Figure 4. M1 protein protects against histones in NETs.
2 × 105 neutrophils were stimulated for 4 h with 25 nM PMA to induce NETs and infected with GAS WT, Δemm1 mutant and complemented strain (Δemm1+pM1) at an MOI of 0.1. Surviving CFUs were calculated relative to initial inoculum after 15 min of infection in (A) untreated cells, (B) cells treated with 100 U/mL DNase 1 to degrade the DNA backbone, or (C) cells treated with blocking anti-histone H2A antibodies. (D) Survival in NETs of GAS WT naïve or anti-M1 antiserum at MOI of 0.1 after 15 min of infection. All results are showing the percentage of bacterial survival relative to the bacterial inoculum. Each dot represents one sample from three independent experiments in triplicates and neutrophils were obtained from three different donors. Results shown represent average ± SEM values and were analyzed by Student’s t-test (n.s., not significant and *P < 0.05, ***P < 0.001).