Ultrasound has received less attention than other imaging modalities for molecular imaging, but has a number of potential advantages. It is cheap, widely available and portable. Using Doppler methods, flow information can be obtained easily and non-invasively. It is arguably the most physiological modality, able to image structure and function with less sedation than other modalities. This means that function is minimally disturbed, and multiple repeat studies or the effect of interventions can easily be assessed. High frame rates of over 200 frames a second are achievable on current commercial systems, allowing for convenient cardiac studies in small animals. It can be used to guide interventional or invasive studies, such as needle placement. Ultrasound is also unique in being both an imaging and therapeutic tool and its value in gene therapy has received much recent interest. Ultrasound biomicroscopy has been used for in utero imaging and can guide injection of virus and cells. Ultrahigh frequency ultrasound can be used to determine cell mechanical properties. The development of microbubble contrast agents has opened many new opportunities, including new functional imaging methods, the ability to image capillary flow and the possibility of molecular targeting using labelled microbubbles. Ultrasound is the most widely used cross-sectional imaging modality worldwide (1). Important recent devel- opments include microbubble contrast agents, quantitative approaches and new signal processing and display methods. In this review, we describe some recent devel- opment of the technology of ultrasound relevant to molecular imaging. These include a number of promising ways of imaging the microcirculation using ultrasound, advances in high and ultrahigh frequency ultrasound, and an overview of the potential of microbubble contrast agents and ultrasound-mediated gene transfection. This review article will first describe some basic principles of ultrasound and then illustrate some applications relevant to molecular imaging. Key areas to be discussed will be developments in high frequency ultrasound, angiogenesis imaging, targeted microbubbles and microbubble ultra- sound mediated gene and drug delivery. Ultrasound refers to the sound at a frequency higher than 20 kHz, and may be broken down by the frequencies used. Clinical diagnostic ultrasound scanners use frequen- cies in the range of 1 MHz to 20 MHz. Ultrasound biomicroscopy (UBM) typically operates at frequencies of 40 MHz to 200 MHz, while scanning acoustic microscopy (SAM) uses frequencies higher than 200 MHz. Currently almost all commercial systems operate in the clinical diagnostic range, although commercial biomicroscopy is now a reality (http://www.visualsonics.com). It is crucial to understand that resolution improves with frequency, while penetration dramatically decreases with frequency. Therefore, the choice of ultrasound frequency is a balance between resolution and penetration depth, with higher frequencies used for more superficial structures. Pulses of ultrasound are sent out in trains from arrays of transducers (probes) and the returning echoes analysed to build up a two-dimensional image of the plane being scanned and displayed using a grey-scale display to code the intensity of the returning echoes. This produces the

The role of ultrasound in molecular imaging

Objective. To investigate the potential efficacy of real-time contrast-enhanced power Doppler sonography in the differentiation of benign and malignant adnexal masses in a pilot study. Methods. Before surgical treatment, adnexal masses were prospectively evaluated with power Doppler sonography before and after injection of a contrast agent. Real-time postinjection sequences were computerized with time-intensity analysis software to determine an enhancement curve and contrast parameters. The intraobserver and interobserver reproducibilities of these criteria were assessed on a subsample. These contrast parameters were compared between benign and malignant tumors using logistic regression. Sensitivity and specificity were used to compare contrast parameters with sonographic and Doppler variables. Results. Ninety-nine women were included, for a total of 101 adnexal masses. There were 23 cases of ovarian malignancies and 78 benign adnexal lesions. Our procedure had excellent intraobserver and interobserver reproducibility, with an average intraclass correlation coefficient of 0.92. The time before enhancement and intensity ratio did not reliably differentiate between the benign and malignant masses. Washout times and areas under the curves were significantly greater in ovarian malignancies than in other benign tumors (P <.001), leading to sensitivity estimates between 96% and 100% and specificity estimates between 83 and 98%. Contrast parameters had slightly higher sensitivity and slightly lower specificity when compared with transvaginal sonographic variables of the resistive index and serum cancer antigen 125 levels. Conclusions. Contrast-enhanced power Doppler imaging may easily and precisely discriminate benign from malignant adnexal lesions. Larger studies are needed to determine the appropriate use and benefits of this new procedure.

Contrast-enhanced sonography helps in discrimination of benign from malignant adnexal masses

PURPOSE: To evaluate the effects of a microbubble contrast agent on the power Doppler ultrasonographic (US) examination of adnexal tumors, with a special focus on the timing of the transit of the microbubble bolus. MATERIALS AND METHODS: Seventy patients who were suspected of having ovarian tumors were examined preoperatively with contrast material–enhanced US. Images obtained during a 5-minute examination were stored digitally, and the behavior of the contrast agent was evaluated objectively with measurement of the time-dependent image intensity at the region of interest with a computer program. A time-intensity curve in each case was derived and analyzed. The Mann-Whitney U test was used to compare intensity changes and tumor parameters in benign and malignant adnexal tumors. RESULTS: Both the baseline and maximum power Doppler intensities, as well as the absolute and relative (percent) rise in intensity, were significantly higher (P < .001) in malignant as compared with benign tumors. The arrival time ...

Kinetics of a US contrast agent in benign and malignant adnexal tumors.

Background—No noninvasive technique is currently capable of “real-time” assessment and monitoring of cerebral blood flow (CBF). We hypothesized that cerebral perfusion could be accurately measured and monitored in “real time” with contrast-enhanced ultrasound (CEU). Methods and Results—Cerebral perfusion was assessed in 9 dogs through a craniotomy with CEU at baseline and during hypercapnia and hypocapnia while normoxia was maintained. Cerebral microvascular blood volume (A), microbubble velocity (), and blood flow (A) were calculated from time-versus–acoustic intensity relations. Compared with baseline, hypercapnia and hypocapnia significantly increased and decreased CBF, respectively, as measured by CEU. These changes in blood flow were mediated by changes in both A and. A good correlation was found between A derived from CEU and CBF measured by radiolabeled microspheres (y0.67x0.04, r0.91, P0.001). Conclusions—Changes in both cerebral microvascular blood volume and red blood cell velocity can be accurately assessed with CEU. Thus, CEU has the potential for bedside measurement and monitoring of cerebral perfusion in real time in patients with craniotomies or burr holes. (Circulation. 2001;104:2582-2587.) n current clinical practice, noninvasive assessment of cerebral blood flow (CBF) can be achieved by use of various techniques, such as CT, MRI, and single photon emission CT (SPECT). These techniques, however, are ill suited for measurement of CBF at the bedside or in the operating room. Furthermore, they cannot be used repeatedly, in rapid succession, to measure changes in CBF that may occur in the natural course of events or in response to treatment. In addition, these accurate, albeit expensive, modalities cannot be used for monitoring regional CBF in patients with acute ischemic stroke or intracranial hemorrhage, or for the serial assessment of global CBF in patients with increased intracranial pressure. We have previously shown that blood flow to various tissues, including the myocardium,1 skeletal muscle,2 and kidney,3 can be measured by contrast-enhanced ultrasound (CEU). This technique, which uses small gas-filled microbubbles, can provide an assessment of microvascular blood volume (MBV) as well as red blood cell velocity (RBCv). These parameters can be obtained during a continuous intravenous infusion of microbubbles. After steady state is achieved, the microbubbles within tissue are destroyed with high-energy ultrasound, and the rate at which they replenish the ultrasound beam is measured, which reflects RBCv. The concentration of microbubbles when the beam is fully replenished is proportional to MBV. We therefore hypothesized that CEU can be used to assess CBF noninvasively. We performed experiments in dogs in which CBF was measured with both CEU and radiolabeled microspheres during rest, hypercapnia, and hypocapnia. The aim was to demonstrate proof of principle so that future technical developments could assist in transferring this method to the clinical setting. Methods

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Quantification of Cerebral Perfusion With “Real-Time” Contrast-Enhanced Ultrasound

To improve the quantification of tissue perfusion using intermittent sonography, a new model describing replenishment kinetics of microbubbles is proposed. The new approach takes into account the variability of blood flow velocities found in vivo , especially in tumors, and consistently describes the refilling process of microbubbles. Based upon this model, blood volume, blood velocity, blood flow and perfusion in 17 experimental tumors were calculated, and compared with the results obtained with the established, phenomenologically derived exponential kinetic model. In contrast to the existing model, our approach describes tissue vascularization more physiologically and allows deduction of a consistent new hyperbolic model for quantification of intermittent sonography. Blood volume and mean blood velocity did significantly correlate between both the new and the established model (k = 0.99; k = 0.94, both p p in vivo can be estimated with the new model. Furthermore, it uses simpler mathematical fitting routines and allows easier data acquisition, which may allow a more practicable clinical application of intermittent sonography. In conclusion, a more valid, detailed and accurate calculation of perfusion parameters, especially of tumors, can be derived in vivo with the new multivessel model of intermittent sonography. (E-mail: m.krix@dkfz.de)

A multivessel model describing replenishment kinetics of ultrasound contrast agent for quantification of tissue perfusion.

Background and Purpose—Through harmonic gray-scale imaging, it is possible to analyze brain tissue perfusion with different ultrasound methods. Methods—In 12 healthy volunteers, 2 doses (0.5 and 1.5 mL) of Optison, a perfluoropropane-containing contrast agent, were injected intravenously and produced a strong increase of brightness in the brain parenchyma. We used harmonic imaging for quantification of ultrasound intensity in the thalamus, ipsilateral temporoparietal white matter (TPWM), and ipsilateral lateral fissure at both sides. Time-intensity curves were calculated, and peak increase (PI) of intensity and the area under the time-intensity curve (AUC) from baseline were compared. Results—We found a significant dose dependence of the AUC in all regions at both sides. PI only showed a significant dose dependence in the TPWM but not in the ipsilateral thalamus and lateral fissure. No side differences for AUC and PI were detected in all regions and doses used. We found a significantly higher value of the...

Harmonic Imaging of the Human Brain Visualization of Brain Perfusion With Ultrasound

By means of harmonic imaging, it is possible to display brain perfusion qualitatively using ultrasound (US) contrast agent (UCA) bolus injection. With UCA continuous infusion reaching a steady state, mean microbubble velocity can be measured, analyzing the reappearance rate after microbubble destruction by US (refill kinetics). We performed an animal pilot study to investigate this new method for the assessment of brain perfusion. Using harmonic grey-scale imaging, five sedated male beagle dogs were investigated through the intact skull with increasing pulsing intervals (250 to 8000 ms) and three UCA infusion rates (0.5, 1.0 and 1.5 mL/min of Optison™). Cerebral blood flow was increased by acetazolamide (30 mg/kg BW). Intensity vs. pulsing interval curves were analyzed using an exponential curve fit [ I(t) = A (1- e −βt ) ] and parameters of the curve were compared. We found that increasing the pulsing interval above 4000 ms led to no further increase of echo enhancement for infusion rates. Mean β values were not influenced by infusion rate ( p = 0.25 and p = 0.55). Mean F values increased nonsignificantly with rising infusion rate ( p = 0.25 and p = 0.86). Acetazolamide led to an increase of mean β and F values ( p = 0.18 and p = 0.025, respectively). It is possible to evaluate changes in brain perfusion through the intact skull by analyzing the UCA refill kinetics after US-induced microbubble destruction. (E-mail:seidel_g@neuro.mu-luebeck.de)

Evaluation of blood flow in the cerebral microcirculation: analysis of the refill kinetics during ultrasound contrast agent infusion

The influence of Doppler system settings on the clearance kinetics of different ultrasound contrast agents

Ultrasound contrast agents improve the signal-to-noise ratio of reflected ultrasound, enhancing the diagnostic value of transcranial Doppler (TCD). In dog studies, we investigated the time course of TCD signal amplitude after application of a phospholipid-containing ultrasound contrast agent (BY963) filled with different gases. The median time of Doppler amplitude enhancement exceeding 5 dB was determined using isoflurane-, isopentane-, trichlortrifluoroethane-, air-, argon-, and perfluoropentane-filled BY963 (69, 72, 75, 78, 88, and 245 seconds respectively). The decrease of time-intensity curve and the duration of signal enhancement showed significant differences comparing the different gases (p = 0.04 and 0.03, respectively). The time course of in vitro stability of BY963 agitated with the different gases measured by absorbance of light (500 nm) showed a retarded decay for perfluoropentane, a rapid decrease for air, isopentane, trichlortrifluoroethane, and argon, and a very rapid decrease using isoflurane. The time course of the different gases depended on the physiochemical properties (lipophilicity and the solubility in water) of the gas encoated in the phospholipid shell. Perfluoropentane-filled BY963 showed the highest in vitro stability and the longest duration of TCD enhancement compared with the other gases used.

The influence of different gases on acoustic properties of a spherosome-based ultrasound contrast agent (BY963). A transcranial Dopplersonography study.

Marion Vidal-Langwasser

Papers

Harmonic Imaging of the Human Brain Visualization of Brain Perfusion With Ultrasound

Evaluation of blood flow in the cerebral microcirculation: analysis of the refill kinetics during ultrasound contrast agent infusion

The influence of Doppler system settings on the clearance kinetics of different ultrasound contrast agents

The influence of different gases on acoustic properties of a spherosome-based ultrasound contrast agent (BY963). A transcranial Dopplersonography study.