FIGURE 3.

Effects of STAT-6 on 15-LOX-1 transcriptional activation. A and B, effects of STAT-6 siRNA transfection on STAT-6 mRNA expression. Caco-2 (A) and SW480 (B) cells were transfected with a pool of four siRNA duplexes against STAT-6 (STAT-6 siRNA), a nonspecific siRNA sequence (Nonspecific siRNA), or transfection medium only (Mock) for 48 h and then treated with 5 nm depsipeptide for 24 h. STAT-6 mRNA levels were measured with real-time reverse transcription-PCR. Expression levels were calculated as the values relative to those of the calibrator samples (Nonspecific siRNA). The values are the means ± S.D. of triplicate experiments. C and D, effects of STAT-6 siRNA transfection on STAT-6 binding to the 15-LOX-1 promoter. Caco-2 (C) and SW480 (D) cells were transfected and treated as described in A and B. The cells were subjected to chromatin immunoprecipitation assays using a specific anti-STAT-6 antibody. STAT-6 antibody + and – indicate immunoprecipitation with and without STAT-6 antibody, respectively, and Input indicates total DNA before immunoprecipitation. E and F, effects of STAT-6 down-regulation by siRNAs on depsipeptide induction of 15-LOX-1 expression. Caco-2 (E) and SW480 (F) cells were transfected and treated as described in A and B. 15-LOX-1 mRNA relative expression levels, real-time reverse transcription-PCR, were calculated as the values relative to those of the calibrator samples (Mock). The values are the means ± S.D. of triplicate experiments.