Figure 1

Nano-SiO₂ or nano-TiO₂ particles trigger active ATP release and IL-1β secretion through purinergic signalling and pannexin/connexin hemichannel activity. Nano-SiO₂ (a) or nano-TiO₂ (b) triggered active release of ATP in the supernatant by PMA-primed THP1 that peaks between 3 and 4 h. This ATP release was correlated with a secretion of IL-1β (a,b). Nano-ZnO did not induce ATP release or IL-1β secretion (c). Apoptotic (PI− anV+) and necrotic (PI+ anV−) cell death of primed THP1 was monitored using the AnnexinV/PI staining (d,e). ARL67156 (50 μM) was added to the supernatant during stimulation to limit ATP catabolism (a–c). IL-1β secretion by nano-SiO₂ or nano-TiO₂ was attenuated in THP1 cells stably expressing shRNA directed against ASC (sh ASC) or NLRP3 (sh NLRP3) in comparison with THP1 transfected with lamin-specific shRNA (sh CTL) (f). Nano-ZnO did not induce IL-1β secretion after 4 h of stimulation (f). Specific inhibition of P2X7R by A740003 partially decreased ATP release and IL-1β secretion by PMA-primed THP1 after 4 h nanoparticle stimulation (g,h). Connexin/pannexin channel blocker carbenoxolone (Cbx) and connexin channel blocker flufenamic acid (FFA) reduced both ATP release and IL-1β secretion upon nano-SiO₂ or nano-TiO₂ (g,h). PMA-primed THP1 stimulated for 4 h with 200 μM ATP, ADP or their stable derivatives ATPγS or ADPβS greatly increased IL-1β production in response to nanoparticles, whereas these nucleotides had no effect alone (i). Nanoparticles are at the concentration of 250 μg/ml (a–h) or 125 μg/ml (i). Data are representative of 2–4 independent experiments. Data are mean±S.D. of triplicates, compared between untreated and nanoparticle-stimulated THP1; ααα, P≤0.001 for THP1, βββ, P≤0.001 for THP1 sh CTL, γγγ, P≤0.001 for THP1 sh ASC, δδδ: P≤0.001 for THP1 sh NLRP3 (f). Data are mean±S.D. of triplicates, compared between nanoparticle-stimulated THP1 and nanoparticles plus inhibitor or agonist; *** and ααα, P≤0.001 for nano-SiO₂ and nano-TiO₂ stimulated THP1, respectively (g–i)