Relative concentration of the main emissive species in the guided ionization wave (conditions: He gas flow = 2slm; voltage frequency = 10kH; voltage amplitude = 5kV; voltage duty cycle = 1%.

<p>Optical fiber placed 5mm after the exit ceramic tube).</p

He-GIW induces a necrotic cell death.

<p><b>(a-c)</b> Presence of apoptosis was assayed by FACS analysis of Annexin V/PI dual staining. <b>(a)</b> An example of flow cytometry profile 6 h after treatment with He-GIW or with staurosporine, a positive control for apoptosis. X axis represents annexin-V-APC staining and Y axis PI staining. <b>(b)</b> Percentage of cells positive for Annexin V only (black), PI only (white) or both (grey) was measured 6 h and 24 h after treatment with He-GIW or staurosporine in standard cell culture conditions (c) or with 1% or 5% FCS. <b>(a-c)</b> are representative of 3 independent experiments. <b>(d)</b> Caspase 3 cleavage (red) was assayed by immunofluorescence 6h after He-GIW or staurosporine treatment. Cell nuclei are labeled with Hoechst and appear blue. <b>(e)</b> Caspase 3 cleavage was monitored by Western Blot for 2 h after He-GIW treatment. <b>(f)</b> Adherent cell percentage 6 h after He-GIW treatment in presence or absence of caspase inhibitor Z-VAD. Error bars represent S.E.M. of three independent experiments. p<0.05 (*), p<0.01 (**) and p<0.001 (***)</p

Main emissive species produced with Helium as plasma carrier gas.

<p>Main emissive species produced with Helium as plasma carrier gas.</p

Device settings and cell culture parameters influence cell fate.

<p><b>(a)</b> hPDL were treated with He-GIW using different voltages and <b>(b)</b> exposure lengths. Adherent cell number was evaluated 24h after treatment and expressed as a percentage of control. <b>(c)</b> Cell morphology was monitored by phase contrast microscopy 6h and 24h after treatment. <b>(d)</b> Cell culture incidence on He-GIW treatment was evaluated using different FCS concentrations and <b>(e)</b> cell confluence. Adherent cell number was assayed 24h after treatment and expressed as percentage of control. Error bars represent S.E.M. of three independent experiments p<0.05 (*), p<0.01 (**) and p<0.001 (***).</p

Mitochondrial membrane depolarization precedes cell membrane alteration.

<p><b>(a)</b> Adherent cell percentage was monitored for 6 h after He-GIW treatment. <b>(b)</b> Cell Δψm was monitored by FACS analysis for 6h using DiOC6 staining. Results of one representative experiment of at least three. <b>(c)</b> Geometric means of DiOC6 staining relative to control 1, 3 or 6 h after He-GIW treatment. <b>(d)</b> Δψm (DiOC6) and plasma cell membrane permeability (7AAD) were monitored by FACS analysis for 6 h after He-GIW treatment. When present, error bars represent S.E.M. of three independent experiments p<0.05 (*), p<0.01 (**) and p<0.001 (***)</p

Experimental set-up.

<p>Experimental set-up.</p

Electrical high voltage and current signals analyzed during one impulse (2a) and during one period (2b) (He gas flow = 2slm, voltage frequency = 10kHz, voltage amplitude = 5kV, voltage duty cycle = 1).

<p>Electrical high voltage and current signals analyzed during one impulse (2a) and during one period (2b) (He gas flow = 2slm, voltage frequency = 10kHz, voltage amplitude = 5kV, voltage duty cycle = 1).</p