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

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

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

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

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

HFD-induced peritoneal macrophage M2 polarization and rosiglitazone treatment potentiated this phenotype.

<p>(A) Dectin-1, MR, CD36, CD11b and TLR2 protein expressions on peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) evaluated by flow cytometry (n = 6). (B) <i>Arginase 1</i>, <i>YM-1</i> and <i>YM-2</i> mRNA expressions on peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) by RT-PCR (n = 6). Data are represented as mean ± SEM *p<0.05 and **p<0.01 compared to NC control (NC C); #p<0.05 and ##p<0.01 compared to HF control (HF C). The data are representative of three independent experiments.</p

Primers sequences used in quantitative PCR experiments.

<p>Primers sequences used in quantitative PCR experiments.</p

Rosiglitazone switches HFD-induced M2b polarization towards M2a.

<p><i>(A) TNF-α</i>, <i>IL-1β</i>, <i>IL-6 and IL-10</i> mRNA expressions in peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) by RT-PCR (n = 6). (B) TNF-α and IL-10 release by peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) by ELISA (n = 6). (C) <i>PPARγ</i>, <i>PPARα</i> and <i>PPARβ/δ</i> mRNA expressions in peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) by RT-PCR (n = 6). (D) PPARγ protein level in peritoneal macrophages from mice fed a chow (NC) or a HFD (HF) and treated or not (C) with rosiglitazone (R) or WY14643 (WY) by western blot. Data are represented as mean ± SEM *p<0.05 and** p<0.01 compared to NC control (NC C); #p<0.05 compared to HF control (HF C). The data are representative of three independent experiments.</p