Mediators of Inflammation / 2011 / Article / Fig 7

Research Article

Dual Specificity Phosphatase 1 Regulates Human Inducible Nitric Oxide Synthase Expression by p38 MAP Kinase

Figure 7

The effect of DUSP1 siRNA on iNOS mRNA and protein expression and NO production in A549 and J774 cells. A549 cells were transfected with DUSP1-specific siRNA 1 or 2 (DUSP1 siRNA 1 or 2), nontargeting control siRNA (control siRNA) or Lamin A/C-specific siRNA (Lamin A/C siRNA), and untransfected cells were used as controls. (a) Cells were stimulated with the cytokine mixture (CM: TNF, IFNγ, and IL-1β; 10 ng/mL each) for 6 h, and iNOS mRNA was determined with quantitative real-time PCR and normalized against GAPDH mRNA. (b) Cells were stimulated with CM for 24 h, and iNOS protein expression was detected by Western blot and NO production as nitrite by Griess reaction. (c) J774 cells were transfected either with DUSP1-specific siRNA (siDUSP1) or nontargeting siRNA (siNEG). Cells were stimulated with LPS (10 ng/mL) for 24 h, and iNOS protein expression was detected by Western blot. In Western blots, chemiluminescent signal was quantified and iNOS expression was normalized against actin. Results are expressed as related to cells treated with CM or LPS. One-way ANOVA with Bonferroni’s posttest was performed, and statistical significance is indicated with ; and , .