HSP70 inhibitors upregulate prostaglandin E(1)-induced synthesis of interleukin-6 in osteoblasts

Interleukin-6 (IL-6) is a pro-inflammatory and bone-resorptive cytokine that also regulates bone formation. We previously showed that prostaglandin E(1) (PGE(1)) induces the synthesis of IL-6 by activating p44/p42 mitogen-activated protein kinase (MAPK), stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), and p38 MAPK in osteoblast-like MC3T3-E1 cells. In the present study, we investigated whether heat shock protein 70 (HSP70), a molecular chaperone that coordinates protein folding and homeostasis, affects PGE(1)-stimulated IL-6 synthesis in MC3T3-E1 cells through the MAPK activation. The osteoblast-like MC3T3-E1 cells were treated with HSP70 inhibitors—VER-155008 and YM-08—, PD98059, SB203580 or SP600125 and then stimulated with PGE(1). IL-6 synthesis was evaluated using an IL-6 enzyme-linked immunosorbent assay kit. IL-6 mRNA expression was measured by real-time RT-PCR. The phosphorylation of p38 MAPK was evaluated by Western blotting. We found that VER-155008, an HSP70 inhibitor, enhanced the PGE(1)-stimulated IL-6 release and IL-6 mRNA expression. YM-08, another HSP70 inhibitor, also enhanced PGE(1)-stimulated IL-6 release. PD98059, a p44/p42 MAPK inhibitor, and SP600125, a SAPK/JNK inhibitor, upregulated PGE(1)-stimulated IL-6 release. On the other hand, SB203580, a p38 MAPK inhibitor, suppressed PGE(1)-stimulated IL-6 release. YM-08 stimulated the PGE(1)-induced phosphorylation of p38 MAPK. SB203580 suppressed the amplification by YM-08 of the PGE(1)-stimulated IL-6 release. Our results suggest that HSP70 inhibitors upregulate the PGE(1)-stimulated IL-6 synthesis through p38 MAPK in osteoblasts and therefore affect bone remodeling.