Fig. 3: Working mechanism of GSDMD-induced pyroptosis and ESCRT III-mediated cell membrane repair.

a Schematic illustration of the pyroptosis-related signaling pathway. b Western blot assay of the pyroptosis signaling pathway in 4T1 cells after treatments of PBS, VNP, GD (GSDMD protein cage), VNP-GD (GSDMD protein cage-conjugated VNP), and VNP-GD+EI-NP (GSDMD protein cage-conjugated VNP + EI-NP). The experiments were repeated three times independently. c Schematic illustration of the calcium influx induced ESCRT III-mediated membrane repair during cancer cell pyroptosis. d Flow cytometry assay of the Ca²⁺ influx detection with Fluo-8 AM in 4T1 cells after treatment with PBS, VNP, GD, VNP-GD, and VNP-GD+EI-NP for 24 h (n = 3 biologically independent samples). e Direct observation of the ESCRT III-mediated 4T1 cancer cell membrane repair during pyroptosis after treatment with VNP-GD and VNP-GD+EI-NP for 24 h (n = 4 biologically independent samples). The cell membrane was stained with Annexin V. The 4T1 cells were genetically engineered to express mCherry-labeled charged multivesicular body protein 3 (CHMP 3) protein, which is the main component of ESCRT III machinery. Scale bar: 5 µm. The experiments were repeated three times independently. Source data are provided as a Source Data file.