Lipid membrane instability and poration driven by capacitive charging

A new model for the interaction of an electric pulse with a lipid membrane is proposed. Using this model we show that when a DC electric pulse is applied to an insulating lipid membrane separating fluids with different conductivities, the capacitive charging current through the membrane drives electrohydrodynamic flow that destabilizes the membrane. The instability is transient and decays as the membrane charges. The bulk conductivity mismatch plays an essential role in this instability because it results in a different rate of charge accumulation on the membrane's physical surfaces. Shearing stresses created by the electric field acting on its own induced free charge are non-zero as long as the charge imbalance exists. Accordingly, the most unstable mode is related to the ratio of membrane charging time and the electrohydrodynamic time.Comment: 4 pages, 4 figure

Post-pulse addition of trans-cyclohexane-1,2-diol improves electrotransfer mediated gene expression in mammalian cells

AbstractElectric field mediated gene transfer is facing a problem in expression yield due to the poor transfer across the nuclear envelope. Trans-cyclohexane-1,2-diol (TCHD) was shown to significantly increase chemically mediated transfection by collapsing the permeability barrier of the nuclear pore complex. We indeed observed a significant increase in expression by electrotransfer when cells are treated post pulse by a low non toxic concentration of TCHD. This was obtained for different pulsing conditions, cell strains and plasmid constructs. An interesting improvement in cell viability can be obtained. This can significantly enhance the non-viral gene electrical delivery

Control by pulse parameters of electric field-mediated gene transfer in mammalian cells

Wolf H, Rols MP, Boldt E, Neumann E, Teissie J. Control by pulse parameters of electric field-mediated gene transfer in mammalian cells. Biophysical Journal. 1994;66(2):524-531.Electric field-mediated gene transfer in mammalian cells (electrotransformation) depends on the pulsing conditions (field intensity, pulse duration, number of pulses). The effect of these parameters was systematically investigated using the transient expression of the chloramphenicol acetyltransferase and the beta-galactosidase activities in Chinese hamster ovary cells. Pulsing conditions inducing reversible permeabilization of the cell plasma membrane are not sufficient to induce gene transfer. The plasmid must be present during the electric pulse if it is to be transferred across the membrane into the cytoplasm. Only the localized part of the cell membrane brought to the permeabilized state by the external field is competent. Pulse duration plays a key role in the magnitude of the transfer. The field induces a complex reaction between the membrane and the plasmid that is accumulated at the cell interface by electrophoretic forces. This leads to an insertion of the plasmid, which can then cross the membrane

The prolate-to-oblate shape transition of phospholipid vesicles in response to frequency variation of an AC electric field can be explained by the dielectric anisotropy of a phospholipid bilayer

The external electric field deforms flaccid phospholipid vesicles into spheroidal bodies, with the rotational axis aligned with its direction. Deformation is frequency dependent: in the low frequency range (~ 1 kHz), the deformation is typically prolate, while increasing the frequency to the 10 kHz range changes the deformation to oblate. We attempt to explain this behaviour with a theoretical model, based on the minimization of the total free energy of the vesicle. The energy terms taken into account include the membrane bending energy and the energy of the electric field. The latter is calculated from the electric field via the Maxwell stress tensor, where the membrane is modelled as anisotropic lossy dielectric. Vesicle deformation in response to varying frequency is calculated numerically. Using a series expansion, we also derive a simplified expression for the deformation, which retains the frequency dependence of the exact expression and may provide a better substitute for the series expansion used by Winterhalter and Helfrich, which was found to be valid only in the limit of low frequencies. The model with the anisotropic membrane permittivity imposes two constraints on the values of material constants: tangential component of dielectric permittivity tensor of the phospholipid membrane must exceed its radial component by approximately a factor of 3; and the membrane conductivity has to be relatively high, approximately one tenth of the conductivity of the external aqueous medium.Comment: 17 pages, 6 figures; accepted for publication in J. Phys.: Condens. Matte

Synchronization modulation increases transepithelial potentials in MDCK monolayers through Na/K pumps

Peer reviewedPublisher PD

Perforin Rapidly Induces Plasma Membrane Phospholipid Flip-Flop

The cytotoxic cell granule secretory pathway is essential for host defense. This pathway is fundamentally a form of intracellular protein delivery where granule proteases (granzymes) from cytotoxic lymphocytes are thought to diffuse through barrel stave pores generated in the plasma membrane of the target cell by the pore forming protein perforin (PFN) and mediate apoptotic as well as additional biological effects. While recent electron microscopy and structural analyses indicate that recombinant PFN oligomerizes to form pores containing 20 monomers (20 nm) when applied to liposomal membranes, these pores are not observed by propidium iodide uptake in target cells. Instead, concentrations of human PFN that encourage granzyme-mediated apoptosis are associated with pore structures that unexpectedly favor phosphatidylserine flip-flop measured by Annexin-V and Lactadherin. Efforts that reduce PFN mediated Ca influx in targets did not reduce Annexin-V reactivity. Antigen specific mouse CD8 cells initiate a similar rapid flip-flop in target cells. A lipid that augments plasma membrane curvature as well as cholesterol depletion in target cells enhance flip-flop. Annexin-V staining highly correlated with apoptosis after Granzyme B (GzmB) treatment. We propose the structures that PFN oligomers form in the membrane bilayer may include arcs previously observed by electron microscopy and that these unusual structures represent an incomplete mixture of plasma membrane lipid and PFN oligomers that may act as a flexible gateway for GzmB to translocate across the bilayer to the cytosolic leaflet of target cells

Electroporation-Induced Electrosensitization

BACKGROUND: Electroporation is a method of disrupting the integrity of cell membrane by electric pulses (EPs). Electrical modeling is widely employed to explain and study electroporation, but even most advanced models show limited predictive power. No studies have accounted for the biological consequences of electroporation as a factor that alters the cell's susceptibility to forthcoming EPs. METHODOLOGY/PRINCIPAL FINDINGS: We focused first on the role of EP rate for membrane permeabilization and lethal effects in mammalian cells. The rate was varied from 0.001 to 2,000 Hz while keeping other parameters constant (2 to 3,750 pulses of 60-ns to 9-µs duration, 1.8 to 13.3 kV/cm). The efficiency of all EP treatments was minimal at high rates and started to increase gradually when the rate decreased below a certain value. Although this value ranged widely (0.1-500 Hz), it always corresponded to the overall treatment duration near 10 s. We further found that longer exposures were more efficient irrespective of the EP rate, and that splitting a high-rate EP train in two fractions with 1-5 min delay enhanced the effects severalfold. CONCLUSIONS/SIGNIFICANCE: For varied experimental conditions, EPs triggered a delayed and gradual sensitization to EPs. When a portion of a multi-pulse exposure was delivered to already sensitized cells, the overall effect markedly increased. Because of the sensitization, the lethality in EP-treated cells could be increased from 0 to 90% simply by increasing the exposure duration, or the exposure dose could be reduced twofold without reducing the effect. Many applications of electroporation can benefit from accounting for sensitization, by organizing the exposure either to maximize sensitization (e.g., for sterilization) or, for other applications, to completely or partially avoid it. In particular, harmful side effects of electroporation-based therapies (electrochemotherapy, gene therapies, tumor ablation) include convulsions, pain, heart fibrillation, and thermal damage. Sensitization can potentially be employed to reduce these side effects while preserving or increasing therapeutic efficiency

Internet of Things in Agricultural Innovation and Security

The agricultural Internet of Things (Ag-IoT) paradigm has tremendous potential in transparent integration of underground soil sensing, farm machinery, and sensor-guided irrigation systems with the complex social network of growers, agronomists, crop consultants, and advisors. The aim of the IoT in agricultural innovation and security chapter is to present agricultural IoT research and paradigm to promote sustainable production of safe, healthy, and profitable crop and animal agricultural products. This chapter covers the IoT platform to test optimized management strategies, engage farmer and industry groups, and investigate new and traditional technology drivers that will enhance resilience of the farmers to the socio-environmental changes. A review of state-of-the-art communication architectures and underlying sensing technologies and communication mechanisms is presented with coverage of recent advances in the theory and applications of wireless underground communications. Major challenges in Ag-IoT design and implementation are also discussed

Nanoelectropulse-driven membrane perturbation and small molecule permeabilization

BACKGROUND: Nanosecond, megavolt-per-meter pulsed electric fields scramble membrane phospholipids, release intracellular calcium, and induce apoptosis. Flow cytometric and fluorescence microscopy evidence has associated phospholipid rearrangement directly with nanoelectropulse exposure and supports the hypothesis that the potential that develops across the lipid bilayer during an electric pulse drives phosphatidylserine (PS) externalization. RESULTS: In this work we extend observations of cells exposed to electric pulses with 30 ns and 7 ns durations to still narrower pulse widths, and we find that even 3 ns pulses are sufficient to produce responses similar to those reported previously. We show here that in contrast to unipolar pulses, which perturb membrane phospholipid order, tracked with FM1-43 fluorescence, only at the anode side of the cell, bipolar pulses redistribute phospholipids at both the anode and cathode poles, consistent with migration of the anionic PS head group in the transmembrane field. In addition, we demonstrate that, as predicted by the membrane charging hypothesis, a train of shorter pulses requires higher fields to produce phospholipid scrambling comparable to that produced by a time-equivalent train of longer pulses (for a given applied field, 30, 4 ns pulses produce a weaker response than 4, 30 ns pulses). Finally, we show that influx of YO-PRO-1, a fluorescent dye used to detect early apoptosis and activation of the purinergic P2X(7 )receptor channels, is observed after exposure of Jurkat T lymphoblasts to sufficiently large numbers of pulses, suggesting that membrane poration occurs even with nanosecond pulses when the electric field is high enough. Propidium iodide entry, a traditional indicator of electroporation, occurs with even higher pulse counts. CONCLUSION: Megavolt-per-meter electric pulses as short as 3 ns alter the structure of the plasma membrane and permeabilize the cell to small molecules. The dose responses of cells to unipolar and bipolar pulses ranging from 3 ns to 30 ns duration support the hypothesis that a field-driven charging of the membrane dielectric causes the formation of pores on a nanosecond time scale, and that the anionic phospholipid PS migrates electrophoretically along the wall of these pores to the external face of the membrane