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. 2022 Dec 8;13:1060441. doi: 10.3389/fimmu.2022.1060441

Figure 2.

Figure 2

LXA4 attenuates the activation of NLRP3 inflammasome induced by MSU crystals in macrophages. BMDMs and PMA-differentiated THP-1 macrophages were primed with LPS (300 ng/ml) for 3 h and then pretreated with LXA4 (50–150 nM) for 1 h, and subsequently, 100 μg/ml of MSU crystal was added to stimulate for 24 h. (A, B) Supernatants were analyzed by ELISA to measure IL-1β levels in BMDMs and PMA-differentiated THP-1 macrophages. (C–F) Supernatants and cell lysates were analyzed by immunoblotting to determine the release of cleaved caspase-1 and matured IL-1β and the cleavage of GSDMD-F into GSDMD-N in BMDMs and PMA-differentiated THP-1 macrophages. PMA-differentiated THP-1 macrophages were primed with LPS (300 ng/ml) for 3 h and then pretreated with LXA4 (50-150 nM) for 1 h, and subsequently, 100 μg/ml MSU crystals were added to stimulate for 6 h. (G) Cell death was detected by propidium iodide (PI) and Hoechst 33342 staining. Scale bars, 50 µm. (H) The percentage of PI-positive cells relative to all cells was calculated in PMA-differentiated THP-1 macrophages. (I) The release of LDH in supernatants was detected in PMA-differentiated THP-1 macrophages (n=4). Data are presented as the mean ± SEM, ** p<0.01 compared with the vehicle group, # p<0.05 compared with the MSU group.