FIGURE 5.
Transfection of an IκBα dominant-negative expression vector blocks FN-f activation of NF-κB and inhibits IL-6 and IL-8 production. Primary human articular chondrocytes from two different donors were transfected with the dominant-negative construct pCMV-IκBαM by nucleofection, as described in Materials and Methods. Cells were stimulated with 0.5 µM FN-f for 30 min to assess phosphorylation of the p65 NF-κB subunit by immunoblotting of cell lysates with an anti-phospho-p65 Ab or nonphosphospecific (total) Ab as control. Medium collected from parallel cultures incubated for 16 h with FN-f was used for immunoblotting for IL-6 and IL-8.