Fig. 2: GSDMD-mediated pyroptosis is activated in keratinocytes treated with psoriasis-like stimulation.

Primary human epidermal keratinocytes (A–E) or primary mouse epidermal keratinocytes (F–H) were treated by M5 (10 ng/mL for each component) for 24 h, respectively. A The secretion of IL-1β, CXCL-2, CCL-20, IL-8 and S100A8/A9 was determined by ELISA (n = 3). B 11.5 h after M5 stimulation, PI was added to the cell supernatant. The video of live-cell imaging was taken by Deltavision Elite Imaging System. Screenshots shows the typical pyroptosis manifestation. C Proteins of interest in cell lysate and culture supernatant were determined by western blotting assay (n = 3). D The ___location of GSDMD was presented by immunofluorescence assay. Nucleus was stained by Hoechst. Actin filaments was stained by phalloidin (n = 3). E PI positive cells (red) and Ki-67 positive cells (green) were detected by immunofluorescence assay (n = 5). F–H Mouse rnGSDMD was added to the cultural supernatant or electroporated into primary mouse keratinocytes, respectively. F Changes of cell morphology were presented. G IL-1β secretion was evaluated by ELISA. H The cleavage of GSDMD was determined by western blotting assay (n = 3) Scale bar represents 50 μm in B. Scale bar represents 30 μm in D. Scale bar represents 100 μm in E and F. *p < 0.05, **p < 0.01, ***p < 0.001. GSDMD-FL GSDMD full length, N-GSDMD N-terminal GSDMD.