Prolonged alcohol exposure augments TLR8 or TLR4 induced TNF alpha expression. Human monocytes (n = 6 or 7) were either treated or not with 25 mM ethanol for 7 days, 2.5 ug/ml CL075 or 0.1 ug/ml LPS for 6 or 24 hours and alcohol treated cells were further stimulated or not with CL075 or LPS for 6 hours (A and C) or 24 hours (B). The cell-free supernatants were collected and analyzed for TNF alpha production by ELISA (A and B, n = 6). RNA was extracted from the cells and real time PCR was performed to quantify TNF alpha gene expression (C, n = 5). Error bars represent ± SEM. unst: unstimulated.