Interferon-gamma and tumor necrosis factor-alpha synergistically enhance the immunosuppressive capacity of human umbilical-cord-derived mesenchymal stem cells by increasing PD-L1 expression

Figure 4 Tumor necrosis factor-alpha increased programmed cell death protein 1 ligand expression by promoting interferon-gamma-induced activation of the JAK/STAT1/interferon regulatory factor 1 pathway. A: Programmed cell death protein 1 ligand (PD-L1) expression was quantified by quantitative real-time PCR (qRT-PCR) in human umbilical-cord-mesenchymal stem cells (hUC-MSCs) after 4 h of incubation with IFN-γ and tumor necrosis factor-alpha (TNF-α) in the presence or absence of ruxolitinib (10 μM) or the NF-κB inhibitor BAY11-7082 (10 μM); B: PD-L1 expression in hUC-MSCs was quantified by flow cytometry after 24 h of incubation with IFN-γ and TNF-α in the presence or absence of ruxolitinib (10 μM); C: Activation of the JAK/STAT1/interferon regulatory factor 1 (IRF1) pathway and the expression of PD-L1 were analyzed by western blot in hUC-MSCs after 24 h of incubation with IFN-γ and TNF-α in the presence or absence of ruxolitinib (10 μM); D and E: PD-L1 expression was quantified by quantitative real-time PCR and flow cytometry in hUC-MSCs transfected with siRNA of NC, STAT1, or IRF1 and subsequently treated with IFN-γ and TNF-α for 4 h or 24 h; F: Activation of the JAK/STAT1/IRF1 pathway was analyzed by western blot in hUC-MSCs treated with IFN-γ (20 ng/mL), TNF-α (10 ng/mL), or a combination of IFN-γ (20 ng/mL) and TNF-α (10 ng/mL) for 24 h. Data were represented as mean ± SEM of n = 3. C: Control; Iso: Isotype; ^cP < 0.0001. IFN-γ: Interferon-gamma; PD-L1: Programmed cell death protein 1 ligand 1; TNF-α: Tumor necrosis factor-alpha.