Fig. 4.

Electronic vapor extract (EVE) exposure inhibits production of neutrophil extracellular traps (NETs). A: extracellular DNA/NET production in response to EVE was quantified using the fluorescent DNA dye PicoGreen (n = 5). B: the PicoGreen method was also used to quantify the effects of EVE exposure on NET production in response to a 2-h treatment with NET inducer phorbol 12-myristate 13-acetate (PMA) (n = 3). C: to assess the concentration dependence of NETosis inhibition by EVE, the effect of a range of concentrations between 1 and 100% on PMA-induced NETosis was tested (n = 7). D: an immunocytochemical approach was used to image NETs produced by untreated neutrophils [control (CTL)] or PMA-treated neutrophils (2 h treatment) incubated in control or EVE-containing media (green, myeloperoxidase; blue, Hoechst-33342; scale bar = 50 µm). Regions were randomly selected for imaging and are representative of 5 separate experiments. E: effect of EVE on noncanonical NETosis induced by nigericin (NGN) was assessed using the PicoGreen method (n = 8). Data shown are expressed as mean values ± SE; where applicable, results were analyzed by 1-way analysis of variance and post hoc Dunnett’s multiple-comparisons test. *P < 0.05 and **P < 0.01 versus PMA or nigericin controls.