Figure 3.

Increased expression of ATG5 in granulocyte-like cells does not affect their ability to form neutrophil extracellular traps (NETs). HL-60-ATG5 and mock control HL-60 cells were differentiated for 5 days with 1 µM ATRA and 70 mM DMF and stimulated with 100 nM phorbol 12-myristate 13-acetate (PMA) or 4 µM calcium ionophore A23187 (CI) or left unstimulated for 3 h. Neutrophil extracellular traps (NETs) formation was assessed qualitatively: (a,b) by conventional fluorescent microscopy and quantitatively (c,d) by fluorometric measurement of DNA release. (a,b) After the stimulation, 100 nM Sytox Green was added to the wells and NETs were visualized under the microscope using fluorescent and transient light, at 10× magnification. Representative images of one out of six independent experiments are shown. Results are shown as means + SEM from 6 independent experiments and were analyzed by one-way ANOVA with post hoc Dunn’s test.