High-voltage 10 ns delayed paired or bipolar pulses for in vitro bioelectric experiments

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Électromagnétisme pour le biomédical : étude des changements induits au niveau cellulaire par des impulsions thermiques générées par les ondes millimétriques

La partie inférieure de la bande de les ondes millimétriques (OMM, c'est-à-dire 20–100 GHz) constitue une alternative attrayante pour le traitement thermique non invasif du mélanome. Le chauffage pulsé induit électromagnétiquement peut entraîner des dommages plus importants dans les cellules par rapport au chauffage continu traditionnel. Dans ce travail, nous étudions les modifications induites au niveau cellulaire dans les cellules de mélanome à la suite d'une exposition au chauffage induit par en onde continue (CW) et ou en régime modulé (PWM) avec avec la même élévation de température moyenne, à 58.4 GHz. Premièrement, l’impact de la convection thermique sur la dynamique de la température dans des modèles représentant des conditions d’exposition in vitro typiques lors du chauffage par CW et PWM est étudié expérimentalement. Deuxièmement, la réponse au choc thermique, médiée par la phosphorylation d'une protéine de choc thermique (HSP27) et l'activation de Caspase-3, indicateur de l'apoptose cellulaire, est quantifiée pour surveiller la réponse biologique en utilisant une approche expérimentale basée sur la microscopie à fluorescence. Deux durées d'impulsion (1.5 s et 6 s) sont considérées. Nos résultats démontrent que les impulsions thermiques sont capables d'induire une réponse cellulaire plus forte dans les cellules de mélanome à la fois en termes de choc thermique et de mortalité cellulaire par rapport à celle induite en CW. Plus la durée de l'impulsion est courte, plus la réponse cellulaire est grande.The lower part of the millimeter wave (MMW) band (i.e., 20–100 GHz) is an attractive alternative for non-invasive thermal treatment of melanoma. Besides, pulsed electromagneticallyinduced heating can lead to stronger damage in cells compared to traditional continuous heating. In in-vitro experiments, continuous-wave (CW) or pulsed-wave (PW) amplitude-modulated MMW can be efficiently used to locally heat cell monolayers with a typical thickness ranging between 3 μm and 10 μm. In this work we investigate the modifications induced at the cellular level in melanoma cells following exposure to CW and PW MMW-induced heating with the same average temperature rise, at 58.4 GHz. First, the impact of thermal convection on temperature dynamics in models representing typical in vitro exposure conditions during CW and PW-induced heating is experimentally investigated. Second, the heat shock response, mediated by phosphorylation of a small heat shock protein (HSP27) and activation of Caspase-3, indicator of cellular apoptosis, are quantified to monitor the biological response using an experimental approach based on fluorescence microscopy. Two pulse durations (1.5 s and 6 s) are considered. Our results demonstrate that thermal pulses are able to induce a stronger cellular response in melanoma cells both in terms of heat shock and cellular mortality compared to the one induced by CW. The shorter the pulse duration, the greater the cellular response

Millimeter-Wave Pulsed Heating in Vitro: Effect of Pulse Duration

International audienceThe aim of this work is to compare the response of A375 melanoma cells following 90 min of exposure to trains of 1.5 or 6 s millimeter-waves (MMW)-induced thermal pulses with the same temperature dynamics. Phosphorylation of heat shock protein 27 (HSP27) and activation of cleaved Caspase-3 were used as markers of cellular stress and apoptosis, respectively. Immunofluorescence was used to observe and precisely quantify the cellular response as a function of the spatial distribution within the exposed area. Results show that cellular response was stronger when cells were exposed to a train of 1.5 s compared to 6 s heat pulses despite the same average temperature dynamics. Cellular apoptosis induced by 1.5 s pulses was about 50% greater compared to the one induced by 6 s pulses in the area of maximal thermal stress. Similarly, HSP27 phosphorylation was approximately 20% stronger than the one induced by 6 s pulses, and mainly focused within a small area of a few mm(2). Cellular response to MMW induced by pulsed heating does not only depend on the peak, average, and minimum temperature. It is a function of combination of the pulse parameters, including duration, peak power, and period. MMW-induced heat pulses can be efficiently used to induce cellular stress and apoptosis in melanoma cells as a promising innovative tool for the treatment of superficial skin cancer. Adaptative therapies might be envisaged by tuning the pulse shape as a function of the desired effect

High Voltage Generator With Adjustable Delay Between Two Nanosecond Pulses

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In vivo functional ultrasound (fUS) imaging of mice brain under radiofrequency exposure: preliminary dosimetric results

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Millimeter-wave Heating in vitro: Local Microscale Temperature Measurements Correlated to Heat Shock Cellular Response

International audienceObjective: Cellular sensitivity to heat is highly variable depending on the cell line. The aim of this paper is to assess the cellular sensitivity of the A375 melanoma cell line to continuous (CW) millimeter-waves (MMW) induced heating at 58.4 GHz, between 37 C and 47 C C to get a deeper insight into optimization of thermal treatment of superficial skin cancer. Methods: Phosphorylation of heat shock protein 27 (HSP27) was mapped within an area of about 30 mm2 to visualize the variation of heat-induced cellular stress as a function of the distance from the waveguide aperture (MMW radiation source). A multiphysics computational approach was then adopted to yield both electromagnetic and thermal field distributions as well as corresponding specific absorption rate (SAR) and temperature elevation. Induced temperature rise was experimentally measured using a micro-thermocouple (TC). Results: Coupling of the incident electromagnetic (EM) field with TC leads was first characterized, and optimal TC placing was identified. HSP27 phosphorylation was induced at temperatures 41 C, and its level increases as a function of the thermal dose delivered, remaining mostly focused within 3 mm2. Conclusion: Phosphorylation of HSP27 represents a valuable marker of cellular stress of A375 melanoma cells under MMW exposure, providing both quantitative and spatial information about the distribution of the thermal stress. Significance: These results may contribute to the design of thermal treatments of superficial melanoma through MMW-induced heating in the hyperthermic temperature range

In Vivo Functional Ultrasound (fUS) Real‐Time Imaging and Dosimetry of Mice Brain Under Radiofrequency Exposure

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Millimeter-wave pulsed heating in vitro: cell mortality and heat shock response

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Effects of Nanosecond Pulsed Electric Field (nsPEF) on a Multicellular Spheroid Tumor Model: Influence of Pulse Duration, Pulse Repetition Rate, Absorbed Energy, and Temperature

International audienceCellular response upon nsPEF exposure depends on different parameters, such as pulse number and duration, the intensity of the electric field, pulse repetition rate (PRR), pulsing buffer composition, absorbed energy, and local temperature increase. Therefore, a deep insight into the impact of such parameters on cellular response is paramount to adaptively optimize nsPEF treatment. Herein, we examined the effects of nsPEF ≤ 10 ns on long-term cellular viability and growth as a function of pulse duration (2–10 ns), PRR (20 and 200 Hz), cumulative time duration (1–5 µs), and absorbed electrical energy density (up to 81 mJ/mm3 in sucrose-containing low-conductivity buffer and up to 700 mJ/mm3 in high-conductivity HBSS buffer). Our results show that the effectiveness of nsPEFs in ablating 3D-grown cancer cells depends on the medium to which the cells are exposed and the PRR. When a medium with low-conductivity is used, the pulses do not result in cell ablation. Conversely, when the same pulse parameters are applied in a high-conductivity HBSS buffer and high PRRs are applied, the local temperature rises and yields either cell sensitization to nsPEFs or thermal damage

Dosimetry of Microelectrodes Array Chips for Electrophysiological Studies Under Simultaneous Radio Frequency Exposures

Studying the response of neuronal networks to radio frequency (RF) signals requires the use of a specific device capable of accessing and simultaneously recording neuronal activity during electromagnetic fields (EMF) exposure. In this study, a microelectrode array (MEA) that records the spontaneous activity of neurons is coupled to an open transverse electromagnetic (TEM) cell that propagates EMF. We characterize this system both numerically and experimentally at 1.8 GHz. Two MEA versions were compared, for the first time, to determine the impact of their design dissimilarities on the response to EMF. Macroscopic and microscopic measurements using, respectively, a fiber-optic probe and a temperature-dependent fluorescent dye (Rhodamine-B) were carried out. Results indicate that one MEA shows more stability toward the changes of the surrounding environment compared to the other MEA. Using a fiber-optic thermometer, the measured specific absorption rate (SAR) probe value in the center of the more stable MEA was 5.5 ± 2.3 W/kg. Using a Rhod-B microdosimetry technique, the measured SAR value at the level of the MEA electrodes was 7.0 ± 1.04 W/kg. SAR values are normalized per 1 W incident power. Due to the additional metallic planes and a smaller chip aperture, this new recording chip is steadier in terms of SAR and temperature stability allowing high exposure homogeneity as required during biological experiments. A typical neuronal activity recording under EMF exposure is reported