HSP70 inhibitor amplifies the bFGF­induced release of IL­6 in osteoblasts.

Heat shock protein 70 (HSP70) functions as an ATP­dependent molecular chaperone under stress and is involved in protein homeostasis, folding and degradation. HSP70 inhibitors amplify TGF­ß­stimulated VEGF synthesis in the mouse osteoblastic MC3T3­E1 cell line. Basic fibroblast growth factor (bFGF) stimulates IL­6 release via p38 MAPK in MC3T3­E1 osteoblast­like cells. In the present study, the effects of HSP70 on the bFGF­stimulated release of IL­6 was evaluated using MC3T3­E1 osteoblast­like cells. IL­6 release and mRNA expression levels were analyzed using ELISA and reverse transcription­quantitative PCR, respectively. Phosphorylation of p38 MAPK and HSP70 was assessed using western blotting. HSP70 inhibitor VER­155008 significantly increased the bFGF­stimulated release of IL­6 in both MC3T3­E1 osteoblastic cells and normal human osteoblasts. Furthermore, VER­155008 significantly enhanced the mRNA expression levels of IL­6 stimulated by bFGF. Western blotting demonstrated a significant increase in the bFGF­stimulated phosphorylation of p38 MAPK in VER­155008­treated MC3T3­E1 cells. A significant increase in the bFGF­stimulated phosphorylation of p38 MAPK was also demonstrated in MC3T3­E1 cells treated with YM­08, another HSP70 inhibitor. VER­155008 or YM­08 did not significantly affect the expression of HSP70 with or without bFGF stimulation. Finally, the specific p38 MAPK inhibitor SB203580 markedly suppressed the enhancing effects of VER­155008 on bFGF­stimulated release of IL­6. Taken together, these results indicated that HSP70 inhibitor amplified bFGF­stimulated release of IL­6 through p38 MAPK activation in the osteoblastic MC3T3­E1 cell line.