Figure 5. HSPC-mediated antigen presentation induces a suppressive phenotype in CD4⁺ T cells.

See also Figure S3 and S4.

(A) Principle component analyses (PCA) of Nanostring (left) and RNA-Seq (right) gene expression of OT-II CD4⁺ T cells activated by HSPCs (T_HSCs) or dendritic cells (T_DCs), n=3-4.

(B) Top T_HSC-enriched gene sets of gene set enrichment analyses (GSEA) of RNA-Seq data from (A), comparing T_HSCs and T_DCs.

(C) Heatmap representing z-scored expressions of co-inhibitory module genes (Chihara et al., 2018).

(D-F and K) Ex vivo CD4⁺ T cell suppression assays. (D) Representative plots for the 1:2 suppressive/bystander naïve CD4⁺ T cell condition. (E) Suppression index for different bystander/suppressive ratios, n=4. (F) Proliferation index of responder CD4⁺ T cells for the 1:2 ratio, n=4.

(G) Ex vivo CD8⁺ T cell suppression assay, n=4.

(H and I) Ex vivo CD8⁺ T cell activation (H) and annexin V cytotoxicity assay (I), n=4. OVA1: ovalbumin 257-264, OVA2: ovalbumin 323-339.

(J) Ex vivo macrophage polarization assay, n=4.

(K) Role of IL-10 in T_HSC-mediated suppression. Activation of WT (left) or Il10rb^−/− (right) bystander T cells in the presence of T_HSCs or T_DCs in a 1:2 suppressive:bystander ratio., n=4.

(L) In vivo suppression assay. Representative plots (left) and quantification of bystander T cell proliferation (right), n=3.

Individual values are shown in A and C, means and SEM are depicted otherwise. No significance = ns, P<0.05 *, P<0.01 **, P<0.001 ***, P<0.0001 ****. One- (F, G, H, I, L) or two-way ANOVA (K) were performed as discovery test. Two-way ANOVA was performed in E and J. If not stated otherwise, unpaired two-tailed t-tests were performed as post-hoc tests.