Fig. 4.

Engagement of P2X₇ receptors. Monocytes were preincubated with KN62 (2.5 μM) or DMSO (0.5%; control; a), oxATP (300 μM) or medium (control; b), and an anti-P2X₇ mAb (5 μg/ml) or the respective IgG2b isotype (5 μg/ml; c) for 25 min before NAD⁺ (200 μM) was added for 80 s. Intracellular Ca²⁺ levels were measured as the change in the 340 nm/380 nm emission ratio. Values obtained in the presence of NAD⁺ were set as the 100% reference. Results are means ± SEM of seven experiments, ***P < 0.001 (Student’s t test; a); four experiments, *P < 0.05 (Student’s t test; b); or three experiments, **P < 0.01 (Student’s t test; c). Monocytes were preincubated with KN62 (2.5 μM) or DMSO (0.5%; control; d), oxATP (300 μM) or medium (control; e), and an anti-P2X₇ mAb (5 μg/ml) or the respective IgG2b isotype (5 μg/ml; f) for 25 min before ATP (100 μM) was added. Shown are Δ ratios 340/380 from one representative measurement. *P < 0.05 (Mann–Whitney rank sum test) Δ ratio 340/380 ATP + DMSO versus ATP + KN62 (n = 4); *P < 0.05 (Mann–Whitney rank sum test) Δ ratio 340/380 ATP + medium versus ATP + oxATP (n = 4); *P < 0.05 (Mann–Whitney rank sum test) Δ ratio 340/380 ATP + isotype versus ATP + P2X₇ mAb (n = 4)