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. 2021 Jan 26;22(3):1194. doi: 10.3390/ijms22031194

Figure 5.

Figure 5

Inhibitory effect of KMU-1170 on LPS-induced activation of NLRP3 inflammasome and anti-inflammatory potentiality of KMU-1170 in THP-1 cells. (A) Cells were differentiated into macrophages for 24 h using PMA (100 nM). And then the cells were treated with LPS (1 μg/mL) and/or ATP (1 mM) for 6 h after pretreatment with 1 μM KMU-1170 for 1 h. Cell lysate (Lysate) and media supernatant (Sup) were isolated and used to measure the protein expression levels of pro-IL-1β and IL-1β for the Sup as well as NLRP3, ASC, pro-caspase-1, pro-IL-1β, and β-actin for the Lysate by Western blot analysis. (B) Image-J software was used to analyze the relative optical density of the NLRP3, ASC, pro-caspase-1, and pro-IL-1β band in the Lysate. (C) Cells were differentiated into macrophages for 24 h using PMA (100 nM). And then the cells were treated with LPS (1 μg/mL) for 6 h after pretreatment with KMU-1170 (1 μM), celecoxib (25 μM), ibrutinib (5 μM), luxolitinib (50 μM), and metotrexate (1 μM) for 1 h. Whole cell lysates were isolated and used to measure the protein expression levels of pro-IL-1β and β-actin by Western blot analysis. (D) Image-J software was used to analyze the relative optical density of the pro-IL-1β band. † p < 0.05, * p < 0.01, and # p < 0.001 compared to LPS alone.