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. 2021 Jul 8;14:3063–3075. doi: 10.2147/JIR.S317672

Figure 4.

Figure 4

Melatonin suppresses MPP+-induced NLRP3 inflammasome activation in vitro. (A‒D) Immunoblot analysis of NLRP3, caspase 1 (full-length and cleaved forms) from mouse BV2 cells primed with MPP+ (500 μM, 6 h), followed by ATP (2.5 mM, 30 min) or nigericin (10 μM, 45 min); or MPP+ (500 μM, 6 h) + melatonin (100 μM, 6 h) followed by ATP (2.5 mM, 30 min) or nigericin (10 μM, 45 min). (E) Quantification of IL-1β in the BV2 cell culture supernatants. (F and G) Intracellular levels of ROS were detected in MPP+-treated (500 μM, 6 h) BV2 cells followed by ATP (2.5 mM, 30 min), with or without melatonin (100 μM, 6 h) pretreatment. Mean fluorescence intensity was quantified by FlowJo in (G). (H) Representative immunofluorescence image of primary microglia treated with PBS or MPP+ followed by ATP, with or without melatonin pretreatment, stained with anti-ASC antibody (green) and anti-IBA1 antibody (red). DAPI represents the nuclear signal (blue). Scale bars = 25 μm. White arrows indicate ASC specks. (I) Quantification of relative ASC speck intensity per IBA1-positive cells. aMT, melatonin; Nig, Nigericin. *P < 0.05, **P < 0.01, ***P<0.001. Data are expressed as the mean ± SEM, all experiments were repeated at least 3 times.