Electro‐mediated gene transfer and expression are controlled by the life‐time of DNA/membrane complex formation

Faurie, CÃ©cile; Reberšek, Matej; Golzio, Muriel; Kandušer, Maša; Escoffre, Jean‐Michel; Pavlin, Mojca; Miklavčič, Damijan; Rols, Marie‐Pierre

doi:10.1002/jgm.1414

Cited by 105 publications

(97 citation statements)

References 48 publications

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“…Pulse parameters were selected by taking into account previous data (Faurie et al 2010). This reference was our guideline for the choice in the delay between successive pulses in a train for the pulse polarity inversion.…”

Section: Resultsmentioning

confidence: 99%

Electric Field Orientation for Gene Delivery Using High-Voltage and Low-Voltage Pulses

et al. 2012

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Electropermeabilization is a biological physical process in response to the presence of an applied electric field that is used for the transfer of hydrophilic molecules such as anticancer drugs or DNA across the plasma membranes of living cells. The molecular processes that support the transfer are poorly known. The aim of our study was to investigate the effect of high-voltage and low-voltage (HVLV) pulses in vitro with different orientations on cell permeabilization, viability and gene transfection. We monitored the permeabilization with unipolar and bipolar HVLV pulses with different train repetition pulses, showing that HVLV pulses increase cell permeabilization and cell viability. Gene transfer was also observed by measuring green fluorescent protein (GFP) expression. The expression was the same for HVLV pulses and electrogenotherapy pulses for in vitro experimentation. As the viability was better preserved for HVLV-pulsed cells, we managed to increase the number of GFP-expressing cells by up to 65% under this condition. The use of bipolar HVLV train pulses increased gene expression to a higher extent, probably by affecting a larger part of the cell surface.

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Section: Resultsmentioning

confidence: 99%

Electric Field Orientation for Gene Delivery Using High-Voltage and Low-Voltage Pulses

et al. 2012

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“…Prior studies have shown that the combination of a short-duration, high-field-strength first pulse, together with a longer-duration, lowerfield-strength second pulse (denoted by 'first pulse' and 'second pulse', respectively) can increase electroporation efficiency and preserve cell viability, especially when delivering larger molecules (MW N 4 kDa [47]) or DNA [33,34,37,39,42,[57][58][59][60][61][62][63][64][65][66]. Pulsing parameters have been well studied, including the strength of the first pulse [33,37,58], the strength and duration of the second pulse [34,39,[57][58][59], and the delay between pulses [34,42,57,58,66,67].…”

Section: Introductionmentioning

confidence: 99%

Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery

Demiryurek

Nickaeen

Zheng

et al. 2015

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Electroporation is of interest for many drug-delivery and gene-therapy applications. Prior studies have shown that a two-pulse-electroporation protocol consisting of a short-duration, high-voltage first pulse followed by a longer, low-voltage second pulse can increase delivery efficiency and preserve viability. In this work the effects of the field strength of the first and second pulses and the inter-pulse delay time on the delivery of two different-sized Fluorescein-Dextran (FD) conjugates are investigated. A series of two-pulse-electroporation experiments were performed on 3T3-mouse fibroblast cells, with an alternating-current first pulse to permeabilize the cell, followed by a direct-current second pulse. The protocols were rationally designed to best separate the mechanisms of permeabilization and electrophoretic transport. The results showed that the delivery of FD varied strongly with the strength of the first pulse and the size of the target molecule. The delivered FD concentration also decreased linearly with the logarithm of the inter-pulse delay. The data indicate that membrane resealing after electropermeabilization occurs rapidly, but that a non-negligible fraction of the pores can be reopened by the second pulse for delay times on the order of hundreds of seconds. The role of the second pulse is hypothesized to be more than just electrophoresis, with a minimum threshold field strength required to reopen nano-sized pores or defects remaining from the first pulse. These results suggest that membrane electroporation, sealing, and re-poration is a complex process that has both short-term and long-term components, which may in part explain the wide variation in membrane-resealing times reported in the literature.

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“…Translocation of the plasmid from the plasma membrane to the cytoplasm and its subsequent passage towards the nuclear envelope take place with a kinetics ranging from minutes to hours (25). When plasmid has reached the nuclei, gene expression can take place and this can be detected up to several weeks later.…”

Section: A Basic Aspects: Modulation Of Membrane Potential Differencementioning

confidence: 99%

How Imaging Molecule Uptake into Cells can Reveal the Mechanisms of Membrane Electropermeabilization

Rols

2016

IFMBE Proceedings

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Abstract-Cell membranes can be transiently permeabilized under application of electric field pulses. This process, called electropermeabilization or electroporation, allows hydrophilic molecules, such as anticancer drugs and DNA, to enter into cells and tissues. Single cell imaging experiments revealed that the uptake of molecules is a complex multi-step that takes place in well-defined membrane regions under processes that affect the entire membrane, depend on the properties of the molecules (size, charge) and in which the cytoskeleton can play a major role.

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Electro‐mediated gene transfer and expression are controlled by the life‐time of DNA/membrane complex formation

Cited by 105 publications

References 48 publications

Electric Field Orientation for Gene Delivery Using High-Voltage and Low-Voltage Pulses

Electric Field Orientation for Gene Delivery Using High-Voltage and Low-Voltage Pulses

Transport, resealing, and re-poration dynamics of two-pulse electroporation-mediated molecular delivery

How Imaging Molecule Uptake into Cells can Reveal the Mechanisms of Membrane Electropermeabilization

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