Inhibition of SAPK/JNK leads to enhanced IL-1-induced IL-6 synthesis in osteoblasts

Stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK)1 iations used: Stress-activated protein kinase/c-Jun N-terminal kinase, SAPK/JNK; Interleukin 1, IL-1; Interleukin 6, IL-6; AMP-activated protein kinase, AMPK; Mitogen-activated protein, MAP; Inhibitor of κB, IκB; Nuclear factor-κB, NF-κB; Vascular endothelial growth factor, VEGF; Enzyme-linked immunosorbent assay, ELISA; Normal Human Osteoblasts, NHOst; Glyceraldehyde-3-phosphate dehydrogenase, GAPDH; α-minimum essential medium, α-MEM; Fetal calf serum, FCS; Sodium dodecyl sulfate, SDS; Polyacrylamide gel electrophoresis, PAGE. h belongs to the MAP kinase superfamily regulates many cellular events. We previously reported that interleukin 1 (IL-1) stimulates the synthesis of interleukin 6 (IL-6) through activation of ERK and p38 MAP kinase in osteoblast-like MC3T3-E1 cells, and that AMP-activated protein kinase (AMPK) negatively regulates the IL-1-induced IL-6 synthesis through IκB/NF-κB pathway. In the present study, we investigated the role of SAPK/JNK in the IL-1-stimulated IL-6 synthesis in these cells. IL-1 induced the phosphorylation of SAPK/JNK. SP600125, an inhibitor of SAPK/JNK, increased the release and the mRNA expression levels of IL-6 induced by IL-1. IL-1-stimulated IL-6 release was significantly up-regulated in SAPK/JNK-knocked down cells. SP600125 remarkably suppressed the IL-1-induced phosphorylation of both IκB and NF-κB, whereas SP600125 failed to affect the IL-1-induced phosphorylation of AMPK, STAT3 or Src. Compound C, an AMPK inhibitor, attenuated the IL-1-induced phosphorylation of SAPK/JNK. SP600125 enhanced IL-1-stimulated IL-6 release also in normal human osteoblasts. These results strongly suggest that SAPK/JNK negatively regulates IL-1-stimulated IL-6 synthesis and acts at the point between AMPK and IκB/NF-κB in osteoblasts.