STAT3 and MEK1/2-ERK1/2 are activated in response to IL-6 in CD45⁻ myeloma cell lines.

(A,B) Activation of STAT3 in myeloma cell lines stimulated with IL-6 for 0, 10, 30, or 90 minutes was determined by Western blot analysis (WB) using antibodies specific for phosphorylated STAT3 at the tyrosine residue 705 (STAT3-P). The activation of the Ras-ERK pathway was assessed by ERK1/2 kinase assay (KA) using Elk-1 as a substrate (A) and by Western blot analysis using the specific antibodies for the phosphorylated MEK1/2 at the serine residues 217 and 221 (MEK1/2-P) (B) for U266 cells (A) and for other myeloma cell lines (B), respectively. (C) Activation of SAPK/JNK and p38 MAPK was analyzed using phosphorylated SAPK/JNK at threonine 183 and tyrosine 185 (SAPK/JNK-P) and phosphorylated p38 MAPK at threonine 180 and tyrosine 182 (p38 MAPK-P) antibodies, respectively. (D) Proliferation of CD45⁺ U266 cells assessed by BrdU incorporation was suppressed by a MEK1 inhibitor, PD98059 (20 μM). Values of BrdU incorporation are indicated by means and SDs that were obtained from 3 independent experiments using triplicate samples.