Elevated H2AX Phosphorylation Observed with kINPen Plasma Treatment Is Not Caused by ROS-Mediated DNA Damage but Is the Consequence of Apoptosis

Figure 3

Dependence of ROS and UV-induced γH2AX expression on intracellular signaling and apoptosis. (a, b) γH2AX expression in cells preincubated with (a) SB202190 (p38 MAPK-inhibitor) or (b) Z-VAD-FMK (pan-caspase inhibitor) 2 h after exposure to various agents. (c) Representative overlay histograms of active caspase 3/7-stain (left) and terminally dead DAPI⁺ (right) in presence of absence of Z-VAD-FMK at 24 h after plasma treatment. (d–e) Quantification of apoptosis in presence or absence of Z-VAD-FMK at (d) 4 h and (e) 24 h after plasma treatment with γH2AX-inducing agents. (f) Confocal laser scanning microscopy (DNA = DAPI, blue; γH2AX = green) of TK6 cells with arrows pointing at apoptotic (cells with fragmented nuclei) cells being positive for γH2AX. Data show box plots (a, b) and single data and mean (d, e) of two to four independent experiments with several replicates each. Statistical analysis was done using -test. Scale bar (f) is 10 μm; n.s. = not significant.

(a)

(b)

(c)

(d)

(e)

(f)