The Experts below are selected from a list of 1473 Experts worldwide ranked by ideXlab platform

Dong Zhang - One of the best experts on this subject based on the ideXlab platform.

  • mechanisms underlying Sonoporation interaction between microbubbles and cells
    Ultrasonics Sonochemistry, 2020
    Co-Authors: Yanye Yang, Xiasheng Guo, Dong Zhang
    Abstract:

    The past several decades have witnessed great progress in "smart drug delivery", an advance technology that can deliver genes or drugs into specific locations of patients' body with enhanced delivery efficiency. Ultrasound-activated mechanical force induced by the interactions between microbubbles and cells, which can stimulate so-called "Sonoporation" process, has been regarded as one of the most promising candidates to realize spatiotemporal-controllable drug delivery to selected regions. Both experimental and numerical studies were performed to get in-depth understanding on how the microbubbles interact with cells during Sonoporation processes, under different impact parameters. The current work gives an overview of the general mechanism underlying microbubble-mediated Sonoporation, and the possible impact factors (e.g., the properties of cavitation agents and cells, acoustical driving parameters and bubble/cell micro-environment) that could affect Sonoporation outcomes. Finally, current progress and considerations of Sonoporation in clinical applications are reviewed also.

  • Sonoporation induced cell membrane permeabilization and cytoskeleton disassembly at varied acoustic and microbubble cell parameters
    Scientific Reports, 2018
    Co-Authors: Maochen Wang, Yi Zhang, Chenliang Cai, Xiasheng Guo, Dong Zhang
    Abstract:

    Sonoporation mediated by microbubbles has being extensively studied as a promising technique to facilitate gene/drug delivery to cells. Previous studies mainly explored the membrane-level responses to Sonoporation. To provide in-depth understanding on this process, various Sonoporation-induced cellular responses (e.g., membrane permeabilization and cytoskeleton disassembly) generated at different impact parameters (e.g., acoustic driving pressure and microbubble-cell distances) were systemically investigated in the present work. HeLa cells, whose α-tubulin cytoskeleton was labeled by incorporation of a green fluorescence protein (GFP)-α-tubulin fusion protein, were exposed to a single ultrasound pulse (1 MHz, 20 cycles) in the presence of microbubbles. Intracellular transport via Sonoporation was assessed in real time using propidium iodide and the disassembly of α-tubulin cytoskeleton was observed by fluorescence microscope. Meanwhile, the dynamics of an interacting bubble-cell pair was theoretically simulated by boundary element method. Both the experimental observations and numerical simulations showed that, by increasing the acoustic pressure or reducing the bubble-cell distance, intensified deformation could be induced in the cellular membrane, which could result in enhanced intracellular delivery and cytoskeleton disassembly. The current results suggest that more tailored therapeutic strategies could be designed for ultrasound gene/drug delivery by adopting optimal bubble-cell distances and/or better controlling incident acoustic energy.

  • cell cycle specific cellular responses to Sonoporation
    Theranostics, 2017
    Co-Authors: Pengfei Fan, Maochen Wang, Yi Zhang, Chenliang Cai, Xiasheng Guo, Dong Zhang, Dongxin Yang, Lawrence A Crum
    Abstract:

    Microbubble-mediated Sonoporation has shown its great potential in facilitating intracellular uptake of gene/drugs and other therapeutic agents that are otherwise difficult to enter cells. However, the biophysical mechanisms underlying microbubble-cell interactions remain unclear. Particularly, it is still a major challenge to get a comprehensive understanding of the impact of cell cycle phase on the cellular responses simultaneously occurring in cell membrane and cytoskeleton induced by microbubble Sonoporation. Methods: Here, efficient synchronizations were performed to arrest human cervical epithelial carcinoma (HeLa) cells in individual cycle phases. The, topography and stiffness of synchronized cells were examined using atomic force microscopy. The variations in cell membrane permeabilization and cytoskeleton arrangement induced by Sonoporation were analyzed simultaneously by a real-time fluorescence imaging system. Results: The results showed that G1-phase cells typically had the largest height and elastic modulus, while S-phase cells were generally the flattest and softest ones. Consequently, the S-Phase was found to be the preferred cycle for instantaneous Sonoporation treatment, due to the greatest enhancement of membrane permeability and the fastest cytoskeleton disassembly at the early stage after Sonoporation. Conclusion: The current findings may benefit ongoing efforts aiming to pursue rational utilization of microbubble-mediated Sonoporation in cell cycle-targeted gene/drug delivery for cancer therapy.

  • interaction between cavitation microbubble and cell a simulation of Sonoporation using boundary element method bem
    Ultrasonics Sonochemistry, 2017
    Co-Authors: Xiasheng Guo, Chenliang Cai, Dong Zhang, Yanye Yang, Pintong Huang
    Abstract:

    Sonoporation has been widely accepted as a significant tool for gene delivery as well as some bio-effects like hemolysis, bringing in high demands of looking into its underlying mechanism. A two-dimensional (2D) boundary element method (BEM) model was developed to investigate microbubble-cell interaction, especially the morphological and mechanical characteristics around the close-to-bubble point (CP) on cell membrane. Based on time evolution analysis of Sonoporation, detailed information was extracted from the model for analysis, including volume expansion ratio of the bubble, areal expansion ratio of the cell, jet velocity and CP displacement. Parametric studies were carried out, revealing the influence of different ultrasound parameters (i.e., driving frequency and acoustic pressure) and geometrical configurations (i.e., bubble-cell distance and initial bubble radius). This model could become a powerful tool not only for understanding bubble-cell interactions, but also for optimizing the strategy of Sonoporation, such that it could be safer and of higher efficiency for biological and medical studies especially in clinics.

  • microbubble induced Sonoporation involved in ultrasound mediated dna transfection in vitro at low acoustic pressures
    Journal of Biomechanics, 2012
    Co-Authors: Yuanyuan Qiu, Chunbing Zhang, Dong Zhang
    Abstract:

    Abstract In the present work, human breast cancer cells MCF-7 mixed with polyethylenimine: deoxyribonucleic acid complex and microbubbles were exposed to 1-MHz ultrasound at low acoustic driving pressures ranging from 0.05 to 0.3 MPa. The Sonoporation pores generated on the cell membrane were examined with scanning electron microscopy. The transfection efficiency and cell viability were evaluated with flow cytometry. The results showed that ultrasound sonication under the current exposure condition could generate cell pores with mean size ranging from about 100 nm to 1.25 μm, and that larger Sonoporation pores would be generated with the increasing acoustic pressure or longer treatment time, leading to the enhancement of transfection efficiency and the reduction of cell viability. The simulations based on the Marmottant model were performed to test the hypothesis that the microstreaming-induced shear stress might be involved in the mechanisms of the low-intensity ultrasound induced Sonoporation. The calculated shear stress resulting from the micro-streaming ranged from 15 to 680 Pa corresponding to the applied acoustic pressures 0.05−0.3 MPa, which is sufficient to induce reversible Sonoporation. This study indicates that the shear stress related bio-effects may provide a base for strategies aimed at targeted drug delivery.

N. De Jong - One of the best experts on this subject based on the ideXlab platform.

  • Opening of endothelial cell–cell contacts due to Sonoporation
    'Elsevier BV', 2020
    Co-Authors: Vegter M., N. De Jong, Lattwein K.r., Mastik F., Beurskens R., Steen, A.f.w. Van Der, Kooiman K.
    Abstract:

    Ultrasound insonification of microbubbles can locally increase vascular permeability to enhance drug delivery. To control and optimize the therapeutic potential, we need to better understand the underlying biological mechanisms of the drug delivery pathways. The aim of this in vitro study was to elucidate the microbubble-endothelial cell interaction using the Brandaris 128 ultra-high-speed camera (up to 25 Mfps) coupled to a custom-built Nikon confocal microscope, to visualize both microbubble oscillation and the cellular response. Sonoporation and opening of cell-cell contacts by single αVβ3-targeted microbubbles (n = 152) was monitored up to 4 min after ultrasound insonification (2 MHz, 100–400 kPa, 10 cycles). Sonoporation occurred when microbubble excursion amplitudes exceeded 0.7 μm. Quantification of the influx of the fluorescent model drug propidium iodide upon Sonoporation showed that the size of the created pore increased for larger microbubble excursion amplitudes. Microbubble-mediated opening of cell-cell contacts occurred as a cellular response upon Sonoporation and did not correlate with the microbubble excursion amplitude itself. The initial integrity of the cell-cell contacts affected the susceptibly to drug delivery, since cell-cell contacts opened more often when cells were only partially attached to their neighbors (48%) than when fully attached (14%). The drug delivery outcomes were independent of nonlinear microbubble behavior, microbubble location, and cell size. In conclusion, by studying the microbubble–cell interaction at nanosecond and nanometer resolution the relationship between drug delivery pathways and their underlying mechanisms was further unraveled. These novel insights will aid the development of safe and efficient microbubble-mediated drug delivery

  • opening of endothelial cell cell contacts due to Sonoporation
    Journal of Controlled Release, 2020
    Co-Authors: Ines Beekers, N. De Jong, Kirby R Lattwein, Antonius F W Van Der Steen, Merel Vegter, Frits Mastik, Robert Beurskens, Martin D Verweij
    Abstract:

    Ultrasound insonification of microbubbles can locally increase vascular permeability to enhance drug delivery. To control and optimize the therapeutic potential, we need to better understand the underlying biological mechanisms of the drug delivery pathways. The aim of this in vitro study was to elucidate the microbubble-endothelial cell interaction using the Brandaris 128 ultra-high-speed camera (up to 25 Mfps) coupled to a custom-built Nikon confocal microscope, to visualize both microbubble oscillation and the cellular response. Sonoporation and opening of cell-cell contacts by single αVβ3-targeted microbubbles (n = 152) was monitored up to 4 min after ultrasound insonification (2 MHz, 100–400 kPa, 10 cycles). Sonoporation occurred when microbubble excursion amplitudes exceeded 0.7 μm. Quantification of the influx of the fluorescent model drug propidium iodide upon Sonoporation showed that the size of the created pore increased for larger microbubble excursion amplitudes. Microbubble-mediated opening of cell-cell contacts occurred as a cellular response upon Sonoporation and did not correlate with the microbubble excursion amplitude itself. The initial integrity of the cell-cell contacts affected the susceptibly to drug delivery, since cell-cell contacts opened more often when cells were only partially attached to their neighbors (48%) than when fully attached (14%). The drug delivery outcomes were independent of nonlinear microbubble behavior, microbubble location, and cell size. In conclusion, by studying the microbubble–cell interaction at nanosecond and nanometer resolution the relationship between drug delivery pathways and their underlying mechanisms was further unraveled. These novel insights will aid the development of safe and efficient microbubble-mediated drug delivery.

  • targeted microbubble mediated Sonoporation of endothelial cells in vivo
    IEEE Transactions on Ultrasonics Ferroelectrics and Frequency Control, 2014
    Co-Authors: Ilya Skachkov, N. De Jong, Ying Luan, Antonius F W Van Der Steen, Klazina Kooiman
    Abstract:

    textabstractUltrasound contrast agents as drug-delivery systems are an emerging field. Recently, we reported that targeted microbubbles are able to sonoporate endothelial cells in vitro. In this study, we investigated whether targeted microbubbles can also induce Sonoporation of endothelial cells in vivo, thereby making it possible to combine molecular imaging and drug delivery. Live chicken embryos were chosen as the in vivo model. αvβ3-targeted microbubbles attached to the vessel wall of the chicken embryo were insonified at 1 MHz at 150 kPa (1 × 10 000 cycles) and at 200 kPa (1 × 1000 cycles) peak negative acoustic pressure. Sonoporation was studied by intravital microscopy using the model drug propidium iodide (PI). Endothelial cell PI uptake was observed in 48% of microbubble-vessel-wall complexes at 150 kPa (n = 140) and in 33% at 200 kPa (n = 140). Efficiency of PI uptake depended on the local targeted microbubble concentration and increased up to 80% for clusters of 10 to 16 targeted microbubbles. Ultrasound or targeted microbubbles alone did not induce PI uptake. This intravital microscopy study reveals that Sonoporation can be visualized and induced in vivo using targeted microbubbles.

  • Sonoporation of endothelial cells by vibrating targeted microbubbles
    Journal of Controlled Release, 2011
    Co-Authors: Klazina Kooiman, N. De Jong, Antonius F W Van Der Steen, Miranda Foppenharteveld
    Abstract:

    Molecular imaging using ultrasound makes use of targeted microbubbles. In this study we investigated whether these microbubbles could also be used to induce Sonoporation in endothelial cells. Lipid-coated microbubbles were targeted to CD31 and insonified at 1 MHz at low peak negative acoustic pressures at six sequences of 10 cycle sine-wave bursts. Vibration of the targeted microbubbles was recorded with the Brandaris-128 high-speed camera (~ 13 million frames per second). In total, 31 cells were studied that all had one microbubble (1.2–4.2 micron in diameter) attached per cell. After insonification at 80 kPa, 30% of the cells (n = 6) had taken up propidium iodide, while this was 20% (n = 1) at 120 kPa and 83% (n = 5) at 200 kPa. Irrespective of the peak negative acoustic pressure, uptake of propidium iodide was observed when the relative vibration amplitude of targeted microbubbles was greater than 0.5. No relationship was found between the position of the microbubble on the cell and induction of Sonoporation. This study shows that targeted microbubbles can also be used to induce Sonoporation, thus making it possible to combine molecular imaging and drug delivery

Cheri X Deng - One of the best experts on this subject based on the ideXlab platform.

  • visualization and quantification of dynamic intercellular coupling in human embryonic stem cells using single cell Sonoporation
    Scientific Reports, 2020
    Co-Authors: Zhenzhen Fan, Xufeng Xue, Cheri X Deng
    Abstract:

    Gap junctions (GJs), which are proteinaceous channels, couple adjacent cells by permitting direct exchange of intracellular molecules with low molecular weights. GJ intercellular communication (GJIC) plays a critical role in regulating behaviors of human embryonic stem cells (hESCs), affecting their proliferation and differentiation. Here we report a novel use of Sonoporation that enables single cell intracellular dye loading and dynamic visualization/quantification of GJIC in hESC colonies. By applying a short ultrasound pulse to excite single microbubbles tethered to cell membranes, a transient pore on the cell membrane (Sonoporation) is generated which allows intracellular loading of dye molecules and influx of Ca2+ into single hESCs. We employ live imaging for continuous visualization of intercellular dye transfer and Ca2+ diffusion in hESC colonies. We quantify cell-cell permeability based on dye diffusion using mass transport models. Our results reveal heterogeneous intercellular connectivity and a variety of spatiotemporal characteristics of intercellular Ca2+ waves in hESC colonies induced by Sonoporation of single cells.

  • characterization of the dynamic activities of a population of microbubbles driven by pulsed ultrasound exposure in Sonoporation
    Ultrasound in Medicine and Biology, 2014
    Co-Authors: Di Chen, Cheri X Deng
    Abstract:

    Abstract Ultrasound-driven microbubble activities have been exploited to transiently disrupt the cell membrane (Sonoporation) for non-viral intracellular drug delivery and gene transfection both in vivo and in vitro . In this study, we investigated the dynamic behaviors of a population of microbubbles exposed to pulsed ultrasound and their impact on adherent cells in terms of intracellular delivery and cell viability. By systematically analyzing the bubble activities at time scales relevant to pulsed ultrasound exposure, we identified two quantification parameters that categorize the diverse bubble activities subjected to various ultrasound conditions into three characteristic behaviors: stable cavitation/aggregation (type I), growth/coalescence and translation (type II) and localized inertial cavitation/collapse (type III). Correlation of the bubble activities with Sonoporation outcome suggested that type III behavior resulted in intracellular delivery, whereas type II behavior caused the death of a large number of cells. These results provide useful insights for rational selection of ultrasound parameters to optimize outcomes of Sonoporation and other applications that exploit the use of ultrasound-driven bubble activities.

  • spatiotemporally controlled single cell Sonoporation
    Proceedings of the National Academy of Sciences of the United States of America, 2012
    Co-Authors: Zhenzhen Fan, Haiyan Liu, Michael Mayer, Cheri X Deng
    Abstract:

    This paper presents unique approaches to enable control and quantification of ultrasound-mediated cell membrane disruption, or Sonoporation, at the single-cell level. Ultrasound excitation of microbubbles that were targeted to the plasma membrane of HEK-293 cells generated spatially and temporally controlled membrane disruption with high repeatability. Using whole-cell patch clamp recording combined with fluorescence microscopy, we obtained time-resolved measurements of single-cell Sonoporation and quantified the size and resealing rate of pores. We measured the intracellular diffusion coefficient of cytoplasmic RNA/DNA from Sonoporation-induced transport of an intercalating fluorescent dye into and within single cells. We achieved spatiotemporally controlled delivery with subcellular precision and calcium signaling in targeted cells by selective excitation of microbubbles. Finally, we utilized Sonoporation to deliver calcein, a membrane-impermeant substrate of multidrug resistance protein-1 (MRP1), into HEK-MRP1 cells, which overexpress MRP1, and monitored the calcein efflux by MRP1. This approach made it possible to measure the efflux rate in individual cells and to compare it directly to the efflux rate in parental control cells that do not express MRP1.

  • effects of shear stress cultivation on cell membrane disruption and intracellular calcium concentration in Sonoporation of endothelial cells
    Journal of Biomechanics, 2011
    Co-Authors: Juyoung Park, Zhenzhen Fan, Cheri X Deng
    Abstract:

    Abstract Microbubble facilitated ultrasound (US) application can enhance intracellular delivery of drugs and genes in endothelial cells cultured in static condition by transiently disrupting the cell membrane, or Sonoporation. However, endothelial cells in vivo that are constantly exposed to blood flow may exhibit different Sonoporation characteristics. This study investigates the effects of shear stress cultivation on Sonoporation of endothelial cells in terms of membrane disruption and changes in the intracellular calcium concentration ([Ca2+]i). Sonoporation experiments were conducted using murine brain microvascular endothelial (bEnd.3) cells and human umbilical vein endothelial cells (HUVECs) cultured under static or shear stress (5 dyne/cm2 for 5 days) condition in a microchannel environment. The cells were exposed to a short US tone burst (1.25 MHz, 8 μs duration, 0.24 MPa) in the presence of DefinityTM microbubbles to facilitate Sonoporation. Membrane disruption was assessed by propidium iodide (PI) and changes in [Ca2+]i measured by fura-2AM. Results from this study show that shear stress cultivation significantly reduced the impact of ultrasound-driven microbubbles activities on endothelial cells. Cells cultured under shear stress condition exhibited much lower percentage with membrane disruption and changes in [Ca2+]i compared to statically cultured cells. The maximum increases of PI uptake and [Ca2+]i were also significantly lower in the shear stress cultured cells. In addition, the extent of [Ca2+]i waves in shear cultured HUVECs was reduced compared to the statically cultured cells.

  • the size of Sonoporation pores on the cell membrane
    Ultrasound in Medicine and Biology, 2009
    Co-Authors: Yun Zhou, Jianmin Cui, Ronald E Kumon, Cheri X Deng
    Abstract:

    Sonoporation uses ultrasound (US) to generate transient nonselective pores on the cell membrane and has been exploited as a nonviral intracellular drug and gene delivery strategy. The pore size determines the size of agents that can be delivered into the cytoplasm using the technique. However, measurements of the dynamic, submicron-scale pores have not been readily available. Electron microscopy or atomic force microscopy has been used to gauge pore size but such techniques are intrinsically limited to post-US measurements that may not accurately reveal the relevant information. As previously demonstrated, changes of the transmembrane current (TMC) of a single cell under voltage clamp can be used for monitoring Sonoporation in real-time. Because the TMC is related to the diffusion of ions through the pores on the membrane, it can potentially provide information of the pore size generated in Sonoporation. Using Xenopus laevis oocytes as the model system, the TMC of single cells under voltage clamp was measured in real-time to assess formation of pores on the membrane in Sonoporation. The cells were exposed to US (0.2 s, 0.3 MPa, 1.075 MHz) in the presence of Definity microbubbles. Experiments were designed to obtain the TMC corresponding to a single pore on the membrane. The size of the pores was estimated from an electro-diffusion model that relates the TMC with pore size from the ion transport through the pores on the membrane. The mean radius of single pores was determined to be 110 nm with standard deviation of 40 nm. This study reports the first results of pore size from the TMC measured using the voltage clamp technique.

Antonius F W Van Der Steen - One of the best experts on this subject based on the ideXlab platform.

  • opening of endothelial cell cell contacts due to Sonoporation
    Journal of Controlled Release, 2020
    Co-Authors: Ines Beekers, N. De Jong, Kirby R Lattwein, Antonius F W Van Der Steen, Merel Vegter, Frits Mastik, Robert Beurskens, Martin D Verweij
    Abstract:

    Ultrasound insonification of microbubbles can locally increase vascular permeability to enhance drug delivery. To control and optimize the therapeutic potential, we need to better understand the underlying biological mechanisms of the drug delivery pathways. The aim of this in vitro study was to elucidate the microbubble-endothelial cell interaction using the Brandaris 128 ultra-high-speed camera (up to 25 Mfps) coupled to a custom-built Nikon confocal microscope, to visualize both microbubble oscillation and the cellular response. Sonoporation and opening of cell-cell contacts by single αVβ3-targeted microbubbles (n = 152) was monitored up to 4 min after ultrasound insonification (2 MHz, 100–400 kPa, 10 cycles). Sonoporation occurred when microbubble excursion amplitudes exceeded 0.7 μm. Quantification of the influx of the fluorescent model drug propidium iodide upon Sonoporation showed that the size of the created pore increased for larger microbubble excursion amplitudes. Microbubble-mediated opening of cell-cell contacts occurred as a cellular response upon Sonoporation and did not correlate with the microbubble excursion amplitude itself. The initial integrity of the cell-cell contacts affected the susceptibly to drug delivery, since cell-cell contacts opened more often when cells were only partially attached to their neighbors (48%) than when fully attached (14%). The drug delivery outcomes were independent of nonlinear microbubble behavior, microbubble location, and cell size. In conclusion, by studying the microbubble–cell interaction at nanosecond and nanometer resolution the relationship between drug delivery pathways and their underlying mechanisms was further unraveled. These novel insights will aid the development of safe and efficient microbubble-mediated drug delivery.

  • viability of endothelial cells after ultrasound mediated Sonoporation influence of targeting oscillation and displacement of microbubbles
    Journal of Controlled Release, 2016
    Co-Authors: Tom Van Rooij, Ilya Skachkov, Ines Beekers, Kirby R Lattwein, Jason Voorneveld, Tom J A Kokhuis, Deep Bera, Ying Luan, Antonius F W Van Der Steen
    Abstract:

    Microbubbles (MBs) have been shown to create transient or lethal pores in cell membranes under the influence of ultrasound, known as ultrasound-mediated Sonoporation. Several studies have reported enhanced drug delivery or local cell death induced by MBs that are either targeted to a specific biomarker (targeted microbubbles, tMBs) or that are not targeted (non-targeted microbubbles, ntMBs). However, both the exact mechanism and the optimal acoustic settings for Sonoporation are still unknown. In this study we used real-time uptake patterns of propidium iodide, a fluorescent cell impermeable model drug, as a measure for Sonoporation. Combined with high-speed optical recordings of MB displacement and ultra-high-speed recordings of MB oscillation, we aimed to identify differences in MB behavior responsible for either viable Sonoporation or cell death. We compared ntMBs and tMBs with identical shell compositions exposed to long acoustic pulses (500-50,000cycles) at various pressures (150-500kPa). Propidium iodide uptake highly correlated with cell viability; when the fluorescence intensity still increased 120s after opening of the pore, this resulted in cell death. Higher acoustic pressures and longer cycles resulted in more displacing MBs and enhanced Sonoporation. Non-displacing MBs were found to be the main contributor to cell death, while displacement of tMBs enhanced reversible Sonoporation and preserved cell viability. Consequently, each therapeutic application requires different settings: non-displacing ntMBs or tMBs are advantageous for therapies requiring cell death, especially at 500kPa and 50,000cycles, whereas short acoustic pulses causing limited displacement should be used for drug delivery.

  • targeted microbubble mediated Sonoporation of endothelial cells in vivo
    IEEE Transactions on Ultrasonics Ferroelectrics and Frequency Control, 2014
    Co-Authors: Ilya Skachkov, N. De Jong, Ying Luan, Antonius F W Van Der Steen, Klazina Kooiman
    Abstract:

    textabstractUltrasound contrast agents as drug-delivery systems are an emerging field. Recently, we reported that targeted microbubbles are able to sonoporate endothelial cells in vitro. In this study, we investigated whether targeted microbubbles can also induce Sonoporation of endothelial cells in vivo, thereby making it possible to combine molecular imaging and drug delivery. Live chicken embryos were chosen as the in vivo model. αvβ3-targeted microbubbles attached to the vessel wall of the chicken embryo were insonified at 1 MHz at 150 kPa (1 × 10 000 cycles) and at 200 kPa (1 × 1000 cycles) peak negative acoustic pressure. Sonoporation was studied by intravital microscopy using the model drug propidium iodide (PI). Endothelial cell PI uptake was observed in 48% of microbubble-vessel-wall complexes at 150 kPa (n = 140) and in 33% at 200 kPa (n = 140). Efficiency of PI uptake depended on the local targeted microbubble concentration and increased up to 80% for clusters of 10 to 16 targeted microbubbles. Ultrasound or targeted microbubbles alone did not induce PI uptake. This intravital microscopy study reveals that Sonoporation can be visualized and induced in vivo using targeted microbubbles.

  • Sonoporation of endothelial cells by vibrating targeted microbubbles
    Journal of Controlled Release, 2011
    Co-Authors: Klazina Kooiman, N. De Jong, Antonius F W Van Der Steen, Miranda Foppenharteveld
    Abstract:

    Molecular imaging using ultrasound makes use of targeted microbubbles. In this study we investigated whether these microbubbles could also be used to induce Sonoporation in endothelial cells. Lipid-coated microbubbles were targeted to CD31 and insonified at 1 MHz at low peak negative acoustic pressures at six sequences of 10 cycle sine-wave bursts. Vibration of the targeted microbubbles was recorded with the Brandaris-128 high-speed camera (~ 13 million frames per second). In total, 31 cells were studied that all had one microbubble (1.2–4.2 micron in diameter) attached per cell. After insonification at 80 kPa, 30% of the cells (n = 6) had taken up propidium iodide, while this was 20% (n = 1) at 120 kPa and 83% (n = 5) at 200 kPa. Irrespective of the peak negative acoustic pressure, uptake of propidium iodide was observed when the relative vibration amplitude of targeted microbubbles was greater than 0.5. No relationship was found between the position of the microbubble on the cell and induction of Sonoporation. This study shows that targeted microbubbles can also be used to induce Sonoporation, thus making it possible to combine molecular imaging and drug delivery

Jennifer Manfan Wan - One of the best experts on this subject based on the ideXlab platform.

  • membrane blebbing as a recovery manoeuvre in site specific Sonoporation mediated by targeted microbubbles
    Journal of the Royal Society Interface, 2015
    Co-Authors: Ruen Shan Leow, Jennifer Manfan Wan
    Abstract:

    Site-specific perforation of the plasma membrane can be achieved through ultrasound-triggered cavitation of a single microbubble positioned adjacent to the cell. However, for this perforation approach (Sonoporation), the recovery manoeuvres invoked by the cell are unknown. Here, we report new findings on how membrane blebbing can be a recovery manoeuvre that may take place in Sonoporation episodes whose pores are of micrometres in diameter. Each Sonoporation site was created using a protocol involving single-shot ultrasound exposure (frequency: 1 MHz; pulse length: 30 cycles; peak negative pressure: 0.45 MPa) which triggered inertial cavitation of a single targeted microbubble (diameter: 1–5 µm). Over this process, live confocal microscopy was conducted in situ to monitor membrane dynamics, model drug uptake kinetics and cytoplasmic calcium ion (Ca2+) distribution. Results show that blebbing would occur at a recovering Sonoporation site after its resealing, and it may emerge elsewhere along the membrane periphery. The bleb size was correlated with the pre-exposure microbubble diameter, and 99% of blebbing cases at Sonoporation sites were inflicted by microbubbles larger than 1.5 µm diameter (analysed over 124 Sonoporation episodes). Blebs were not observed at irreversible Sonoporation sites or when Sonoporation site repair was inhibited via extracellular Ca2+ chelation. Functionally, the bleb volume was found to serve as a buffer compartment to accommodate the cytoplasmic Ca2+ excess brought about by Ca2+ influx during Sonoporation. These findings suggest that membrane blebbing would help sonoporated cells restore homeostasis.

  • single site Sonoporation disrupts actin cytoskeleton organization
    Journal of the Royal Society Interface, 2014
    Co-Authors: Xian Chen, Ruen Shan Leow, Jennifer Manfan Wan
    Abstract:

    Sonoporation is based upon an ultrasound–microbubble cavitation routine that physically punctures the plasma membrane on a transient basis. During such a process, the actin cytoskeleton may be disrupted in tandem because this network of subcellular filaments is physically interconnected with the plasma membrane. Here, by performing confocal fluorescence imaging of single-site Sonoporation episodes induced by ultrasound-triggered collapse of a single targeted microbubble, we directly observed immediate rupturing of filamentary actin (F-actin) at the Sonoporation site (cell type: ZR-75-30; ultrasound frequency: 1 MHz; peak negative pressure: 0.45 MPa; pulse duration: 30 cycles; bubble diameter: 2–4 µm). Also, through conducting a structure tensor analysis, we observed further disassembly of the F-actin network over the next 60 min after the onset of Sonoporation. The extent of F-actin disruption was found to be more substantial in cells with higher uptake of Sonoporation tracer. Commensurate with this process, cytoplasmic accumulation of globular actin (G-actin) was evident in sonoporated cells, and in turn the G-actin : F-actin ratio was increased in a trend similar to drug-induced (cytochalasin D) actin depolymerization. These results demonstrate that Sonoporation is not solely a membrane-level phenomenon: organization of the actin cytoskeleton is concomitantly perturbed.

  • membrane perforation and recovery dynamics in microbubble mediated Sonoporation
    Ultrasound in Medicine and Biology, 2013
    Co-Authors: Jennifer Manfan Wan
    Abstract:

    Transient Sonoporation can essentially be epitomized by two fundamental processes: acoustically induced membrane perforation and its subsequent resealing. To provide insight into these processes, this article presents a new series of direct evidence on the membrane-level dynamics during and after an episode of Sonoporation. Our direct observations were obtained from anchored fetal fibroblasts whose membrane topography was imaged in situ using real-time confocal microscopy. To facilitate controlled Sonoporation at the single-cell level, microbubbles that can passively adhere to the cell membrane were first introduced at a 1:1 cell-to-bubble ratio. Single-pulse ultrasound exposure (1-MHz frequency, 10-cycle pulse duration, 0.85-MPa peak negative pressure in situ) was then applied to trigger microbubble pulsation/collapse, which, in turn, instigated membrane perforation. With this protocol, five membrane-level phenomena were observed: (i) localized perforation of the cell membrane was synchronized with the instant of ultrasound pulsing; (ii) perforation sites with temporal peak area 100 μm(2)) or in the absence of extracellular calcium ions. These findings serve to underscore the spatiotemporal complexity of membrane-level dynamics in Sonoporation.

  • induction of endoplasmic reticulum stress by Sonoporation linkage to mitochondria mediated apoptosis initiation
    Ultrasound in Medicine and Biology, 2013
    Co-Authors: W Zhong, Jennifer Manfan Wan, Xian Chen, Pingping Jiang, Peng Qin
    Abstract:

    Abstract The use of cavitational means to create transient membrane pores on living cells ( i.e., Sonoporation) may potentially induce a broad range of downstream bio-effects that disrupt the functioning of various organelles. Here we observed that on HL-60 leukemia cells, Sonoporation may induce endoplasmic reticulum (ER) stress on a time-lapse basis and, in turn, signal the mitochondria to commit a cell toward apoptosis. Our observations were derived from in vitro ultrasound exposure experiments performed on HL-60 cells in the presence of lipid-shelled microbubbles (1:1 cell-to-bubble ratio; 1-MHz frequency; 0.45-MPa in situ peak negative pressure; 100-cycle pulse length; 1-kHz pulse repetition frequency; 60-s exposure period). Using flow cytometry, we found that sonoporated cells exhibited a progressive loss of functional ER mass over a 6-h period. Also, post-exposure Western blot assays (between 0 and 24 h) revealed various indications of post-Sonoporation ER stress: (i) upregulation of ER-resident enzymes responsible for catalyzing protein folding; (ii) activation of trans-ER-membrane stress sensors; (iii) increased expression of ER-induced regulatory proteins that mediate pro-apoptotic signals to the mitochondria. These results corresponded to flow cytometry observations that depicted a progressive depolarization of a sonoporated cell’s mitochondrial outer membrane potential. They were also consistent with another Western blot assay that found, in sonoporated cells, a time-lapse increase of caspase-9 (a mitochondria-activated apoptosis initiator protein). Taken together, our findings indicate that Sonoporation may upset ER homeostasis, and this may ultimately result in initiation of apoptosis.

  • Sonoporation as a cellular stress induction of morphological repression and developmental delays
    Ultrasound in Medicine and Biology, 2013
    Co-Authors: Xian Chen, Jennifer Manfan Wan
    Abstract:

    For Sonoporation to be established as a drug/gene delivery paradigm, it is essential to account for the biological impact of this membrane permeation strategy on living cells. Here we provide new insight into the cellular impact of Sonoporation by demonstrating in vitro that this way of permeating the plasma membrane may inadvertently induce repressive cellular features even while enhancing exogenous molecule uptake. Both suspension-type (HL-60) and monolayer (ZR-75-30) cells were considered in this investigation, and they were routinely exposed to 1-MHz pulsed ultrasound (pulse length, 100 cycles; pulse repetition frequency, 1 kHz; exposure period, 60 s) with calibrated field profile (spatial-averaged peak negative pressure, 0.45 MPa) and in the presence of microbubbles (cell:bubble ratio, 10:1). The post-exposure morphology of sonoporated cells (identified as those with calcein internalization) was examined using confocal microscopy, and their cell cycle progression kinetics were analyzed using flow cytometry. Results show that for both cell types investigated, sonoporated cells exhibited membrane shrinkage and intra-cellular lipid accumulation over a 2-h period. Also, as compared with normal cells, the deoxyribonucleic acid synthesis duration of sonoporated cells was significantly lengthened, indicative of a delay in cell cycle progression. These features are known to be characteristics of a cellular stress response, suggesting that Sonoporation indeed constitutes as a stress to living cells. This issue may need to be addressed in optimizing Sonoporation for drug/gene delivery purposes. On the other hand, it raises opportunities for developing other therapeutic applications via Sonoporation.