Ultrasound-mediated disruption of cell membranes. I. Quantification of molecular uptake and cell viability

Guzmán, Héctor R; Nguyen, Daniel X.; Khan, Sohail; Prausnitz, Mark R.

doi:10.1121/1.1376131

Cited by 186 publications

(151 citation statements)

References 24 publications

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“…Increased intracellular delivery is believed to occur due to the formation of transient pores of nanometer to micron dimensions [25,43,44]. Consistent with this expectation, our flow cytometry results show that ultrasound increased intracellular DNA delivery for both naked DNA and PEI:DNA complexes (Fig.…”

Section: Discussionsupporting

confidence: 87%

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Synergistic effect of ultrasound and PEI on DNA transfection in vitro

Deshpande

Prausnitz

2007

Journal of Controlled Release

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Ultrasound and poly(ethylenimine) (PEI) have each separately been shown to increase DNA transfection efficiency. This study tested the hypothesis that the combination of ultrasound and PEI can have a synergistic effect to increase DNA transfection. This in vitro study assessed transfection efficiency of two different DNA plasmids encoding green fluorescent protein and firefly luciferase in two different cells types, a primary culture of human aortic smooth muscle cells and an immortal line of human prostrate cancer cells. We found that ultrasound sonication increased transfection up to 18-fold, DNA complexation with PEI increased transfection up to 90-fold, and the combination of ultrasound and PEI synergistically increased transfection up to 200-fold, which resulted in reporter gene expression by 34% of cells. Kinetic measurements found that the effects of ultrasound alone acted quickly, whereas increased transfection by PEI either alone or in combination with ultrasound strongly benefited from a 4-h incubation with the DNA plasmid after sonication. Although serum reduced absolute expression levels, it did not affect the relative increase in transfection when ultrasound was added to PEI enhancement. Flow cytometry measurements showed that sonication increased intracellular uptake of labelled DNA complexed to PEI by 55% relative to PEI complexation alone. Electrophoresis assay showed no damage to DNA or PEI-DNA complexes after sonication. Overall, these results suggest that the combination of ultrasound and PEI can have a synergistic effect to increase DNA transfection.

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Section: Discussionsupporting

confidence: 87%

“…Ultrasound equipment-As previously described in detail [25], the ultrasound apparatus consisted of a focused, 500-kHz, piezoelectric transducer with an 8.9-cm focal length and a beam width of 3 mm (−6 db intensity area) at the focal beam point (Fig. 1).…”

Section: 13mentioning

confidence: 99%

Synergistic effect of ultrasound and PEI on DNA transfection in vitro

Deshpande

Prausnitz

2007

Journal of Controlled Release

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“…By voltage clamp techniques, an increase in transmembrane current simultaneously with ultrasound application was reported, indicating ion flux through pores [10]. Moreover, several papers demonstrated the uptake of cell-impermeable molecules when exposing cells to ultrasound [11][12][13]. However, more recently, Meijering et al [14] stated that besides pore formation, enhanced endocytosis also contributes to ultrasound mediated delivery.…”

Section: Introductionmentioning

confidence: 99%

Ultrasound and microbubble mediated drug delivery: Acoustic pressure as determinant for uptake via membrane pores or endocytosis

Cock

Zagato

Braeckmans

et al. 2015

Journal of Controlled Release

208

229

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Although promising results are achieved in ultrasound mediated drug delivery, its underlying biophysical mechanisms remain to be elucidated. Pore formation as well as endocytosis has been reported during ultrasound application. Due to the plethora of ultrasound settings used in literature, it is extremely difficult to draw conclusions on which mechanism is actually involved. To our knowledge, we are the first to show that acoustic pressure influences which route of drug uptake is addressed, by inducing different microbubble-cell interactions. To investigate this, FITC-dextrans were used as model drugs and their uptake was analyzed by flow cytometry. In fluorescence intensity plots, two subpopulations arose in cells with FITCdextran uptake after ultrasound application, corresponding to cells having either low or high uptake. Following separation of the subpopulations by FACS sorting, confocal images indicated that the low uptake population showed endocytic uptake. The high uptake population represented uptake via pores. Moreover, the distribution of the subpopulations shifted to the high uptake population with increasing acoustic pressure. Real-time confocal recordings during ultrasound revealed that membrane deformation by microbubbles may be the trigger for endocytosis via mechanostimulation of the cytoskeleton. Pore formation was shown to be caused by microbubbles propelled towards the cell. These results provide a better insight in the role of acoustic pressure in microbubble-cell interactions and the possible consequences for drug uptake. In addition, it pinpoints the need of a more rational, microbubble behavior based choice of acoustic parameters in ultrasound mediated drug delivery experiments.

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“…To cite a few of them, there is clear evidence that the permeability of the cell membrane to large molecules (both drugs and genes) is increased when suspensions of cells are exposed to ultrasound in the presence of an UCA (Bao et al 1997;Brayman et al 1999;Greenleaf et al 1998;Guzmán et al 2001a;Guzmán et al 2001b;Kinoshita and Hynynen 2005;Ward et al 2000). Inertial cavitation (IC) is believed to be the key mechanism for such effect.…”

Section: Introductionmentioning

confidence: 99%

Cavitation Threshold of Microbubbles in Gel Tunnels by Focused Ultrasound

Sassaroli

Hynynen

2007

Ultrasound in Medicine & Biology

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The investigation of inertial cavitation in micro-tunnels has significant implications for the development of therapeutic applications of ultrasound such as ultrasound-mediated drug and gene delivery. The threshold for inertial cavitation was investigated using a passive cavitation detector with a center frequency of 1 MHz. Micro-tunnels of various diameters (90 to 800 μm) embedded in gel were fabricated and injected with a solution of Optison™ contrast agent of concentrations 1.2% and 0.2% diluted in water. An ultrasound pulse of duration 500 ms and center frequency 1.736 MHz was used to insonate the microbubbles. The acoustic pressure was increased at one second intervals until broadband noise emission was detected. The pressure threshold at which broadband noise emission was observed was found to be dependent on the diameter of the micro-tunnels, with an average increase of 1.2 to 1.5 between the smallest and the largest tunnels, depending on the microbubble concentration. The evaluation of inertial cavitation in gel tunnels rather than tubes provides a novel opportunity to investigate microbubble collapse in a situation that simulates in vivo blood vessels better than tubes with solid walls do.

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Ultrasound-mediated disruption of cell membranes. I. Quantification of molecular uptake and cell viability

Cited by 186 publications

References 24 publications

Synergistic effect of ultrasound and PEI on DNA transfection in vitro

Synergistic effect of ultrasound and PEI on DNA transfection in vitro

Ultrasound and microbubble mediated drug delivery: Acoustic pressure as determinant for uptake via membrane pores or endocytosis

Cavitation Threshold of Microbubbles in Gel Tunnels by Focused Ultrasound

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