Comparative study of temperature measurements in ex vivo swine muscle and a tissue-mimicking material during high intensity focused ultrasound exposures.

Tissue-mimicking materials (TMMs) can provide a convenient, stable, and reproducible means for testing high intensity focused ultrasound (HIFU) devices. When TMMs containing thermal sensors are used to measure ultrasound-induced temperature rise, it is important that measurement results reasonably represent those that occur in biological tissue. Therefore the aim of this paper is to compare the thermal behavior of the TMM under HIFU exposure to that of ex vivo tissue. This was accomplished using both a previously developed TMM and fresh ex vivo swine muscle that were instrumented with bare 50 µm thin wire thermocouples. HIFU at 825 kHz was focused at the thermocouple junction. 30 s exposures of increasing peak negative pressure (1 to 5 MPa) were applied and the temperature profile during and after sonication was recorded. B-mode imaging was used to monitor bubble activity during sonication. If bubble formation was noted during the sonication, the sonication was repeated at the same pressure levels two more times at 20 min intervals. Temperature traces obtained at various pressure levels demonstrated similar types of heating profiles in both the tissue and TMM, the exact nature of which depended on whether bubbles formed during the HIFU exposure. The onset of bubble activity occurred at lower ultrasonic pressures in the TMM, but the basic temperature rise features due to HIFU exposure were essentially the same for both materials.

Spatially resolved fluorescence lifetime mapping of enzyme kinetics in living cells.

Traditional cuvette-based enzyme studies lack spatial information and do not allow real-time monitoring of the effects of modulating enzyme functions in vivo. In order to probe the realistic timescales of steric modifications in enzyme-substrate complexes and functional binding-unbinding kinetics in living cells without losing spatial information, it is imperative to develop sensitive imaging strategies that can report enzyme kinetics in real time over a wide dynamic range of timescales. Here we present a multi-photon excitation-based, ultra-fast photon detection using a streak camera and Laguerre expansion-based fast deconvolution approach for achieving high spatio-temporal resolution in monitoring real-time enzyme kinetics in single cells. In particular, we report spatially resolved, nanosecond-scale fluorescence dynamics associated with binding-unbinding kinetics of endogenous metabolic co-factor nicotinamide adenine dinucleotide with enzymes in intact living cells. By monitoring real-time kinetics of NAD(P)H-enzyme kinetics in primary hepatocytes isolated from young and aged mouse models, we observed that the mechanism of inhibition of mitochondrial respiration at complex I site is mediated by redistribution of free and protein-bound nicotinamide adenine dinucleotide pools and that this equilibrium redistribution is affected by age-related modifications in mitochondrial function. We describe unique advantages of Laguerre deconvolution algorithm in comparison with conventional lifetime analysis approaches. Non-invasive monitoring of metabolic dysfunctions in intact animal models is an attractive strategy for gaining insight into the dynamics of tissue metabolism in health and in various metabolic syndromes such as cancer, diabetes and aging-induced metabolic dysfunctions. Besides the example demonstrated above, we envisage that the proposed method can find applications in a variety of other situations where intensity-based approaches fall short owing to spectroscopic artefacts.

Theoretical study of steady-state temperature rise within the eye due to ultrasound insonation.

The soft tissue thermal index (TIS), as defined in the AIUM/NEMA Output Display Standard, may not be relevant with respect to eye exposure, primarily because of differences in actual vs. assumed acoustic and thermal properties. Therefore, a theoretical study of temperature rise within the eye due to ultrasound insonation was undertaken to compare the TIS with more exact calculations. At each plane in the direction of propagation, the focused ultrasound beam was modeled as a disc of uniform intensity. Each disc becomes a heat source, and integration over all discs provides the total temperature rise at any axial position. Calculations were done assuming the ultrasound beam intersects the lens of the eye as well as for the case in which the beam does not intersect the lens. Results were found for frequencies of 7.0 MHZ to 40 MHZ, transducer diameters of 0.2 cm to 1.0 cm, and focal lengths ranging from 0.2 cm to 3.0 cm. Perfusion was assumed negligible and thermal and acoustic parameters were taken from reported studies. For every case, the ratio of maximum temperature rise to the TIS (assuming constant output power) was calculated. For the lens case, the ratio varied from 7.35 to 0.8. For the no-lens case, the ratio varied from 4.1 to 0.4. These results indicate that the TIS is not adequate to represent the temperature rise occurring within the eye upon insonation.

Maximum likelihood estimation of receiver operating characteristic (ROC) curves from continuously-distributed data.

We show that truth-state runs in rank-ordered data constitute a natural categorization of continuously-distributed test results for maximum likelihood (ML) estimation of ROC curves. On this basis, we develop two new algorithms for fitting binormal ROC curves to continuously-distributed data: a true ML algorithm (LABROC4) and a quasi-ML algorithm (LABROC5) that requires substantially less computation with large data sets. Simulation studies indicate that both algorithms produce reliable estimates of the binormal ROC curve parameters a and b, the ROC-area index Az, and the standard errors of those estimates.

Statistical comparison of two ROC-curve estimates obtained from partially-paired datasets.

The authors propose a new generalized method for ROC-curve fitting and statistical testing that allows researchers to utilize all of the data collected in an experimental comparison of two diagnostic modalities, even if some patients have not been studied with both modalities. Their new algorithm, ROCKIT, subsumes previous algorithms as special cases. It conducts all analyses available from previous ROC software and provides 95% confidence intervals for all estimates. ROCKIT was tested on more than half a million computer-simulated datasets of various sizes and configurations representing a range of population ROC curves. The algorithm successfully converged for more than 99.8% of all datasets studied. The type I error rates of the new algorithm's statistical test for differences in Az estimates were excellent for datasets typically encountered in practice, but diverged from alpha for datasets arising from some extreme situations.

Study of an acoustic technique to detect cavitation produced by a tilting disc valve.

BACKGROUND AND AIM OF THE STUDY: Transient cavitation has been directly observed near operating mechanical heart valves in vitro and inferred in vivo via the observation of pitting on explanted clinically used valves. Visual detection of cavitation bubbles, however, cannot be accomplished in vivo or when any opaque fluid, e.g. blood, is used. METHODS: This study examines a passive acoustic technique for detecting cavitation caused by a 27 mm tilting disc valve. We captured, valve closing sounds in vitro and attempted to detect a shift of energy into higher frequencies due to emission of broad-band noise caused by collapsing bubbles. The valve tester consists of a piston pump which directly drives the valve, a 16 cm diameter cylindrical lucite atrium and an air chamber in parallel to provide compliance. Water was used as a blood analog fluid. The cycle rate was altered to vary valve loading and produce cavitation. Acoustic signals were detected by a miniature hydrophone and a large area transducer and the waveforms were spectrum analyzed to 200 kHz and 500 kHz respectively. Cavitation onset was determined rising a high speed video camera. RESULTS: It was found that even under non-cavitating conditions, significant energy was produced at frequencies greater than 100 kHz, and this energy increased with increased load. The proportion of energy in high frequency bands, however, remained fairly constant when cavitation was not present and began to rise only after the cavitation threshold was reached. To isolate cavitation as an independent variable, data were taken with all system parameters constant, but using water under two different conditions. Degassed 17 degrees C water produced no visualizable bubbles, while aerated tap water at 43 degrees C showed a high degree of cavitation. CONCLUSIONS: The results indicate that cavitation, while causing a shift of energy to higher frequencies, is not the only mechanism responsible for the shift of energy into higher frequencies.

An interlaboratory comparison of the FDA protocol for the evaluation of cavitation potential of mechanical heart valves.

Five laboratories carried out measurements of cavitation threshold for a common set of six mechanical prosthetic heart valves, two each from three different manufacturers. This study was intended to evaluate to what extent FDA's current guidance for cavitation testing would lead to consistent results in a variety of laboratory settings and to seek areas for improvement in the recommended test protocol. The inter-laboratory study protocol specified: (1) characterization of the test fluid by oxygen content and electrical conductivity, (2) location and frequency response of pressure sensors, (3) determination of ventricular and atrial pressures (P) and loading rates (dP/dt) averaged over the time period of valve closure and over the time periods of 1 ms, 5 ms, and 20 ms prior to video visualization. The protocol did not specify: (1) the fluid pumping equipment to be used to generate cavitation, (2) the pump or fluid parameters adjusted to raise or lower the loading rate, (3) the equipment, technique, or sensitivity used to visualize cavitation, and (4) a specific definition of the threshold for cavitation. Results from the five laboratories are reported. Significant differences in results were observed in dP/dt and in the pressure difference across the valves during closure at cavitation threshold. Specific differences in test systems included a wide range of ventricular compliance and single valved versus double valved test systems. Three single valve systems with compliant ventricles produced results in reasonable agreement with one another. Further similarity in test equipment should be specified to assure adequate interlaboratory reliability for cavitation testing. Areas needing better specification include the design of the valve mount, the design of the cavitation generators, and qualitative criteria for detection of threshold cavitation.

A protocol for the evaluation of the cavitation potential of mechanical heart valves.

To assure the safety and effectiveness of prosthetic heart valves, the Food and Drug Administration requires performance testing prior to human use. Guidance for such testing is currently undergoing revision to include in vitro tests for cavitation potential. This work describes a test procedure to determine the minimum ventricular dp/dt associated with bubble formation. Cavitation bubbles as small as 0.5 mm diameter existing for 0.02 msec should be detectable by video imaging techniques.

Effect on verapamil on ventricular function: studies in denervated human heart.

Verapamil has complex influences on ventricular function owing to its direct myocardial effects, vasodilation, and reflex activation of the sympathetic nervous system. To investigate the direct myocardial effects of verapamil in humans independent of reflex sympathetic stimulation, we administered the drug to 13 recent heart transplant recipients with denervated ventricles. Hemodynamics and radionuclide angiograms were recorded at baseline, with altered loading conditions, and after intravenous (i.v.) verapamil (median dose 4 mg). Left ventricular (LV) systolic and diastolic function was analyzed by systolic pressure-volume relations (SPVR) and peak filling rate (PFR), respectively. Verapamil caused a decrease in blood pressure (BP) and heart rate (HR) with increases in right atrial pressure (RAP 6 +/- 3-8 +/- 3, p < 0.01) and pulmonary artery wedge pressure (PAWP, 9 +/- 3-11 +/- 3 mm Hg, p < 0.01) pressures. LV ejection fraction (EF) decreased (69 +/- 7-66 +/- 8%, p < 0.02) in association with an increase in LV end-systolic counts (3.45 +/- 1.27 to 4.72 +/- 1.78 kcts, p < 0.001). In 11 of 13 patients, the SPV point after verapamil administration was decreased from the line established during altered loading conditions. PFR (4.05 +/- 0.81 to 4.11 +/- 0.76 EDV/s) was unchanged. In the denervated ventricle, verapamil has negative chronotropic and inotropic effects with minimal effects on PFR.

Diastolic dysfunction in cardiac transplant recipients: an important role in the response to increased afterload.

We evaluated the hemodynamic and functional response to acute elevations in left ventricular (LV) afterload in 22 recent recipients of cardiac transplants to determine whether abnormalities in LV diastolic function influence the response to this intervention. In seven patients (group 1) LV ejection fraction decreased significantly from baseline values (> or = 5%) during methoxamine infusion, whereas in 15 patients (group 2) LV ejection fraction was maintained. Peak filling rate was lower in group 1 versus group 2 (3.36 +/- 0.46 vs 4.23 +/- 0.68 end-diastolic volumes/sec, p < 0.01). In addition, patients in group 1 did not have LV dilatation during methoxamine (percentage change in end-diastolic counts, -3.4 +/- 6.9%) and had a large increase in pulmonary artery wedge pressure. In contrast, patients in group 2 had LV dilatation (percentage change in end-diastolic counts, +10.7 +/- 14.7%) and a smaller increase in pulmonary artery wedge pressure. There was a relationship between the baseline peak filling rate and the change in LV ejection fraction during methoxamine (r = 0.65, p = 0.001). Therefore in a subset of cardiac transplant patients, abnormalities in LV filling can have an impact on the response to increased afterload.

Effectiveness of latex condoms as a barrier to human immunodeficiency virus-sized particles under conditions of simulated use.

Condoms were tested in an in vitro system simulating key physical conditions that can influence viral particle leakage through condoms during actual coitus. The system quantitatively addresses pressure, pH, temperature, surfactant properties, and anatomical geometry. A suspension of fluorescence-labeled, 110-nm polystyrene microspheres models free human immunodeficiency virus (HIV) in semen, and condom leakage is detected spectrofluorometrically. Leakage of HIV-sized particles through latex condoms was detectable (P less than 0.03) for as many as 29 of the 89 condoms tested. Worst-case condom barrier effectiveness (fluid transfer prevention), however, is shown to be at least 10(4) times better than not using a condom at all, suggesting that condom use substantially reduces but does not eliminate the risk of HIV transmission.

PIP: Physical science researchers tested the ability of 89 undamaged latex condoms manufactured in the US to prevent passage of HIV=size particles under simulated physiologic conditions at their Food and Drug Administration laboratory in Rockville, Maryland. The design of the test system considered particle size, pH, surface tension, and time. A suspension of polystyrene 110 nm microspheres labeled with fluorescent dye served as the HIV-sized particle model in semen. They challenged each condom with this suspension for 30 minutes. The test did not include motion since stretching over the penis accounts for most pore stretching. Leakage of fluorescent dye occurred in 29 condoms (p .03). 21 condoms leaked at minimum leak rates 1 nl/s, 7 at 1-6 nl/s, and 1 at around 10 nl/s. Assuming the leakage occurred through the only pore in each condom, the pore diameters ranged from 2 to 7 mcm. Also assuming an even more conservative criterion, the qualitative results were the same: 11 condoms with leak rates were nl/s vs. 6 condoms with leak rates 1-9 nl/s (p .002). The widely used 300 ml water test did not indicate any pores in any of the condoms. In the extreme and highly unlikely scenario of all the fluid being pumped out of the condom, the transfer rate would be about 0.1 mcl after 10 minutes of thrusting after ejaculation filled the condom with semen (i.e., 0.01% of a typical 3 ml ejaculate). Thus proper use of latex condoms would result in exposure reduction from HIV of at least 4 orders of magnitude. These findings demonstrated that use of latex condoms can significantly reduce the risk of HIV transmission, but it does not eliminate that risk.

Effects of acute alterations in left ventricular loading conditions on peak filling rate in the denervated (transplanted) ventricle.

Peak filling rate is an indicator of left ventricular (LV) diastolic function. It is influenced by heart rate, loading conditions, sympathetic nervous system activity, ejection fraction and other factors. To determine the effect of altered loading conditions on peak filling rate, independent of heart rate and sympathetic nervous system activity, 12 patients were studied 3 weeks after orthotopic heart transplantation. Plasma catecholamine level, heart rate and ejection fraction were not changed by any maneuver. Nitroglycerin caused a decrease in pulmonary artery wedge pressure (9 +/- 2 to 6 +/- 1 mm Hg, p less than 0.001) and in absolute peak filling rate (46.0 +/- 3.0 to 42.8 +/- 2.5 kcts/s, p less than 0.01), but no change in normalized peak filling rate. Volume infusion increased pulmonary artery wedge pressure (9 +/- 2 to 12 +/- 2 mm Hg, p less than 0.001) and absolute peak filling rate (46.0 +/- 3.0 to 51.5 +/- 5.3 kcts/s, p less than 0.01), but peak filling rate normalized to stroke volume was unchanged. During nitroglycerin and volume infusions, there was a high correlation between changes in pulmonary artery wedge pressure and absolute peak filling rate (r = 0.82, p less than 0.001). With normalization of peak filling rate, these variables correlated less well. With methoxamine, 4 patients demonstrating systolic dysfunction had a decrease in absolute and normalized peak filling rate despite a large increase in pulmonary artery wedge pressure. The other 8 patients without systolic dysfunction had an increase in pulmonary artery wedge pressure with increased absolute but unchanged normalized peak filling rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Test method for evaluating the permeability of intact prophylactics to viral-size microspheres under simulated physiologic conditions.

The alarming number of AIDS cases has increased the attention given to barrier devices such as condoms. The authors describe a new test method that evaluates the permeability of the intact condom when subjected to simulated physiologic conditions. Fluorescent-labelled polystyrene microspheres (110 nm diameter) are used to model cell-free virus. Physical and chemical conditions that are present during coitus, such as pressure, pH, and temperature, are considered in the design of the method. The testing chamber is designed to be continuously monitored for changes in fluorescence due to leakage across the condom surface. The sensitivity of the system is 1 x 10(-5) of the original concentration of microsphere solution (3.4 x 10(11) particles/mL), which corresponds to leak rates as small as .001 microL/sec. The test provides an in vitro test of barrier material permeability relevant to actual use.

Digital acoustical analysis of normal and bimodal Björk-Shiley 60 degrees convexo-concave heart valves.

Fracture of the outlet strut of the Björk-Shiley 60 degrees convexo-concave (BS60CC) valve has been attributed to a bimodal closing pattern in certain valves in which the closing disk rotates about the inlet strut, causing upward displacement of the outlet strut and its eventual fracture. This article reports the in vivo studies of the normal BS60CC valve and the in vitro studies of the normal and bimodal BS60CC valves, using a digital acoustical signal processing technique, in which the individual collisions (impact history) of the occluder disk with the components of the valve body are revealed during each closing cycle. In vitro analysis of the closing acoustical signals of normal BS60CC valves showed impact history cluster width (IHCW) means of 2.07 +/- 0.85 ms (standard error), not significantly different from those of 1.86 +/- 0.58 ms (standard error) observed in 38 clinically normal patients with BS60CC valves (p greater than 0.1). The bimodal valves showed IHCW of 6.14 +/- 0.98 ms (standard error), in vitro, which was significantly greater than those observed in the normal in vitro valve group and in the normal patient population (p less than 0.0001).

The effects of a glycerin-based blood analog on the testing of bioprosthetic heart valves.

Detailed comparisons of aortic valvular flow using saline, with that using a glycerin-based blood analog in a pulse duplicator are reported. The experiments were carried out to determine whether exposure to glycerin caused stiffening of bioprosthetic valve leaflets. For two pericardial bioprostheses and for a mechanical valve we observed a fluid-dependent systolic volume flow, a fluid-dependent regurgitation volume, and fluid-dependent systolic pressure differences. Volume flow changes, both forward and reverse, are independent of valve type. The observed pressure differences, while proportional to fluid density for the mechanical valve, are fluid dependent in a more complicated way for the pericardial valves. However, no trend of changing valvular performance was observed over as much as 80 days of glycerin exposure, indicating that it is unlikely that the fluid-dependent performance was caused by glycerin absorption by the valve leaflets. We conclude that valid performance comparisons between mechanical and bioprosthetic valves may be made using a glycerin-based fluid. Furthermore, it appears that any detailed analysis of the physical mechanisms of valvular flow dissipation will require a properly matched blood analog.

Interlaboratory comparison of ultrasonic attenuation and speed measurements.

A set of test samples, all containing ultrasonically equivalent tissue-mimicking material, was produced and measurements of ultrasonic speed and ultrasonic attenuation coefficients were made at seven laboratories using various techniques. The ultrasonic speed values agree well with one another, having a spread of about 0.3 per cent; thus, speed values for tissue parenchyma appearing in the literature are likely to be accurate. Values of ultrasonic attenuation coefficients agree fairly well with one another, with differences between individual values and the group mean of generally less than 20 per cent of the group mean.

Dynamics and topographical distribution of surface glycoproteins during myoblast fusion: a resonance energy transfer study.

We have investigated changes in topography and lateral translational mobility of concanavalin A (Con A) receptors on the surface of cultured chick muscle cells during the period of myoblast fusion. A temporal correlation between these phenomena and the alteration in membrane fluidity known to occur during this time period is established. Receptor topography and mobility are studied by means of a resonance energy transfer technique employing pyrene- and FITC-Con A conjugates. All measurements are performed through a microscope on single cells. Our results reveal that during the period of myoblast fusion Con A receptors undergo a dramatic redistribution on the cell surface. Furthermore, our data suggest that the changes in membrane fluidity observed during muscle differentiation serve to modulate the lateral mobility of these receptors.