Figure 4
From: Chemically different non-thermal plasmas target distinct cell death pathways
Necrostatin-1 (Nec-1, a potent and selective inhibitor of necroptosis) antagonizes the He NTP-induced cytotoxicity. Cell viability as detected by the WST-1 assay of (a) 3T3 fibroblasts and (b) MSCs treated with air, helium NTPs or ozone for indicated time periods with supplementation of 10 µM Nec-1, measured 24 h after exposure. Readings were done in quadruplicates. The data present the mean values of four independent experiments. Data are expressed as means ± SEM (n = 4), *P < 0.05 **P < 0.01. (c) He NTP and ozone treatment induces RIP1 and RIP3 upregulation (full blots of RIP1 and RIP3 are presented in Fig. S5 in Supporting Information) without concomitant activation of caspase-8. 3T3 fibroblasts and MSCs were treated with air, helium NTPs or ozone for 30 s. Cells were analyzed by Western immunoblotting 4 h after treatment. Actin – control of equal protein loading. The graphs show densitometric quantification of RIP1 and RIP3 Western immunoblots, *P < 0.05 **P < 0.01, mean ± SEM, n = 3. Representative blots out of three independent experiments are shown. (d) RIP3 complexes isolated by immunoprecipitation from either 3T3 fibroblasts or MSCs treated with air, He NTPs and ozone for 30 s. The graph shows densitometric quantification of the respective immunoblots, *P < 0.05 **P < 0.01, mean ± SEM, n = 3. Representative blots out of three independent experiments are shown.
