Figure 8.

S1P₂ regulates intracellular ROS production from LPS-primed BMDMs followed by S1P exposure. Cells were treated with JTE013 (10 μM) or NAC (5 mM) for 30 min. BMDMs were then primed with LPS (500 ng/mL) for 4 h and exposed to S1P (1 μM) for 1 h. (A,B) Effects of JTE013 on intracellular ROS production were analyzed by DCF-DA staining of LPS/S1P-treated BMDMs (LPS + S1P). Representative images of cells bearing DCF (A) and quantification of ROS levels (B). Scale bar, 10 μm. n = 4 per group. *** p < 0.001 versus control BMDMs (Veh). ### p < 0.001 versus LPS/S1P-treated BMDMs (LPS+S1P). (C,D) Effects of NAC on NLRP3 upregulation were determined by Western blot in LPS/S1P-treated BMDMs (LPS+S1P). Representative blot of NLRP3 (C) and quantification (D). n = 4 per group. *** p < 0.001 versus control BMDMs (Veh). (E–G) Effects of NAC on NLRP3 inflammasome activation were analyzed by Western blot in LPS/S1P-treated BMDMs (LPS+S1P). Representative blots of pro-caspase-1, cleaved caspase-1, pro-IL-1β, and mature IL-1β € and quantification (F, caspase-1 activation; G, IL-1β maturation). n = 4 per group. *** p < 0.001 versus control BMDMs (Veh). ### p < 0.001 versus LPS/S1P-treated BMDMs (LPS + S1P).