Extended Data Fig. 1: The cell death of T_FH cells is not regulated by apoptosis or pyroptosis.

a, b, As indicated by the schematic of bone marrow (BM) reconstitution to generate chimeric mice, indicated mixtures of BM cells were transferred into lethally irradiated Rag1^–/– recipients. After 6 weeks, chimeric mice were subcutaneously immunized with NP-OVA in Alum. b, At day 14 post immunisation, frequencies of T_FH and B_GC cells derived from WT or transgenic were analysed by flow cytometry (n = 5 for control and n = 6 for Bcl-2-Tg and Casp1/11-KO). c, C57/BL6/J mice were subcutaneously immunized with NP-OVA in Alum. Cell size of T_FH and non-T_FH cells at day 7 post immunisation was analysed by flow cytometry (n = 6). d, Congenic marked C57/BL6/J mice were adoptively transferred of OT-II cells and then intraperitoneally immunised with OVA in Alum. At day 7 post immunisation, cell size of T_FH and non-T_FH OT-II cells was analysed by flow cytometry (n = 6). e, Mice were intraperitoneally immunised with SRBC, T_FH and non-T_FH cells were sorted at day 8 post immunisation. Lipid ROS production in T_FH and non-T_FH cells was analysed by flow cytometry using C11-BODIPY (n = 5). f, Flow cytometric analysis of lipid ROS production in purified tonsillar T_FH and non-T_FH cells (n = 4). Each dot represents one individual. g, Purified naïve CD4⁺ T, non-T_FH, and T_FH cells from human tonsils were stimulated with αCD3/CD28 and treated with different doses of RSL-3 or H₂O₂ for 12 h. Frequencies of Annexin V^–7-AAD^– viable cells were assessed by flow cytometry (n = 3). Data are representative of three independent experiments. For b, g, data are expressed as mean ± SEM. Data are analysed by two-sided Student’s t-test (b), paired-sample t-test (c-f), or one-way ANOVA (g). For g, bonferroni correction was used to adjust the significance level by using a value of 0.017 for each comparison. Data are representative of three (c–e) experiments with at least three mice per group.

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