Transcriptional regulation of MMP-9 expression in stromal cells of human giant cell tumor of bone by tumor necrosis factor-alpha

We determined whether certain factor(s) secreted by multinucleated giant ce lls, which is of monocyte/ macrophage lineage in giant cell tumor of bone ( GCT), regulate the induction of matrix metalloproteinase (MMP)-9 expression in mononucleated stromal cells. Our data derived using enzyme linked immun osorbant assays (ELISAs) suggest that the GCT cells in primary culture prod uce both MMP-9 and tumor necrosis factor-alpha (TNF-alpha). Further, the MM P-9 expression in GCT primary cultures was partially abrogated by neutraliz ing antibody to TNF-alpha, suggesting that TNF-alpha secretion by the multi nucleated giant cells may be one of the factors responsible for the product ion of MMP-9 by the stromal cells in vivo. In order to confirm this we exam ined the role of TNF-alpha on the induction of MMP-9 expression in bone GCT stromal cells. These cells express MMP-2, but not MMP-9. However, treatmen t of these cells with TNF-alpha induced the expression of MMP-9 in a concen tration-dependent manner. Kinetic experiments revealed that the secretion o f MMP-9 peaked 12 h post TNF-alpha stimulation. Immunofluorescence studies confirmed the expression of MMP-9 after stimulation of GCT stromal cells wi th TNF-alpha. Further, TNF-alpha-induced MMP-9 expression was completely bl ocked with neutralizing antibody to TNF-alpha, thereby demonstrating the sp ecificity. In addition, the induction of MMP-9 expression by TNF-alpha was completely abrogated in the presence of cycloheximide, a protein synthesis inhibitor, suggesting that de novo protein synthesis may be required. Nucle ar run-on analysis demonstrated that treatment of GCT stromal cells signifi cantly enhanced the MMP-9 gene transcription. Together, our data suggest th at TNF-alpha secreted by the multinucleated giant cells up-regulates MMP-9 expression in GCT stromal cells by the induction of certain transcription f actors, which in turn enhanced the rate of transcription of MMP-9 gene. The se studies also suggest the existence of an essential cell-cell interaction in the regulation of MMP-9 expression in GCT.