FIGURE 4.

Effect of PKC inhibitors on NETosis. Neutrophils were incubated with A23187 ionophore in the presence of 5 μM chelerythrine (A), or 20 μM chelerythrine (B). Antibody binding to deiminated histone H3 (green) and the DNA contained in cells or released as NETs (blue) was observed in 5 μM chelerythrine, whereas 20 μM inhibitor precluded deimination and most other morphologic features of NETosis. Cells incubated with PKCα/β pseudosubstrate peptide inhibitor deiminated histone H3 and proceeded to NET release (C). The PKCζ inhibitor peptide suppressed histone deimination and NET release but earlier stages of NETosis, such as a partial heterochromatin decondensation and nuclear de lobulation, were observed (D). As in Figure 1E, untreated neutrophils showed typical multilobed nuclei without apparent evidence of NETosis (E).