Fig. 3: Knockdown of TIGIT in healthy CD4⁺ T cells recapitulates the proinflammatory phenotype of PM TIGIT^low CD4⁺ T cells.

a, b Knockdown of TIGIT was confirmed by flow cytometry. Data from five biologically independent replicates. c Knockdown of TIGIT in CD4⁺ T cells from HC or PM was confirmed by western blot. The experiment was repeated three times independently with similar results. d–g CD4⁺ T cells were labeled with CFSE and stimulated with αCD3/CD28 for 3 d. T-cell proliferation was evaluated by flow cytometry. Data from five biologically independent replicates. h, i IFNγ, and IL-17A expression in CD4⁺ T cells was measured by flow cytometry. Data from five biologically independent replicates. d–r Naive CD4⁺ T cells from HC were stimulated by αCD3/CD28 for 1 d, and then transfected with scramble or TIGIT siRNA by electroporation. Following transfection, cells were subjected to culture conditions for Th1, Th2, Th17, and Treg cell differentiation for an additional 4 d, respectively. j–x Naive CD4⁺ T cells from HC were stimulated by αCD3/CD28 for 1 d, and then transfected with scramble or TIGIT siRNA by electroporation. Following transfection, cells were subjected to culture conditions for Th1, Th2, Th17 and Treg cell differentiation for an additional 4 d, respectively. j, k Knockdown efficacy of TIGIT was confirmed by flow cytometry. Data from five biologically independent replicates. l–x Th1 condition: T-bet and IFNγ (l–n), Th2 condition: GATA3 and IL-4 (o–r), Th17 condition: RORγt and IL-17A (s–v), and Treg condition: FoxP3 (w, x) expression in CD4⁺ T cells were measured by flow cytometry. Representative contour plots or histograms were shown. Data from five biologically independent replicates. All data are mean ± SEM. Statistics were done by a two-tailed unpaired Student’s t test.