Fig 1: T cells respond to influenza vaccine and virus antigens with different functional abilities(A) UMAP (generated from scRNA-seq data) of sorted CD4 T cells from organoids collected on days 4, 7, 10, and 14 from n = 4 donors (1 experiment).(B) Bubble plot of top differentially expressed genes used to identify T cell clusters. Bubble size indicates percentage of cells expressing the marker and color indicates magnitude of expression.(C) Frequencies of key T cell clusters in organoids at different time points poststimulation with influenza antigen modalities (n = 4 donors, 1 experiment).(D) Frequencies of GC TFH following organoid stimulation (n = 12 donors, 1 experiment) as quantified by flow cytometry.(E) GC TFH frequencies at the peak of CD4 T cell activation (day 7) as quantified by flow cytometry (n = 5 donors, 1 experiment).(F) Top gene ontology (GO) terms of genes upregulated in LAIV relative to IIV-stimulated organoids in TFH and GC TFH on day 7.(G) Violin plot of candidate genes (mapping to “cytokine-mediated signaling pathway”) upregulated with LAIV relative to IIV stimulation in TFH and GC TFH on day 7.(H) Cytokine production by GC TFH under different stimulation conditions on day 7 as quantified by intracellular staining (n = 12 donors, 1 experiment).(I) Frequencies of Th1- (CXCR3+CCR4-CCR6-), Th2- (CXCR3-CCR4+CCR6-), and Th17-like (CXCR3-CCR4+CCR6+) GC TFH on day 7 as quantified by flow cytometry (n = 12 donors, 1 experiment). *p < 0.05, **p < 0.01, and ***p < 0.001 using a Kruskal-Wallis test followed by paired Mann-Whitney U tests to compare groups. p values shown are for comparisons against the unstimulated control unless otherwise indicated by lines.(J) Bar graphs comparing Chao diversity metric within activated (OX40+) T cells, TFH, and GC TFH on day 7 poststimulation.(K) Secreted cytokines and chemokines in organoid supernatants on day 4, measured using Luminex (n = 4 donors, 1 experiment). Boxplots show the median, with hinges indicating the first and third quartiles and whiskers indicating the highest and lowest value within 1.5 times the interquartile range of the hinges.See also Tables 1, S2, and S3 and Figures S4–S6.