(A) MDMs (n=4) were transfected with scrambled, Bmi1 or c-Maf siRNA for 24h. Fold mRNA expression normalized to scrambled siRNA-transfected MDMs + SEM. (B) MDMs (n=4) were left untreated (for acute) or pre-treated with 100μg/ml MDP for 24h, then transfected with scrambled, c-Maf or Bmi1 siRNA, alone or in combination, and 24h later (total 48h MDP pre-treatment), MDMs were stimulated with 100μg/ml MDP for 24h (acute) and cytokine secretion was assessed + SEM. Numbers on the bars are the ratios of cytokine secretion upon MDP treatment of the corresponding pre-treated versus non-pretreated MDMs. Statistical significance above the knockdown sample bars is compared to its corresponding scrambled siRNA sample. (C) MDMs were left untreated (for acute) or pre-treated with 100μg/ml MDP for 24h, then transfected with scrambled, or a combination of Twist1 and Twist2 siRNA ± vectors expressing c-Maf or Bmi1 (or empty vector control), and 24h later (total 48h MDP pre-treatment), MDMs were treated with 100μg/ml MDP for an additional 24h (acute) and supernatants were examined for cytokines. Mean concentration + SEM for n=4. Numbers on the bars are the ratios of cytokine secretion upon MDP treatment of pre-treated versus non-pretreated MDMs. *, p<0.01; **, p<0.01; ***, p<0.001; †, p<1×10−4; ††, p<1×10−5. Tx, treatment; scr, scrambled.