Jinhyoung Park

Multimodality Imaging In Vivo for Preclinical Assessment of Tumor-Targeted Doxorubicin Nanoparticles

This study presents a new multimodal imaging approach that includes high-frequency ultrasound, fluorescence intensity, confocal, and spectral imaging to improve the preclinical evaluation of new therapeutics in vivo. Here we use this approach to assess in vivo the therapeutic efficacy of the novel chemotherapy construct, HerDox during and after treatment. HerDox is comprised of doxorubicin non-covalently assembled in a viral-like particle targeted to HER2+ tumor cells, causing tumor cell death at over 10-fold lower dose compared to the untargeted drug, while sparing the heart. Whereas our initial proof-of-principle studies on HerDox used tumor growth/shrinkage rates as a measure of therapeutic efficacy, here we show that multimodal imaging deployed during and after treatment can supplement traditional modes of tumor monitoring to further characterize the particle in tissues of treated mice. Specifically, we show here that tumor cell apoptosis elicited by HerDox can be monitored in vivo during treatment using high frequency ultrasound imaging, while in situ confocal imaging of excised tumors shows that HerDox indeed penetrated tumor tissue and can be detected at the subcellular level, including in the nucleus, via Dox fluorescence. In addition, ratiometric spectral imaging of the same tumor tissue enables quantitative discrimination of HerDox fluorescence from autofluorescence in situ. In contrast to standard approaches of preclinical assessment, this new method provides multiple/complementary information that may shorten the time required for initial evaluation of in vivo efficacy, thus potentially reducing the time and cost for translating new drug molecules into the clinic.

A high-frequency annular-array transducer using an interdigital bonded 1-3 composite

This paper reports the design, fabrication, and characterization of a 1-3 composite annular-array transducer. An interdigital bonded (IB) 1-3 composite was prepared using two IB operations on a fine-grain piezoelectric ceramic. The final composite had 19-μm-wide posts separated by 6-μm-wide polymer kerfs. A novel method to remove metal electrodes from polymer portions of the 1-3 composite was established to eliminate the need for patterning and aligning the electrode on the composite to the electrodes on a flexible circuit. Unloaded epoxy was used for both the matching and backing layers and a flexible circuit was used for interconnect. A prototype array was successfully fabricated and tested. The results were in reasonable agreement with those predicted by a circuit-analogous model. The average center frequency estimated from the measured pulse-echo responses of array elements was 33.5 MHz and the -6-dB fractional bandwidth was 57%. The average insertion loss recorded was 14.3 dB, and the maximum crosstalk between the nearest-neighbor elements was less than -37 dB. Images of a wire phantom and excised porcine eye were obtained to show the capabilities of the array for high-frequency ultrasound imaging.

Hemodynamics and ventricular function in a zebrafish model of injury and repair.

Myocardial infarction results in scar tissue and irreversible loss of ventricular function. Unlike humans, zebrafish has the capacity to remove scar tissue after injury. To assess ventricular function during repair, we synchronized microelectrocardiogram (μECG) signals with a high-frequency ultrasound pulsed-wave (PW) Doppler to interrogate cardiac hemodynamics. μECG signals allowed for identification of PW Doppler signals for passive (early [E]-wave velocity) and active ventricular filling (atrial [A]-wave velocity) during diastole. The A wave (9.0±1.2 cm·s(-1)) is greater than the E wave (1.1±0.4 cm·s(-1)), resulting in an E/A ratio <1 (0.12±0.05, n=6). In response to cryocauterization to the ventricular epicardium, the E-wave velocity increased, accompanied by a rise in the E/A ratio at 3 days postcryocauterization (dpc) (0.55±0.13, n=6, p<0.001 vs. sham). The E waves normalize toward the baseline, along with a reduction in the E/A ratio at 35 dpc (0.36±0.06, n=6, p<0.001 vs. sham) and 65 dpc (0.2±0.16, n=6, p<0.001 vs. sham). In zebrafish, E/A<1 at baseline is observed, suggesting the distinct two-chamber system in which the pressure gradient across the atrioventricular valve is higher compared with the ventriculobulbar valve. The initial rise and subsequent normalization of E/A ratios support recovery in the ventricular diastolic function.

Combined chirp coded tissue harmonic and fundamental ultrasound imaging for intravascular ultrasound: 20–60 MHz phantom and ex vivo results

The application of chirp coded excitation to pulse inversion tissue harmonic imaging can increase signal to noise ratio. On the other hand, the elevation of range side lobe level, caused by leakages of the fundamental signal, has been problematic in mechanical scanners which are still the most prevalent in high frequency intravascular ultrasound imaging. Fundamental chirp coded excitation imaging can achieve range side lobe levels lower than -60dB with Hanning window, but it yields higher side lobes level than pulse inversion chirp coded tissue harmonic imaging (PI-CTHI). Therefore, in this paper a combined pulse inversion chirp coded tissue harmonic and fundamental imaging mode (CPI-CTHI) is proposed to retain the advantages of both chirp coded harmonic and fundamental imaging modes by demonstrating 20-60MHz phantom and ex vivo results. A simulation study shows that the range side lobe level of CPI-CTHI is 16dB lower than PI-CTHI, assuming that the transducer translates incident positions by 50μm when two beamlines of pulse inversion pair are acquired. CPI-CTHI is implemented for a proto-typed intravascular ultrasound scanner capable of combined data acquisition in real-time. A wire phantom study shows that CPI-CTHI has a 12dB lower range side lobe level and a 7dB higher echo signal to noise ratio than PI-CTHI, while the lateral resolution and side lobe level are 50μm finer and -3dB less than fundamental chirp coded excitation imaging respectively. Ex vivo scanning of a rabbit trachea demonstrates that CPI-CTHI is capable of visualizing blood vessels as small as 200μm in diameter with 6dB better tissue contrast than either PI-CTHI or fundamental chirp coded excitation imaging. These results clearly indicate that CPI-CTHI may enhance tissue contrast with less range side lobe level than PI-CTHI.

High frequency photoacoustic imaging for in vivo visualizing blood flow of zebrafish heart.

A technique on high frame rate(28fps), high frequency co-registered ultrasound and photoacoustic imaging for visualizing zebrafish heart blood flow was demonstrated. This approach was achieved with a 40MHz light weight(0.38g) ring-type transducer, serving as the ultrasound transmitter and receiver, to allow an optic fiber, coupled with a 532nm laser, to be inserted into the hole. From the wire target study, axial resolutions of 38µm and 42µm were obtained for ultrasound and photoacoustic imaging, respectively. Carbon nanotubes were utilized as contrast agents to increase the flow signal level by 20dB in phantom studies, and zebrafish heart blood flow was successfully observed.

Non-contact acoustic radiation force impulse microscopy via photoacoustic detection for probing breast cancer cell mechanics

We demonstrate a novel non-contact method: acoustic radiation force impulse microscopy via photoacoustic detection (PA-ARFI), capable of probing cell mechanics. A 30 MHz lithium niobate ultrasound transducer is utilized for both detection of phatoacoustic signals and generation of acoustic radiation force. To track cell membrane displacements by acoustic radiation force, functionalized single-walled carbon nanotubes are attached to cell membrane. Using the developed microscopy evaluated with agar phantoms, the mechanics of highly- and weakly-metastatic breast cancer cells are quantified. These results clearly show that the PA-ARFI microscopy may serve as a novel tool to probe mechanics of single breast cancer cells.

High-frequency dual mode pulsed wave Doppler imaging for monitoring the functional regeneration of adult zebrafish hearts

Adult zebrafish is a well-known small animal model for studying heart regeneration. Although the regeneration of scars made by resecting the ventricular apex has been visualized with histological methods, there is no adequate imaging tool for tracking the functional recovery of the damaged heart. For this reason, high-frequency Doppler echocardiography using dual mode pulsed wave Doppler, which provides both tissue Doppler (TD) and Doppler flow in a same cardiac cycle, is developed with a 30 MHz high-frequency array ultrasound imaging system. Phantom studies show that the Doppler flow mode of the dual mode is capable of measuring the flow velocity from 0.1 to 15 cm s−1 with high accuracy (p-value = 0.974 > 0.05). In the in vivo study of zebrafish, both TD and Doppler flow signals were simultaneously obtained from the zebrafish heart for the first time, and the synchronized valve motions with the blood flow signals were identified. In the longitudinal study on the zebrafish heart regeneration, the parameters for diagnosing the diastolic dysfunction, for example, E/Em < 10, E/A < 0.14 for wild-type zebrafish, were measured, and the type of diastolic dysfunction caused by the amputation was found to be similar to the restrictive filling. The diastolic function was fully recovered within four weeks post-amputation.

Linear power amplifier for high frequency ultrasound coded excitation imaging

Commercial function generators and power amplifiers are generally used in high frequency coded excitation imaging systems. In order to minimize size, reduce external noise and deliver transmit energy to transducer efficiently in a system with the coded excitation, digital waveform generator and power amplifier should be placed on a single board. The pulser with capability of doing arbitrary waveform amplification was custom-designed and developed for a high frequency coded excitation imaging system. The matching of input stage was achieved by transmission line transformer (TLT) with ferrite core. Using TLT, matching circuit became simple otherwise it would require matching circuit using lumped components. Transformers were used between stages. The 6dB bandwidth of developed amplifier was from 10MHz to 110MHz. In the pass band, the gain of amplifier fluctuated within 4dB. Wire target measurements and ex-vivo measurements on tissues clearly demonstrated its superior performance.

High frequency, high frame rate pulse inversion chirp coded tissue harmonic imaging

The pulse inversion method, which alternatively transmits a burst and a subsequent one with its amplitude inversed, has been utilized to cancel out fundamental signal by summing up the pulse pair for second harmonic imaging. The approach was implemented with chirp coded excitation for a high frame rate mechanical sector scanner or an ultrasound biomicroscope (UBM), to suppress range side lobe level (RSLL) and to enhance spatial resolution and echo signal to noise ratio (eSNR). For this purpose, beam sequences and transmission timing strategy were optimized to reduce moving artifacts caused by the motor movement. The system was evaluated by scanning wire targets at a scanning speed of 68 Hz employing a 40 MHz lithium niobate (LiNbO3) single element light weight transducer. The wire phantom results showed that pulse inversion chirp coded tissue harmonic imaging (PI-CTHI) achieved a 4 dB higher eSNR than conventional pulse inversion tissue harmonic imaging (PI-THI), while maintaining the spatial resolution. RSLL of PI-CTHI was improved by 4 dB compared to band-pass filtered (F-CTHI).

Stand-alone front-end system for high- frequency, high-frame-rate coded excitation ultrasonic imaging

A stand-alone front-end system for high-frequency coded excitation imaging was implemented to achieve a wider dynamic range. The system included an arbitrary waveform amplifier, an arbitrary waveform generator, an analog receiver, a motor position interpreter, a motor controller and power supplies. The digitized arbitrary waveforms at a sampling rate of 150 MHz could be programmed and converted to an analog signal. The pulse was subsequently amplified to excite an ultrasound transducer, and the maximum output voltage level achieved was 120 Vpp. The bandwidth of the arbitrary waveform amplifier was from 1 to 70 MHz. The noise figure of the preamplifier was less than 7.7 dB and the bandwidth was 95 MHz. Phantoms and biological tissues were imaged at a frame rate as high as 68 frames per second (fps) to evaluate the performance of the system. During the measurement, 40-MHz lithium niobate (LiNbO3) single-element lightweight (<;0.28 g) transducers were utilized. The wire target measure- ment showed that the -6-dB axial resolution of a chirp-coded excitation was 50 μm and lateral resolution was 120 μm. The echo signal-to-noise ratios were found to be 54 and 65 dB for the short burst and coded excitation, respectively. The contrast resolution in a sphere phantom study was estimated to be 24 dB for the chirp-coded excitation and 15 dB for the short burst modes. In an in vivo study, zebrafish and mouse hearts were imaged. Boundaries of the zebrafish heart in the image could be differentiated because of the low-noise operation of the implemented system. In mouse heart images, valves and chambers could be readily visualized with the coded excitation.