TAK1-MAPK-NFkappaB Signaling Pathways Regulates Induction of TNF-alpha by Wear Particles In Vitro

摘要

Purpose: Wear particles may induce inflammatory osteolysis by activating the nuclear factor kB (NFkB) and mitogen activated protein kinases (MAPKs). Previous studies have shown that transforming growth factor-beta activated kinase 1 (TAK1) functions as an adaptor molecule in the interaction between tumor necrosis factor receptor-associated factor 6(TRAF6) and downstream molecules such as NF-kappaB, p38MAPK signaling cascades. This study investigated the roles of TAK1 pathway in TNF-alpha production after exposure to titanium particles. Methods: RAW 264.7 murine macrophages were incubated with endotoxin-free titanium particles in the presence and absence of TAK1 inhibitors 5Z-7-oxozeaenol. TAK1 activation was assessed with use of Western blot. p38MAPK and NFkB activation was evaluated by detecting endogenous levels of Phospho-p38alpha MAPK and Phospho-NFkappaBp65 protein using enzymelinked immunosorbent assay (ELISA). TNF-alpha mRNA and protein in cell culture medium was qualified using quantitative real time reverse transcriptase polymerase chain reaction and ELISA respectively. Results: Endogenous TAK1 was phosphorylated upon simulation of titanium particles in RAW 264.7 cells. We also showed that 5Z-7-oxozeaenol decreased the amount of phospho-p38alpha and phospho-NFkappaBp65 which were activated by LPS and the titanium particles. Furthermore, we demonstrated that TNF-alpha was completely blocked by the TAK1 inhibitor in RAW 264.7 cells. Conclusions: In the present study, we conclude that TAK1- MAPK-NFkappaB signaling pathways are involved in the induction of TNF-alpha in RAW264.7 cells exposed to titanium particles. These results suggest that TAKI-MAPK-NFkappaB signaling pathway may be a potential pharmacological target to modulate wear particle- induced inflammation.