Nanosecond pulsed electric field effects on human cells

Electric fields have been used on intact cells for decades, most notably for effects on plasma membranes for cell fusion and transient electroporation to allow the entry of xenomolecules. Compared to electroporation pulses, new pulse power technology generates non-thermal pulses with shorter durations and higher electric fields (nsPEF) such that the charging time of the plasma membrane is not reached, but subcellular membranes and intracellular functions are affected independent of heating. Here we investigated nsPEF-induced programmed cell death (apoptosis) in intact human cells with pulse durations between 10-300 nanoseconds over a range of electric field intensities and energy densities. nsPEF effects are observed on the plasma membrane integrity and caspase activation using fluorescent markers, flow cytometry, biochemical assays. The results indicate that once a critical electric field intensity threshold is reached, nsPEF-induced apoptosis is dependent on the pulse duration and is independent of the electric field intensity. Relatively long pulses (300 ns), which have greater plasma membrane effects and lesser intracellular effects, recruit apoptosis mechanisms that rapidly advance through apoptosis to loss of membrane integrity in minutes. Shorter pulses (10 ns), which have greater intracellular effects and lesser plasma membrane effects, induce apoptosis, but require longer times (tens of minutes to hours). We conclude that nsPEF-induced apoptosis does not require plasma membrane effects related to electroporation. nsPEF affect a continuum of cellular mechanisms from the plasma membrane to intracellular targets. Longer pulses recruit multiple and perhaps synergistic pathways while shorter pulses recruit more time-dependent pathways. These studies promote a better understand of the effects of nsPEF on basic functions during cellular responses to intrinsic and extrinsic stresses and suggest the potential for nsPEF applications for therapeutic purposes.