Fig. 1

Pyroptosis of CD4⁺ T cells is increased in RA patients. A Representative immunoblots showing ASC, NLRP3, cleaved Caspase-1, and GSDMD-N expression in CD4⁺ T cells from NCs and RA patients. B Quantification of ASC, NLRP3, cleaved Caspase-1, and GSDMD-N protein expression shown in A (n = 3). C Concentrations of secreted IL-1β measured by ELISAs (n = 5). D CD4⁺ T cell death was quantified for control and RA CD4⁺ T cells on day 3 after stimulation by flow cytometric analysis of Annexin V⁺ 7AAD⁺ CD4⁺ T cells (n = 5). Representative flow plots of Annexin V and 7AAD staining of CD4⁺T cells and a histogram of the percentage of Annexin V⁺ 7AAD⁺ CD4⁺T cells are shown. E CD4⁺ T cell death was quantified by measuring LDH release (n = 5). F Cleaved Caspase-1 was identified by FLICA staining and flow cytometric analysis. Representative scatterplots of FLICA⁺ CD4⁺ T cells in the controls and RA patients are shown. G NLRP3, Caspase-1, GSDMD, and ASC transcripts compared in NCs and RA CD4 + T cells. All data are presented as the mean ± SD. Statistical significance was determined by Student’s t test; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001. NC, normal control; RA, rheumatoid arthritis; ASC, apoptosis-associated speck-like protein containing a CARD; NLRP3: NOD-like receptor protein 3; CASP1, Caspase-1; GSDMD-N, N-terminal pore-forming domain of GSDMD