Fig 1: TNFa does not affect zafirlukast-induced cell death. (A) mRNA expression levels of CysLTR1, TNFR1 and TNFa in the indicated cell lines were determined using qRT-PCR. Data are normalized to GAPDH expression and presented as x-fold mRNA expression compared to control. Mean and SD of four independent experiments performed in triplicate are shown. * p < 0.05, ** p < 0.01, *** p < 0.001; (B) Western blot analysis of CysLTR1 and TNFR1 expression in 293T, 786-O, 786-O-EV and 786-O-VHL cells. Vinculin serves as loading control. Representative blots of at least three different independent experiments are shown; (C) 786-O and 293T cells were pre-treated with 100 µg/mL of Etanercept (Enbrel) for 1 h, followed by treatment with DMSO or 100 µM zafirlukast for 24 h, after which cell death was determined by quantifying PI uptake. Data are presented as percentage of PI-positive cells and mean and SD of three independent experiments performed in triplicate are shown. *** p < 0.001, n.s.: non-significant.