Figure 3.

OSCC secretome promote PD-1 and TIGIT expression and inhibit cytokine production in comparison with control secretome. (A) Representative dot plots of PD-1 and TIGIT expression in Tregs and Teff after co-culture with control or OSCC secretome. Briefly, sorted memory Tregs and Teff obtained from peripheral blood from 4 healthy donors were pre-activated with anti-CD3/CD28 beads (1:5) in the presence of IL-2 (1000U). After activation, 2x10⁵ Tregs and Teff were co-cultured with secretomes from OSCC or control samples for 48h and cells were stained with PD-1 and TIGIT, whereas the supernatants were used to measure cytokines using cytokine bead array. Expression of both suppressive molecules was measured by flow cytometry. (B) Comparison of PD-1 expression between memory Tregs and Teff co-cultured with OSCC or control secretome. (C) Comparison of TIGIT expression between memory Tregs and Teff co-cultured with OSCC or control secretome. (D) Comparison of secreted Th cytokines from Tregs and Teff co-cultured with OSCC or control secretome. Data are presented as mean ± SEM using bars with scatter dot plots for phenotype and scatter dot plots for cytokine secretion (Unpaired t test). For all statistical tests, ^∗∗∗∗p < 0.0001, ^∗∗∗p < 0.001, ^∗∗p < 0.01 and ^∗p < 0.05 were considered significant. ns, not significant.