Figure 3
From: Epithelial expression of TLR4 is modulated in COPD and by steroids, salmeterol and cigarette smoke
Corticosteroids downregulate TLR4 expression in respiratory epithelial cells. A549 cells (3 × 105) were seeded onto 6-well plates and grown to confluence. Cells were washed, placed in low serum (1% FCS) medium and were left untreated or incubated with fluticasone propionate or dexamethasone over the dose ranges 10-9 to 10-6 Molar for 16 hours. A & B. Total RNA was extracted, reverse transcribed into cDNA and used as a template in PCR reactions using, TLR4 and GAPDH gene-specific primers. Products were electrophoresed in 1.5% TBE agarose gels containing 0.5 μg/ml ethidium bromide and visualised under UV. Gels are representative of three independent experiments. C. Western blot analysis of membrane extracts (10 μg) from A549 cells probed with an anti-TLR4 or anti-Actin antibody. Equal protein loading and transfer efficiency was confirmed by Ponceau S staining. Data are representative of three separate experiments.