Signal transduction induced by BAFF or APRIL in myeloma cells. (A) XG-13, XG-14, and RPMI8226 cells were starved overnight and cultured without cytokine, or with either IL-6 (30 ng/mL), BAFF (800 ng/mL), or APRIL (800 ng/mL) for 10 and 30 minutes at 37°C. Cell lysates were analyzed by Western blotting with antisera against phospho-STAT3 (pSTAT3), phospho-ERK1/2 (pMAPK), and phospho-AKT (pAKT). Immunoblotting for STAT3, MAPK, and AKT confirmed equal protein loading. Western blots are of one representative experiment of 3. (B) XG-13, XG-14, and RPMI8226 cells were starved overnight and cultured without cytokine, or with either IL-6 (3 ng/mL), TNF-α (20 ng/mL), BAFF (200 ng/mL), or APRIL (200 ng/mL) for 30 minutes at 37°C. NF-κB activity was detected by ELISA according to the manufacturer's instructions. (C) XG-13 cells were IL-6 starved for 3 hours and cultured without cytokine or in the presence of BAFF (200 ng/mL) or APRIL (200 ng/mL). When indicated, Ly 294002 (25 μM), SN50 (100 μg/mL), or the SN50 inactive peptide (100 μg/mL) was added. Results are the mean values ± SD of the tritiated thymidine incorporation determined on sextuplet culture wells and are expressed as the percentage of the proliferation obtained with APRIL and SN50 inactive peptide. ^*Mean value is statistically significantly different from that obtained with BAFF or APRIL using a Student t test (P ≤ .05).