Figure 4.

NF-κB signaling pathway might be involved in celastrol-suppressed chemokine production. (A) Total cell lysates of RA–FLS1 and 2 cells treated with or without celastrol (1 μM) were examined by Western blotting. (B) Subcellular fractionation was performed as described in the Materials and Methods Section; nuclear and cytoplasmic lysates of RA–FLSs with or without celastrol treatment were examined by Western blotting. (C) After RA–FLS1 and 2 cells were treated with celastrol (1 μM) or celastrol (1 μM) together with pretreatment of two NF-κB activators (PMA and TNF-α), real-time PCR analysis was carried out to examine the mRNA expression of CCL2 and CXCL12. Data bars represent the mean value ± SD from triplicate wells from three separate experiments using RA–FLS1 and 2 cells. * p < 0.05 versus RA-FLS1/2 treated with celastrol.