FIGURE 3.

STAT1 silencing impairs RET/PTC-induced IRF1 and IDO1 overexpression. PTC3–5 cells were transfected with rat STAT1 siRNA (25 nm) (siSTAT1) or with non-target siRNA (25 nm) (siNC). Twenty-four hours after transfection, cells were stimulated with Dox (1 μg/ml) for 48 h before cell harvesting for total RNA and cell lysate preparation. After total RNA extraction and cDNA synthesis, STAT1 (A), IRF1 (B), and IDO1 (C) mRNA expression levels were evaluated by quantitative PCR. STAT1, IRF1, and IDO1 mRNA expression resulted significantly in impaired STAT1-silenced cells compared with controls. The quantitative PCR data are presented as means of arbitrary units. D, immunoblottings for RET, IDO1, total STAT1, and tubulin were performed. In STAT1-silenced cells, despite a comparable expression of RET/PTC3, STAT1 and IDO1 protein expression was significantly impaired compared with controls. All experiments were repeated twice and the more representative results are depicted. p values were calculated applying the two-tailed unpaired Student's t test.