Figure 2

Cordycepin efficiently reduces VCAM-1 expression in response to LPS, but not TNF-α, in A549 cells, and such effects are associated with a reduction in p65 NF-κB phosphorylation but not its nuclear translocation. (A) A549 cells were treated for 6 h with 1 μg/mL LPS or 10 ng/mL TNF-α in the absence or presence of the indicated concentrations of cordycepin. Total RNA was subjected to RT-PCR analysis with primers to VCAM-1, ICAM-1 or β-actin. (B) A549 cells were treated with TNF-α for 12 h in the absence or presence of the indicated concentrations of cordycepin. Protein extracts were prepared and subjected to immunoblot analysis with antibodies to VCAM-1, ICAM-1 or GAPDH. (C) A549 cells were treated with TNF-α for 5 min in the absence or presence of the indicated concentrations of cordycepin. Protein extracts were prepared and subjected to immunoblot analysis with antibodies to p65 NF-κB, its serine-536 phosphorylated form, p50 NF-κB or GAPDH. (D) A549 cells were treated with TNF-α for 30 min in the absence or presence of the indicated concentrations of cordycepin. Cells were then fixed and subjected to immunofluorescence with antibodies to p65 or p50 NF-κB. Note that all cells were pretreated with the indicated drug 30 min before treatment with either LPS or TNF-α in the continued presence of the drug.