Activation of Adenosine A3 Receptor Alleviates TNF-α-Induced Inflammation through Inhibition of the NF-κB Signaling Pathway in Human Colonic Epithelial Cells
Figure 4
Effect of 2-Cl-IB-MECA on TNF-α-induced NF-κB p65 activation in HT-29 cells. HT29 cells were treated with various concentrations (10 nM, 30 nM, and 50 nM) of 2-Cl-IB-MECA for 30 min and then incubated with TNF-α (10 ng/mL) for 30 min. After that, cytosolic and nuclear proteins were extracted and analyzed by western blot. (a, b) TNF-α resulted in a reduction of NF-κB p65 in cytosolic extracts and an increase of NF-κB p65 in nuclear extracts from HT-29 cells, demonstrating TNF-α-induced NF-κB p65 translocation from the cytoplasm to the nucleus. (c, d) Compared with the TNF-α-only-treated group, 2-Cl-IB-MECA pretreatment reversed the effects of TNF-α, leading to a significant increase in NF-κB p65 in the cytoplasm and a decrease in NF-κB p65 in the nucleus, both in a concentration-dependent manner. Data are shown as the mean ± SD from three independent experiments. (e–h) Representative pictures of NF-κB p65 protein expression in indicated groups detected by western blot. and compared with the NC group; compared with the TNF-α-only-treated group; between indicated groups.
