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. 2021 Oct 27;10(11):1706. doi: 10.3390/antiox10111706

Figure 5.

Figure 5

S1P2 knockdown attenuates S1P-induced NLRP3 inflammasome activation in LPS-primed BMDMs. BMDMs transfected with non-target control siRNA (siCON) or S1P2-specific siRNA (siS1P2) were primed with LPS (500 ng/mL) for 4 h and exposed to S1P (1 μM) for 1 h. (A) Effects of S1P2 knockdown on ASC speck formation were determined by double immunofluorescence for ASC (green) and NLRP3 (red) in LPS/S1P-treated BMDMs (LPS+S1P). Representative images of ASC speck formation in NLRP3-positive BMDMs. Scale bar, 10 μm. (B–D) Effects of S1P2 knockdown on caspase-1 activation and IL-1β maturation were analyzed by Western blot. (B) Representative blots of pro-caspase-1, cleaved caspase-1, pro-IL-1β, and mature IL-1β. Quantification of caspase-1 activation (C) and IL-1β maturation (D). (E) Effects of S1P2 knockdown on IL-1β secretion were measured by ELISA. n = 4 per group. *** p < 0.001 versus control BMDMs transfected with control siRNA (siCON + Veh). ## p < 0.01 and ### p < 0.001 versus LPS/S1P-treated BMDMs transfected with control siRNA (siCON + LPS + S1P).