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. 2016 Apr 28;7(22):31772–31789. doi: 10.18632/oncotarget.9091

Figure 2. Isoflurane impairs zymosan-induced NF-κB activation, iNOS expression and NO/ONOO− production in neutrophils.

Figure 2

A. Neutrophils were treated with increasing concentrations of zymosan for indicated time periods, and were subjected to Western blot analysis. B. Neutrophils were incubated with zymosan for 15 min, post-treated with 0.7% isoflurane for 15 min, followed by a continuous incubation with zymosan for a total of 8 h before Western blot analysis. β-actin was used as an inner control for whole cell lysates. C. Neutrophils were co-transfected with or without an iNOS-luc reporter plasmid and indicated siRNAs. The promoter activity of iNOS was determined in the cell lysates after zymosan treatment for 6 h. D., E. Neutrophils were pretreated with NAI (D) or transfected with NF-κB p65 siRNA (E), followed by incubation with zymosan for 8 h. The nuclear or whole cell lysates were prepared for Western blot analysis. Lamin B was used as an inner control for the lysates of the nuclear fraction. F. Neutrophils were treated as described in (D), followed by incubation with zymosan for 6 h prior to RT-PCR analysis. G. Neutrophils were treated as described in (D), followed by measurement of NO and ONOO− production. H. Neutrophils were treated as described in (B), and the nuclear lysates were prepared for Western blot analysis. Data are represented as the mean ± SEM of 3 replicates or representative of 3 independent experiments. *P < 0.05, #P < 0.01, as compared with the cells exposed to vehicle alone (A). *P < 0.05, #P < 0.01, as compared with the cells exposed to zymosan alone (B-H), or zymosan + scrambled siRNA (E).