Jean-Christophe Béra

Piezoelectric osteotomy: a new technique for bone surgery-advantages in craniofacial surgery.

AbstractIntroductionUltrasonic bone cutting is a new surgical technique used in dentistry to section hard tissues without damaging adjacent soft tissues. We hypothesized that such a device could also be useful in craniofacial surgery, particularly during the removal of the superior orbital roof during craniofaciostenosis surgery.Materials and methodsAn ultrasonic device was employed in different craniofacial surgical procedures: 1.to remove the superior orbital roof in 30 cases of craniofaciostenosis,2.to perform a Le Fort III osteotomy for the treatment of Crouzon syndrome in two patients,3.to cut the parietal and frontal bone in 30 cases of craniofaciostenosis. The integrity of soft tissues and surgical time was evaluated.ResultsFunctional results were good without any soft tissue damage appreciated. The overall operative time, however, was increased.ConclusionsPiezosurgery is a new technical procedure, which can be advantageous for bone cutting in multiple situations with minimal to no damage in adjacent soft tissues.

Transfection of cells in suspension by ultrasound cavitation

Sonoporation holds many promises in developing an efficient, reproducible and permanent gene delivery vector. In this study, we evaluated sonoporation as a method to transfect nucleic acids in suspension cells, including the human follicular lymphoma cell line RL and fresh human Chronic Lymphocytic Leukemia (CLL) cells. RL and CLL cells were exposed to continuous ultrasound waves (445 kHz) in the presence of either plasmid DNA coding for green fluorescent protein (GFP) or fluorescent siRNA directed against BCL2L1. Transfection efficiency and cell viability were assessed using fluorescent microscopy and flow cytometry analysis, respectively. Knock-down of target protein by siRNA was assessed by immunoblotting. Moreover, sonoporation was used to stably transfect RL cells with a plasmid coding for luciferase (pGL3). These cells were then used for the non-invasive monitoring of tumorigenesis in immunodeficient SCID mice. Sonoporation allows a highly efficient transfection of nucleic acid in suspension cells with a low rate of mortality, both in a tumor cell line and in fresh human leukemic cells. It also allowed efficient transfection of BCL2L1 siRNA with efficient reduction of the target protein level. In conclusion, ultrasound cavitation represents an efficient method for the transfection of cells in suspension, including fresh human leukemic cells.

Effect of spectral smearing on the perceptual segregation of vowel sequences

Although segregation of both simultaneous and sequential speech items may be involved in the reception of speech in noisy environments, research on the latter is relatively sparse. Further, previous studies examining the ability of hearing-impaired listeners to form distinct auditory streams have produced mixed results. Finally, there is little work investigating streaming in cochlear implant recipients, who also have poor frequency resolution. The present study focused on the mechanisms involved in the segregation of vowel sequences and potential limitations to segregation associated with poor frequency resolution. An objective temporal-order paradigm was employed in which listeners reported the order of constituent vowels within a sequence. In Experiment 1, it was found that fundamental frequency based mechanisms contribute to segregation. In Experiment 2, reduced frequency tuning often associated with hearing impairment was simulated in normal-hearing listeners. In that experiment, it was found that spectral smearing of the vowels increased accurate identification of their order, presumably by reducing the tendency to form separate auditory streams. These experiments suggest that a reduction in spectral resolution may result in a reduced ability to form separate auditory streams, which may contribute to the difficulties of hearing-impaired listeners, and probably cochlear implant recipients as well, in multi-talker cocktail-party situations.

In vitro sonodynamic cytotoxicity in regulated cavitation conditions.

Sonodynamic toxicity has always been linked to the cavitation phenomenon. In this work, sonodynamic effect with Photofrin was evaluated with a new ultrasound device: a regulated cavitation generator. In this way, acoustic intensity was substituted with cavitation level as ultrasound parameter. Photofrin potentiated significantly the cavitation cytotoxicity even for low setpoints where no inertial cavitation appeared. Therefore sonodynamic mechanism was principally mechanical, facilitated by the Photofrin insertion in cellular cytoplasmic membranes. This assertion was also supported by the fact that sonodynamic cytotoxicity was independent from the Photofrin presence or absence in the extracellular medium. Reproducible sonodynamic efficiency was perfectly obtained with this new regulated cavitation generator.

Experimental Assessment of Calvarial Bone Defect Re-Ossification Stimulation Using Low-Intensity Pulsed Ultrasound

Ultrasound treatment has been proposed by several authors to enhance the repair of long bone injury. The present study investigated in a murine model the treatment by low-intensity pulsed ultrasound (LIPUS) of calvarial flat bone defect. The animals were operated to create bone defect and exposed to ultrasound for 5 min per day, 5 d per week, during two weeks. Two intensities of ultrasound (1 MHz, 100 Hz pulse repetition frequency and 20% duty cycle) were investigated: 100 and 300 mW/cm(2) spatial-averaged, time-averaged. Re-ossification surface and volume were determined after 30 and 60 days using computerized X-ray tomography in all animals of the control and treated groups. The results showed a significant increase of bone re-ossification in the group treated with the higher-intensity ultrasound (mean value of 18% volume reconstruction), whereas lower bone reconstruction was observed in the lower-intensity and control groups (respective mean values of 10 and 12% volume reconstruction).

Feedback loop process to control acoustic cavitation.

Applications involving acoustic cavitation mechanisms, such as sonoporation, are often poorly reproducible because of the unstationary behavior of cavitation. For this purpose, this study proposes to work at a fixed cavitation level instead of a fixed acoustic intensity. A regulated cavitation generator has been developed in an in vitro configuration of standing wave field. This system implements the regulation of the cavitation level during sonication by modulating the applied acoustic intensity with a feedback loop based on acoustic measurements. The experimental setup consists of a plane piezoelectric transducer for sonication (continuous wave, frequency 445 kHz) and a hydrophone pointing to the sonicated medium. The cavitation level is quantified every 5 ms from a spectral analysis of the acoustic signal. The results show that the regulation device generates reproducible mean cavitation levels with a standard deviation lower than 1.6% in the applied intensity range (from 0.12 to 3.44 W/cm(2)), while this standard deviation can reach 76% without regulation. The feedback loop process imposes precise cavitation level even in low applied acoustic intensity.

Streaming of vowel sequences based on fundamental frequency in a cochlear-implant simulation.

Cochlear-implant (CI) users often have difficulties perceiving speech in noisy environments. Although this problem likely involves auditory scene analysis, few studies have examined sequential segregation in CI listening situations. The present study aims to assess the possible role of fundamental frequency (F(0)) cues for the segregation of vowel sequences, using a noise-excited envelope vocoder that simulates certain aspects of CI stimulation. Obligatory streaming was evaluated using an order-naming task in two experiments involving normal-hearing subjects. In the first experiment, it was found that streaming did not occur based on F(0) cues when natural-duration vowels were processed to reduce spectral cues using the vocoder. In the second experiment, shorter duration vowels were used to enhance streaming. Under these conditions, F(0)-related streaming appeared even when vowels were processed to reduce spectral cues. However, the observed segregation could not be convincingly attributed to temporal periodicity cues. A subsequent analysis of the stimuli revealed that an F(0)-related spectral cue could have elicited the observed segregation. Thus, streaming under conditions of severely reduced spectral cues, such as those associated with CIs, may potentially occur as a result of this particular cue.

Promoting inertial cavitation by nonlinear frequency mixing in a bifrequency focused ultrasound beam

Enhancing cavitation activity with minimal acoustic intensities could be interesting in a variety of therapeutic applications where mechanical effects of cavitation are needed with minimal heating of surrounding tissues. The present work focuses on the relative efficiency of a signal combining two neighbouring frequencies and a one-frequency signal for initiating ultrasound inertial cavitation. Experiments were carried out in a water tank, using a 550kHz piezoelectric composite spherical transducer focused on targets with 46μm roughness. The acoustic signal scattered, either by the target or by the cavitation bubbles, is filtered using a spectral and cepstral-like method to obtain an inertial cavitation activity measurement. The ultrasound excitations consist of 1.8ms single bursts of single frequency f(0)=550kHz excitation, in the monofrequency case, and of dual frequency f(1)=535kHz and f(2)=565kHz excitation, in the bifrequency case. It is shown that depending on the value of the monofrequency cavitation threshold intensity the bifrequency excitation can increase or reduce the cavitation threshold. The analysis of the thresholds indicates that the mechanisms involved are nonlinear. The progress of the cavitation activity beyond the cavitation threshold is also studied. The slope of the cavitation activity considered as a function of the acoustic intensity is always steeper in the case of the bifrequency excitation. This means that the delimitation of the region where cavitation occurs should be cleaner than with a classical monofrequency excitation.

Stabilizing in vitro ultrasound-mediated gene transfection by regulating cavitation.

It is well known that acoustic cavitation can facilitate the inward transport of genetic materials across cell membranes (sonoporation). However, partially due to the unstationary behavior of the initiation and leveling of cavitation, the sonoporation effect is usually unstable, especially in low intensity conditions. A system which is able to regulate the cavitation level during sonication by modulating the applied acoustic intensity with a feedback loop is implemented and its effect on in vitro gene transfection is tested. The regulated system provided better time stability and reproducibility of the cavitation levels than the unregulated conditions. Cultured hepatoma cells (BNL) mixed with 10 μg luciferase plasmids are exposed to 1-MHz pulsed ultrasound with or without cavitation regulation, and the gene transfection efficiency and cell viability are subsequently assessed. Experimental results show that for all exposure intensities (low, medium, and high), stable and intensity dependent, although not higher, gene expression could be achieved in the regulated cavitation system than the unregulated conditions. The cavitation regulation system provides a better control of cavitation and its bioeffect which are crucial important for clinical applications of ultrasound-mediated gene transfection.

A time splitting projection scheme for compressible two-phase flows. Application to the interaction of bubbles with ultrasound waves

This paper is focused on the numerical simulation of the interaction of an ultrasound wave with a bubble. Our interest is to develop a fully compressible solver in the two phases and to account for surface tension effects.As the volume oscillation of the bubble occurs in a low Mach number regime, a specific care must be paid to the effectiveness of the numerical method which is chosen to solve the compressible Euler equations. Three different numerical solvers, an explicit HLLC (Harten-Lax-van Leer-Contact) solver 48, a preconditioning explicit HLLC solver 14 and the compressible projection method 21,53,55, are described and assessed with a one dimensional spherical benchmark. From this preliminary test, we can conclude that the compressible projection method outclasses the other two, whether the spatial accuracy or the time step stability are considered.Multidimensional numerical simulations are next performed. As a basic implementation of the surface tension leads to strong spurious currents and numerical instabilities, a specific velocity/pressure time splitting is proposed to overcome this issue. Numerical evidences of the efficiency of this new numerical scheme are provided, since both the accuracy and the stability of the overall algorithm are enhanced if this new time splitting is used. Finally, the numerical simulation of the interaction of a moving and deformable bubble with a plane wave is presented in order to bring out the ability of the new method in a more complex situation.