Further Insights in the Mechanisms of Interleukin-1β Stimulation of Osteoprotegerin in Osteoblast-Like Cells

The mechanisms of IL-1β stimulation of OPG were studied in more detail. Whereas p38 and ERK activation was confirmed to be needed, NF-κB was not necessary for this regulation. We also found that OPG production after IL-1β stimulation was not sufficient to block TRAIL-induced apoptosis in MG-63 cells. Introduction: Osteoprotegerin (OPG) plays a key role in the regulation of bone resorption and is stimulated by interleukin (IL)-1β. Herein, we defined the mechanisms of IL-1β stimulation of OPG focusing on the potential involvement of MAPK and NF-κB. We also examined whether OPG production in response to IL-1β influences TRAIL-induced apoptosis in MG-63 cells. Materials and Methods: OPG mRNA levels in MG-63 cells were quantified by real-time RT-PCR and protein levels of OPG and IL-6 by ELISA. Cell viability was assessed using the methyltetrazidium salt (MTS) reduction assay. The role of the MAPK pathway was studied by both Western blotting and the use of specific chemical inhibitors. NF-κB function was studied using BAY 11-7085 and by siRNA transfection to inhibit p65 synthesis. Transcription mechanisms were analyzed by transiently transfecting MG-63 cells with OPG promoter constructs. Post-transcriptional effects were examined by using cycloheximide and actinomycin D. Results: MG-63 cells treatment with IL-1β resulted in the phosphorylation of c-Jun NH2-terminal kinase (JNK), p38, and extracellular signal-regulated kinase (ERK). The use of the specific inhibitors showed that p38 and ERK but not JNK were needed for IL-1β–induced OPG production. In contrast, NF-κB was not essential for IL-1β induction of OPG. We also showed a small transcriptional and a possible post-transcriptional or translational regulation of OPG by IL-1β. Exogenous OPG blocked TRAIL-induced apoptosis, but IL-1β induction of OPG did not influence TRAIL-induced cell death. Conclusions: IL-1β stimulates OPG production by mechanisms dependent on p38 and ERK. In contrast, NF-κB was not essential for this regulation. Although the relevance of IL-1β stimulation of OPG is still not fully understood, our data showed that IL-1β stimulation of OPG does not modify TRAIL-induced cell death.