycle excitation produced greater values of SHS than the 10-cycle excitation. The Point UCAs (a) produced the lowest SHS, indicating relatively weak and infrequent subharmonic events. The Point SHS indicated that peak subharmonic activity occurred at approximately 0.8 MPa for both pulse durations. All four Philips UCAs exhibited significantly higher values of SHS than the Point UCA. In the pressure range that was investigated, none of the Phillips UCAs indicated a peak of SHS. The subharmonic onset was estimated from the pressure required to reach an SHS of 10 times the initial value by fitting the curves to a power law and are indicated by the vertical black lines.

Fig. 7

Diameter of the ultrasound-induced destruction zone of the polymer-shelled ultrasound contrast agents (UCAs) with respect to incident acoustic pressure. (Data for the figure courtesy of Böhmer et al. [47].) The damage area increased with increase in incident pressure, but decreased with an increase in shell-thickness-to-radius ratio (STRR).

Fig. 8

Theoretically predicted scattering cross-section of the ultrasound contrast agent (UCA) populations for two cases: (a) nominal shell thickness inferred using shell-thickness-to-radius ratio (STRR) and the measured mean UCA size, i.e., an intact, homogenous UCA; and (b) a shell of effective bending thickness estimated by fitting the overpressure-rupture data to the Ru model (3), i.e., an inhomogeneous UCA with shell imperfections. The UCA parameters employed for these calculations are presented in Table I. The resonance frequencies of the Philips UCAs obtained for the homogenous shell are up to 4 times greater than the excitation frequency of 20 MHz. When estimated using the inhomogeneous shell, the resonance frequencies shift close to 20 MHz. The resonance frequency of the homogenous Point UCAs was estimated to be 24 MHz, which indicated that the Point UCA might require a lower acoustic pressure to induce nonlinear oscillations.