Posttranscriptional regulation of macrophage tissue factor expression by antioxidants

Tissue factor (TF) expression by cells of monocyte/macrophage lineage represents an important mechanism underlying the initiation of fibrin deposition at sites of extravascular inflammation. Recent evidence suggests a role for oxidant stress in the signalling pathway of various cell types by virtue of its ability to induce DNA binding of various transcription factors, including nuclear factor kappa B and AP-1. The effect of antioxidant treatment on lipopolysaccharide (LPS)-induced TF expression was examined in murine peritoneal macrophages and human monocytes. Both pyrrolidine dithiocarbamate, an oxidant scavenger, and N-acetyl-cysteine, a precursor of the endogenous antioxidant glutathione, inhibited stimulation of macrophage procoagulant activity by LPS. Northern blot analysis showed that neither of these agents reduced LPS-stimulated TF mRNA accumulation, thereby suggesting a posttranscriptional mechanism for the effect. Immunofluorescence studies of human monocytes using polyclonal anti-TF antibody showed that N-acetyl-cysteine treatment prevented the characteristic plasmalemmal localization of TF antigen that occurs in response to LPS. Western blot analysis showed that N-acetyl-cysteine reduced the accumulation of the 47-kD mature glycoprotein in LPS-treated cells, a finding consistent with the results of the immunofluorescence studies. Furthermore, these conditions did not result in an accumulation of the less mature forms of TF. When considered together, these data suggest that antioxidants exert their effects by impairing translation and/or by causing degradation of newly translated protein. The effect of antioxidants on tumor necrosis factor appeared to be species specific, with no effect on LPS-induced tumor necrosis factor in murine cells, but with inhibition in human monocytes. The posttranscriptional effect of antioxidants on TF expression data suggests a novel mechanism whereby these agents might modulate monocyte/macrophage activation.