Eitan Kimmel

Intramembrane cavitation as a unifying mechanism for ultrasound-induced bioeffects

The purpose of this study was to develop a unified model capable of explaining the mechanisms of interaction of ultrasound and biological tissue at both the diagnostic nonthermal, noncavitational (<100 mW·cm−2) and therapeutic, potentially cavitational (>100 mW·cm−2) spatial peak temporal average intensity levels. The cellular-level model (termed “bilayer sonophore”) combines the physics of bubble dynamics with cell biomechanics to determine the dynamic behavior of the two lipid bilayer membrane leaflets. The existence of such a unified model could potentially pave the way to a number of controlled ultrasound-assisted applications, including CNS modulation and blood–brain barrier permeabilization. The model predicts that the cellular membrane is intrinsically capable of absorbing mechanical energy from the ultrasound field and transforming it into expansions and contractions of the intramembrane space. It further predicts that the maximum area strain is proportional to the acoustic pressure amplitude and inversely proportional to the square root of the frequency () and is intensified by proximity to free surfaces, the presence of nearby microbubbles in free medium, and the flexibility of the surrounding tissue. Model predictions were experimentally supported using transmission electron microscopy (TEM) of multilayered live-cell goldfish epidermis exposed in vivo to continuous wave (CW) ultrasound at cavitational (1 MHz) and noncavitational (3 MHz) conditions. Our results support the hypothesis that ultrasonically induced bilayer membrane motion, which does not require preexistence of air voids in the tissue, may account for a variety of bioeffects and could elucidate mechanisms of ultrasound interaction with biological tissue that are currently not fully understood.

Investigations into Pulsed High-Intensity Focused Ultrasound–Enhanced Delivery: Preliminary Evidence for a Novel Mechanism

Pulsed high-intensity focused ultrasound (HIFU) exposures without ultrasound contrast agents have been used for noninvasively enhancing the delivery of various agents to improve their therapeutic efficacy in a variety of tissue models in a nondestructive manner. Despite the versatility of these exposures, little is known about the mechanisms by which their effects are produced. In this study, pulsed-HIFU exposures were given in the calf muscle of mice, followed by the administration of a variety of fluorophores, both soluble and particulate, by local or systemic injection. In vivo imaging (whole animal and microscopic) was used to quantify observations of increased extravasation and interstitial transport of the fluorophores as a result of the exposures. Histological analysis indicated that the exposures caused some structural alterations such as enlarged gaps between muscle fiber bundles. These effects were consistent with increasing the permeability of the tissues; however, they were found to be transient and reversed themselves gradually within 72 h. Simulations of radiation force-induced displacements and the resulting local shear strain they produced were carried out to potentially explain the manner by which these effects occurred. A better understanding of the mechanisms involved with pulsed HIFU exposures for noninvasively enhancing delivery will facilitate the process for optimizing their use.

ULTRASOUND-INDUCED CAVITATION DAMAGE TO EXTERNAL EPITHELIA OF FISH SKIN

Transmission electron microscopy was used to show the effects of therapeutic ultrasound (< or = 1.0 W/cm2, 1 MHz) on the external epithelia of fish skin. Exposures of up to 90 s produced damage to 5 to 6 of the outermost layers. Negligible temperature elevations and lack of damage observed when using degassed water indicated that the effects were due to cavitation. The minimal intensity was determined for inducing cellular damage, where the extent and depth of damage to the tissues was correlated to the exposure duration. The results may be interpreted as a damage front, advancing slowly from the outer cells inward, presumably in association with the slow replacement of the perforated cell contents with the surrounding water. This study illustrates that a controlled level of microdamage may be induced to the outer layers of the tissues.

Preliminary investigations of ultrasound induced acoustic streaming using particle image velocimetry

Particle image velocimetry was used to investigate ultrasound-induced acoustic streaming in a system for the enhanced uptake of substances from the aquatic medium into fish. Four distinct regions of the induced streaming in the system were observed and measured. One of the regions was identified as an preferential site for substance uptake, where the highest velocities in proximity to the fish surface were measured. A positive linear relationship was found between the ultrasound intensity and the maximum streaming velocity, where a unitless geometric factor, specific to the system, was calculated for correcting the numerical relationship between the two parameters. The results are part of a comprehensive study aimed at improving mass transdermal administrations of substances (e.g. vaccines, hormones) into fish from the aquatic medium.

Ultrasound-induced intercellular space widening in fish epidermis

Transmission electron microscopy was employed to determine the effects of therapeutic ultrasound (US) (I(sata) < or =2.2 W cm(-2), 3 MHz), sonicated at different angles and durations, on the external epithelia of fish skin. Sonication at 1.7 W cm(-2) (90 s), where the ultrasonic beam was perpendicular to the skin surface, produced minor intercellular space widening (ICSW), as well as the disruption of desmosomes connecting between the cells. Increasing the intensity to 2.2 W cm(-2) increased ICSW, the extent of which was positively correlated to the duration of exposure (30 to 90 s). Perpendicular sonication produced ICSW, almost exclusively between cells of the two outermost cell layers, parallel to the skin surface. Sonicating at 45 degrees (2.2 W cm(-2), 90 s) produced ICSW in deeper cell layers in the tissues, in which the spaces were at seemingly random orientations. Mucous cells and macrophages were also found to be damaged, as were apoptotic epidermal cells. The suggested mechanism for ICSW is the formation of transverse (shear) waves at the interface between the aquatic medium and the skin surface. The waves, which are damped out within a few cell layers, give rise to shear stresses that, in turn, cause strains that act to separate between cells and damage some of the relatively weaker cells.

Ultrasound-facilitated transport of silver chloride (AgCl) particles in fish skin.

Electron-dense nano-particles in aqueous suspension were administered by immersion into the epidermis of fish using ultrasound in the therapeutic range. Enhanced permeability of the tissues to the particles was achieved by acoustic cavitation, which induced a controlled level of necrosis in the outer cell layers, and by non-cavitational exposures, which widened intercellular spaces of non-necrosed tissue in deeper regions of the epidermis. Both particle concentration and penetration depth were quantified using transmission electron microscopy. While cavitation-induced perforation was necessary for particles to penetrate into the tissues, non-cavitational exposures during immersions increased the particle flux towards the skin surface, as well as the diffusion rate of the particles within the epidermis and their depth of penetration. The technique described above may potentially be applied for non-stressful, mass-administration of substances into aquatic animals, as well as the relatively new field of ultrasound-facilitated delivery in moist epithelial tissues in humans.

Intramembrane Cavitation as a Predictive Bio-Piezoelectric Mechanism for Ultrasonic Brain Stimulation

Low-intensity ultrasonic waves can remotely and nondestructively excite central nervous system (CNS) neurons. While diverse applications for this effect are already emerging, the biophysical transduction mechanism underlying this excitation remains unclear. Recently, we suggested that ultrasound-induced intramembrane cavitation within the bilayer membrane could underlie the biomechanics of a range of observed acoustic bioeffects. In this paper, we show that, in CNS neurons, ultrasound-induced cavitation of these nanometric bilayer sonophores can induce a complex mechanoelectrical interplay leading to excitation, primarily through the effect of currents induced by membrane capacitance changes. Our model explains the basic features of CNS acoustostimulation and predicts how the experimentally observed efficacy of mouse motor cortical ultrasonic stimulation depends on stimulation parameters. These results support the hypothesis that neuronal intramembrane piezoelectricity underlies ultrasound-induced neurostimulation, and suggest that other interactions between the nervous system and pressure waves or perturbations could be explained by this new mode of biological piezoelectric transduction.

Behavioral response of carp (Cyprinus carpio) to ammonia stress

Abstract The behavioral responses of schools of young Koi fish ( Cyprinus carpio ) to sub-lethal ammonia concentrations were monitored using CCD cameras and computer image processing. Several geometrical parameters of the schools such as the position of the center of gravity, and the distribution of the fish were calculated continuously and plotted versus time. Swimming speed and activity were measured indirectly through a novel parameter—the Projected Mobility Picture (PMP). The immediate response of the treated fish was to dive to the bottom of the tank and stay there for a time period which increased with the ammonia concentration. Later, the fish rose to depths which increased with ammonia concentration. At high ammonia concentration, the fish approached the surface. Decreased activity of the treated fish was accompanied by a smaller mean distribution. Blood glucose levels of the treated fish increased, and they became indifferent to food. In general, this remote method of continuous monitoring of alterations in fish behavior under stress has the potential to be developed into a method for detecting early signs of stress in populations of fish.

Shear stress induced by a gas bubble pulsating in an ultrasonic field near a wall

Some of the effects that therapeutic ultrasound has in medicine and biology may be associated with steady oscillations of gas bubbles in liquid, very close to tissue surface. The bubble oscillations induce on the surface steady shear stress attributed to microstreaming. A mathematical simulation of the problem for both free and capsulated bubbles, known as contrast agents, is presented here. The simulation is based on a solution of Laplaces equation for potential flow and existing models for microstreaming. The solution for potential flow was obtained numerically using a boundary integral method. The solution provides the evolution of the bubble shape, the distribution of the velocity potential on the surface, and the shear stress along the surface. The simulation shows that significant shear stresses develop on the surface when the bubble bounces near the tissue surface. In this case, pressure amplitude of 20 kPa generates maximal steady shear stress of several kilo Pascal. Substantial shear stress on the tissue surface takes place inside a circular zone with a radius about half of the bubble radius. The predicted shear stress is greater than stress that causes hemolysis in blood and several orders of magnitude greater than the physiological stress induced on the vessel wall by the flowing blood.

Histoarchitecture of schistosomal granuloma development and involution: morphogenetic and biomechanical approaches

The authors present morphogenetic and biomechanical approaches on the concept of the Schistosoma mansoni granulomas, considering them as organoid structures that depend on cellular adhesion and sorting, forming rearrangement into hierarchical concentric layers, creating tension-dependent structures, aiming to acquire round form, since this is the minimal energy form, in which opposing forces pull in equally from all directions and are in balance. From the morphogenetic point of view, the granulomas function as little organs, presenting maturative and involutional stages in their development with final disappearance (pre-granulomatous stages, subdivided in: weakly and/or initial reactive and exudative; granulomatous stages: exudative-productive, productive and involutional). A model for the development of granulomas was suggested, according to the following stages: encapsulating, focal histolysis, fiber production, orientation and compacting and involution and disintegration. The authors concluded that schistosomal granuloma is not a tangled web of individual cells and fibers, but an organized structure composed by host and parasite components, which is not formed to attack the miracidia, but functions as an hybrid interface between two different phylogenetic beings.