Fig. 5
From: Neutrophil extracellular trap formation requires OPA1-dependent glycolytic ATP production
Reduced glycolytic ATP production in neutrophils of Opa1N∆ mice is a consequence of reduced mitochondrial complex I activity. a ATP-dependent luciferase activity assay. Control human neutrophils were incubated with the indicated inhibitors for 70 min in a concentration-dependent manner. Relative luciferase unit (RLU) values are means ± SEM. **p < 0.01; ***p < 0.001; n = 3. b ATP/ADP bioluminescent assay. Neutrophils of Opa1N∆ and control mice were pre-treated with 3 mM 2-DG for 70 min and the ATP/ADP ratio was measured in the presence and absence of GM-CSF/C5a. RLU values are means ± SEM. **p < 0.01; n = 6. c Complex I (CI), complex III (CIII) and citrate synthase (CS) activities of mitochondria isolated from resting and GM-CSF/C5a-activated Opa1N∆ and control Hoxb8 neutrophils were assessed in the presence and absence of 10 µM rotenone (Rot) using a spectrophotometer. Values are means ± SEM. n.s., not significant; *p < 0.05; **p < 0.01; n = 3. d NAD/NADH bioluminescent assay. NAD+/NADH ratios in resting and GM-CSF/C5a-activated neutrophils of Opa1N∆ and control mice were measured after 70 min incubation in the presence and absence of 10 µM Rot using a NAD/NADH determination kit. RLU values are means ± SEM. *p < 0.05; **p < 0.01; ****p < 0.0001; n = 5. e Lactate bioluminescent assay. Lactate levels in resting and GM-CSF/C5a activated neutrophils of Opa1N∆ and control mice were measured after 70 min incubation in the presence and absence of 3 mM 2-DG using a lactate determination kit. RLU values are means ± SEM. n.s., not significant; *p < 0.05; **p < 0.01; ***p < 0.001; n = 4. ECAR and OCR data are provided in Supplementary Fig. 9
