BIRB 796 and dasatinib suppress the production of TNFαand SEAP transcripts. T-shNT and T-shFC were maintained in medium alone (control), or medium treated with R848 (30μM) for 24 hours after 6 hours of pretreatment with either BIRB 796 (500nM) or dasatinib (500nM). Total RNA was prepared and TNFα (A) and SEAP (B) mRNAs quantified using real-time qRT-PCR. Data are expressed as the mean ± SD -fold change relative to transcripts in unexposed (control) cells and based on 3 technical replicates normalized to 18S rRNA. Dasatinib suppressed TNFα mRNA levels to a greater degree than did BIRB 796. (C-D) Neither BIRB 796 nor dasatinib enhanced TNFα mRNA degradation. T-shFC cells were either cultured in medium alone or treated with BIRB 796 (500nM; C) or dasatinib (500nM; D) for 6 hours, after which time all cells were stimulated with R848 (30μM) for 24 hours. After adding actinomycin D (5 μg/mL), total RNA was collected at 0, 60, and 120 minutes and TNFα mRNA was measured by real-time qRT-PCR. For each drug, 1 representative of 2 experiments with identical results is shown.