FIGURE 4.

Effects of the reactive oxygen scavenger NAC and Caspase1 inhibitor VX765 on oxidative stress and pyroptosis after luteolin treatment. (A) Changes in intracellular ROS levels. Red: control group, Blue: luteolin group, Green: NAC group, Orange: VX765 group (B) Changes in SOD content. (C) Changes in GSH content. (D) Changes in H₂O₂ content. (E) The contents of IL-1β in the cell supernatants. (F) Changes in mitochondrial membrane potential. (G) The control, luteolin-treated and VX765-treated HT-29 cells were collected and analysed by WB to assess the expression levels of NLRP3, ASC, Pro-Caspase1, Caspase1, GSDMD, Cleaved-GSDMD (N-T), and IL-1β. GAPGH, β -Tubulin and β-actin served as internal controls. (H) Immunofluorescence colocalization of NLRP3 with GSDMD in the control and luteolin-treated cells. Cells were double stained for NLRP3 (Cy3) and GSDMD (FITC). DAPI was used to stain the nucleus. The cells that were labelled with three colours are those where pyroptosis occurred. The results are presented as the mean ± SEM, n = 3. *p < 0.05, **p < 0.01, and ****p < 0.0001 vs. the control group.