Research Paper
Extracellular histones induce inflammation and senescence of vascular smooth muscle cells by activating the AMPK/FOXO4 signaling pathway
Figure 2. Extracellular histones facilitate NLRP3 inflammasome assembly. VSMCs were treated with various concentrations of histones (0, 10, 25, 50, and 100 μg/mL) for 6 h. (A) Western blotting was performed to analyze NLRP3, ASC, and caspase-1 inflammasome protein expression. (B) Quantitative analysis of (A). (C) RT-qPCR was performed to determine the mRNA expressions of NLRP3, ASC, and caspase-1. (D) Double immunofluorescence staining was performed to explore ASC and NLRP3 expression and colocation (green, ASC; red, NLRP3; blue, DAPI). Bar = 25 μm. (E) Quantitative analysis of the fluorescence intensity of ASC and NLRP3 from (D). (F) VSMCs were treated with or without histones (100 μg/mL), and CoIP was performed to examine the interaction of NLRP3 and ASC. For (B, C and E), data are from three independent experiments; mean ± SEM; Student's t-test, *P < 0.05, **P < 0.01, and ***P < 0.001 vs. the corresponding control.