Fibronectin-bound TNF-α stimulates monocyte matrix metalloproteinase-9 expression and regulates chemotaxis

Abstract

Abstract Tumor necrosis factor α (TNF-α) is a proinflammatory cytokine implicated in the stimulation of matrix metalloproteinase (MMP) production by several cell types. Our previous studies demonstrated that TNF-α avidly binds fibronectin (FN) and laminin, major adhesive glycoproteins of extracellular matrix (ECM) and basement membranes. These findings suggested that TNF-α complexing to insoluble ECM components may serve to concentrate its activities to distinct inflamed sites. Herein, we explored the bioactivity and possible function of ECM-bound TNF-α by examining its effects on MMP-9 secretion by monocytes. Immunofluorescent staining indicated that LPS-activated monocytes deposited newly synthesized TNF-α into ECM-FN. FN-bound TNF-α (FN/TNF-α) significantly up-regulated MMP-9 expression and secretion by the human monocytic cell line MonoMac-6 and peripheral blood monocytes. Such secretion could be inhibited by antibodies that block TNF-α activity and binding to its receptors TNF RI (p55) and TNF RII (p75). Chemotaxis through ECM gels in the presence of soluble or bound TNF-α was inhibited by a hydroxamic acid inhibitor of MMPs (GM6001). It is interesting that, although the adhesion of MonoMac-6 cells to FN/TNF-α required functional activated β1 integrins, FN/TNF-α-induced MMP-9 secretion was independent of binding to β1 integrins, since MMP-9 secretion was unaffected by: (1) neutralizing mAb to α4, α5, and β1 subunits, which blocked cell adhesion; (2) a mAb that stimulated β1 integrin-mediated adhesion; and (3) binding TNF-α to the 30-kDa amino-terminal fragment of FN, which lacks the major cell adhesive binding sites. Thus, in addition to their cell-adhesive roles, ECM glycoproteins, such as FN, may play a pivotal role in presenting proinflammatory cytokines to leukocytes within the actual inflamed tissue, thereby affecting their capacities to secrete ECM-degrading enzymes. These TNF-α-ECM interactions may serve to limit the cytokine’s availability and bioactivity to target areas of inflammation.