Nanosecond, high‐intensity pulsed electric fields induce apoptosis in human cells

Beebe, Stephen J.; Fox, P.; Rec, Laura J.; Willis, E Lauren K; Schoenbach, K.H.

doi:10.1096/fj.02-0859fje

Cited by 418 publications

(355 citation statements)

References 38 publications

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“…The electric fields administered were 4, 6.5, 10, and 15 kV/cm. The generator consisted of a 10-ohm pulse-forming network (five 50-ohm cables in parallel) and a spark gap in atmospheric air as a nanosecond closing switch (16). Post-pulse, the cell suspension was removed from the nsPEF cuvette and assayed.…”

Section: Methodsmentioning

confidence: 99%

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Stimulation of Capacitative Calcium Entry in HL-60 Cells by Nanosecond Pulsed Electric Fields

White

Blackmore

Schoenbach

et al. 2004

Journal of Biological Chemistry

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Nanosecond pulsed electric fields (nsPEFs) are hypothesized to affect intracellular structures in living cells providing a new means to modulate cell signal transduction mechanisms. The effects of nsPEFs on the release of internal calcium and activation of calcium influx in HL-60 cells were investigated by using real time fluorescent microscopy with Fluo-3 and fluorometry with Fura-2. nsPEFs induced an increase in intracellular calcium levels that was seen in all cells. With pulses of 60 ns duration and electric fields between 4 and 15 kV/cm, intracellular calcium increased 200 -700 nM, respectively, above basal levels (ϳ100 nM), while the uptake of propidium iodide was absent. This suggests that increases in intracellular calcium were not because of plasma membrane electroporation. nsPEF and the purinergic agonist UTP induced calcium mobilization in the presence and absence of extracellular calcium with similar kinetics and appeared to target the same inositol 1,4,5-trisphosphate-and thapsigargin-sensitive calcium pools in the endoplasmic reticulum. For cells exposed to either nsPEF or UTP in the absence of extracellular calcium, there was an electric field-dependent or UTP dose-dependent increase in capacitative calcium entry when calcium was added to the extracellular media. These findings suggest that nsPEFs, like ligand-mediated responses, release calcium from similar internal calcium pools and thus activate plasma membrane calcium influx channels or capacitative calcium entry.

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Section: Methodsmentioning

confidence: 99%

“…Previous studies showed that nsPEF treatment increased apoptosis signaling mechanisms in both Jurkat and HL-60 cells through assays for caspase activation, annexin binding, and cytochrome c localization (16). Other studies using electric fields weaker than those that induce apoptosis have been shown to cause a proliferative effect on cells.…”

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confidence: 99%

Stimulation of Capacitative Calcium Entry in HL-60 Cells by Nanosecond Pulsed Electric Fields

White

Blackmore

Schoenbach

et al. 2004

Journal of Biological Chemistry

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225

157

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“…They provide a new approach for physically targeting intracellular organelles with many applications, including the initiation of apoptosis in cultured cells [12][13][14] and tumors [6], enhancement of gene transfection efficiency [13,14], and inhibiting tumor growth [6]. During the past year, we have treated over 300 murine melanomas in 120 mice with 40 kV/cm electric field pulses 300 ns in duration with dramatic results.…”

Section: Introductionmentioning

confidence: 99%

Nanosecond pulsed electric fields cause melanomas to self-destruct

Nuccitelli

Pliquett

Chen

et al. 2006

Biochemical and Biophysical Research Communications

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We have discovered a new, drug-free therapy for treating solid skin tumors. Pulsed electric fields greater than 20 kV/cm with rise times of 30 ns and durations of 300 ns penetrate into the interior of tumor cells and cause tumor cell nuclei to rapidly shrink and tumor blood flow to stop. Melanomas shrink by 90% within two weeks following a cumulative field exposure time of 120 μs. A second treatment at this time can result in complete remission. This new technique provides a highly localized targeting of tumor cells with only minor effects on overlying skin. Each pulse deposits 0.2 J and 100 pulses increase the temperature of the treated region by only 3 °C, ten degrees lower than the minimum temperature for hyperthermia effects. KeywordsSkin cancer; Cancer therapy; Tumor; Pulsed electric fields; Pyknosis; Inhibiting angiogenesis; DNA; Nucleus Electric fields have been employed in several different types of cancer therapy. Some of these involve radiofrequency or microwave devices that heat the tumor to greater than 43 °C to kill the cells via hyperthermia [1,2]. Others use pulsed electric fields to permeabilize the tumor cells to allow the introduction of toxic drugs or DNA [3][4][5]. We have discovered that ultrashort electrical pulses can be used as a purely electrical cancer therapy that kills tumors without hyperthermia or drugs. Previous work from this laboratory found that fibrosarcoma tumors treated in vivo with ten 300 ns pulses exhibited a reduced growth rate compared to control tumors in the same animal [6]. Here, we report that when melanoma tumors are treated with four hundred of these pulses, tumors shrink by 90% within two weeks and a subsequent treatment can result in complete remission.The main characteristics of these nanosecond pulsed electric fields (nsPEF) are their low energy that leads to very little heat production and their ability to penetrate into the cell to permeabilize intracellular organelles [7,8] and release calcium [9][10][11] from the endoplasmic reticulum [11]. They provide a new approach for physically targeting intracellular organelles with many applications, including the initiation of apoptosis in cultured cells [12][13][14] and tumors [6], enhancement of gene transfection efficiency [13,14], and inhibiting tumor growth [6]. During the past year, we have treated over 300 murine melanomas in 120 mice with 40 kV/cm electric field pulses 300 ns in duration with dramatic results. Every tumor exposed to 400 such pulses exhibits rapid pyknosis and reduced blood flow and shrinks by an average of 90% within two The efficacy of this nsPEF treatment depends on two separate electric field parameters: pulse duration and amplitude. The effect of pulse duration can be understood by considering the process of membrane charging when the cell is placed in an electric field. Ions in the cell interior will respond to the electric field by moving in the field direction and charging the highly resistive membrane until they experience no further force. By definition this will only occur ...

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“…Increased permeabilization of the plasma membrane, typically demonstrated by measuring propidium iodide (PI) uptake, due to nsPEFs was delayed compared to traditional electroporation [5,6], indicating that it was an indirect effect. These nsPEFs induced cellular calcium release [7,8] and apoptosis in cells [9,10] or tumors [11,12]. These effects appeared to be due to the charging of interior organelle membranes and the ensuing creation of nanopores.…”

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confidence: 99%

Nanosecond electric pulses penetrate the nucleus and enhance speckle formation

Chen

Garner

Chen

et al. 2007

Biochemical and Biophysical Research Communications

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Nanosecond electric pulses generate nanopores in the interior membranes of cells and modulate cellular functions. Here, we used confocal microscopy and flow cytometry to observe Smith antigen antibody (Y12) binding to nuclear speckles, known as small nuclear ribonucleoprotein particles (snRNPs) or intrachromatin granule clusters (IGCs), in Jurkat cells following one or five 10 ns, 150 kV/cm pulses. Using confocal microscopy and flow cytometry, we observed changes in nuclear speckle labeling that suggested a disruption of pre-messenger RNA splicing mechanisms. Pulse exposure increased the nuclear speckled substructures by 2.5-fold above basal levels while the propidium iodide (PI) uptake in pulsed cells was unchanged. The resulting nuclear speckle changes were also cell cycle dependent. These findings suggest that 10 ns pulses directly influenced nuclear processes, such as the changes in the nuclear RNA-protein complexes. Published by Elsevier Inc.Keywords: Nucleus; Nuclear speckles; Small nuclear ribonucleoprotein particles/intrachromatin granule clusters; Intracellular electromanipulation; Nanosecond electric pulsesWe have been exploring a new nanosecond (ns) time domain in cell biology. When cells are exposed to intense (50-300 kV/cm) pulsed electric fields (PEFs) with rise times faster than the time it takes for charges on the plasma membrane to redistribute, the electric field can penetrate into the interior of the cell and organelles. If these fields exceed the breakdown threshold of these membranes, nanopores will form in the organelle membranes [1,2]. To better understand the mechanism involved, mathematical models for the cellular electropermeabilization due to nsPEFs have been developed [3,4]. The resulting changes in cellular structures and functions differ significantly from traditional electroporation [microseconod to millisecond pulsed electric fields (PEFs)] [1,5,6]. Increased permeabilization of the plasma membrane, typically demonstrated by measuring propidium iodide (PI) uptake, due to nsPEFs was delayed compared to traditional electroporation [5,6], indicating that it was an indirect effect. These nsPEFs 0006-291X/$ -see front matter Published by Elsevier Inc.

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Nanosecond, high‐intensity pulsed electric fields induce apoptosis in human cells

Cited by 418 publications

References 38 publications

Stimulation of Capacitative Calcium Entry in HL-60 Cells by Nanosecond Pulsed Electric Fields

Stimulation of Capacitative Calcium Entry in HL-60 Cells by Nanosecond Pulsed Electric Fields

Nanosecond pulsed electric fields cause melanomas to self-destruct

Nanosecond electric pulses penetrate the nucleus and enhance speckle formation

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