The role of gel-phase domains in electroporation of vesicles

Abstract

Transient permeabilisation of the cell membrane is a critical step to introduce drugs or DNA into living cells, yet challenging for both biological research and therapeutic applications. To achieve this, electroporation (or electropermeabilisation) has become a widely used method due to its simplicity to deliver almost any biomolecule to any cell type. Although this method demonstrates promise in the field of drug/gene delivery, the underlying physical mechanisms of the response of the heterogeneous cell membrane to strong electric pulses is still unknown. In this study, we have investigated the role of gel-phase lipids in the electroporation of binary giant unilamellar vesicles (GUVs), composed from DPPC (gel-phase) and DPhPC (fluid-phase) lipids (molar ratio 8:2 and 2:8). We have observed that the exposure to electric pulses leads to expel of fluid-phase lipids and concomitant decrease in GUV size, whereas the gel-phase domains become buckled. Based on experiments on pure fluid-phase and gel-phase GUVs, we have found that fluid-phase lipids can be expelled by electrical forces and the highly viscous gel-phase lipids cannot. Moreover, our analyses suggest that pore formation occurs primarily in fluid-phase domains and that the pore size is similar in all GUVs containing fluid-phase lipids, irrespective of the gel-phase percentage.

Self‐Assembly of Giant Unilamellar Vesicles by Film Hydration Methodologies

“…Besides the setup, the conditions can vary between different studies without any obvious reasons ( Table 1 ). In terms of frequency, regardless of ITO/Pt electrodes or lipid/polymer amphiphiles, most studies uses 10 Hz, which is sometimes followed by a decrease of the frequency to 4–5 Hz, said to be required for GUV detachment . Some experiments also use frequencies as low as 1 Hz or as high as 500 Hz for lipids .…”

“…Dimova et al was able to prove that lowering the voltage to 800 mV also generated polymersome GUVs but required longer times to form, 3 h, contrary to 15 min at 9 V . Sometimes the change in frequency for detaching the GUVs from the electrode is associated with a change in voltage to higher or lower values . In terms of duration, most electroformation methodologies require only a few minutes, several hours or are left overnight (≈12 h) .…”

“…The formation of vesicles by electroformation is always carried out above the transition temperature ( T m ) of the lipids, as liposomes seem to only self‐assemble in the lipid‐disordered state . For example, 1,2‐dipalmitoyl‐sn‐glycero‐3‐phosphocholine (DPPC) has a T m of 41 °C, therefore electroformation is always carried out at ≥50 °C . Using high temperatures can denature the proteins and impair their activity and can become problematic when reconstituting proteins in GUVs.…”

“…In terms of lipids, zwitterionic (PC, PE), negatively charged (PS, phosphatidylglycerol (PG)) and positively charged (DOTAP) lipids have all been successfully used. These lipids can be used alone or in complex mixture more relevant to cell membranes notably when also incorporating cholesterol . For polymers, diblock and triblock amphiphilic polymers have successfully formed GUVs by electroformation using a variety of hydrophobic blocks (PEE, PDMS, PB, polyethylene (PEt)) and a few hydrophilic blocks (PEO or poly‐2‐methyl‐2‐oxazoline (PMOXA)) .…”

Unraveling the response of a biomimetic actin cortex to electric pulses in vesicles

“…the actin-supported bilayer has a higher electrical stability ( Figure 5B). This could be caused by the increased surface viscosity of the bilayer due to the presence of the actin layer 37 .…”