Fig. 4. LAC induced necroptosis in neutrophil.

a, b Human neutrophils were stimulated with PBS, LAC (MOI 10), LAC culture supernatant, or HKLAC. Level of phosphorylated MLKL (pMLKL), MLKL, and β-actin in neutrophil lysate were detected by western blot. c, e Human neutrophils pretreated with 1, 3, and 10 μM necrosulfonamide (NSA) or 50, 100, and 200 μM Nec for 1 h at 37 °C and 5% CO₂, and then were stimulated with bacterial supernatant for 1 h. The release of LDH by neutrophils was detected (n = 3). d, f Levels of pMLKL, MLKL, and β-actin in neutrophil lysate in (c) or (e) treatment were detected by western blot. Data were shown as mean ± SE (c, e). *P < 0.05, ***P < 0.0001 vs. supernatant treated group; ^###P < 0.0001 vs. PBS-treated group