SEA induces IL-10 via a mechanism that is dependent on the Jak3/Stat3 pathway in malignant but not benign T cells. (A-B) Malignant (SeAx) and benign (MF1850) T cells were mono- and cocultured in the presence of SEA (50 ng/mL) together with vehicle (dimethylsulfoxide), (A) a pan Jak inhibitor (JAKI, 1 µM) or (B) a Jak3 inhibitor (Tofacitinib, 0.3 µM). After 24 hours of culture, the concentrations of IL-10 in the cell culture supernatants were determined by ELISA. Error bars represent SEM of 3 independent experiments. (C) Representative western blot of Jak3 and Erk1/2 expression after transient transfection of malignant (SeAx) and benign (MF1850) T cells with Jak3 or nontargeting (NT) siRNA. (D) Malignant (SeAx) and benign (MF1850) T cells were transiently transfected with NT or Jak3-specific siRNA and monocultured for 24 hours. Then, the transfected cells were washed and cocultured in the presence of SEA (50 ng/mL) for another 24 hours before the concentrations of IL-10 in the cell culture supernatants were determined by ELISA. Shown is the percent IL-10 expression relative to cocultures of malignant and benign cells transfected with NT siRNA. Error bars represent SEM of 3 independent experiments. (E) Representative western blot of Stat3 and Erk1/2 expression after transient transfection of malignant (SeAx) and benign (MF1850) T cells with Stat3 or NT siRNA. (F) Malignant (SeAx) and benign (MF1850) T cells were transiently transfected with NT or Stat3-specific siRNA and monocultured for 24 hours. Then, the transfected cells were washed and cocultured in the presence of SEA (50 ng/mL) for another 24 hours before the concentrations of IL-10 in the cell culture supernatants were determined by ELISA. Shown is the percent IL-10 expression relative to cocultures of malignant and benign T cells transfected with NT siRNA. Error bars represent SEM of 3 independent experiments. DMSO, dimethylsulfoxide.