Enhancement of nuclear factor-κB activation and protein tyrosine phosphorylation by a tyrosine phosphatase inhibitor, pervanadate, involves reactive oxygen species in silica-stimulated macrophages

Reactive oxygen species (ROS) and phosphorylation events mediated by tyrosine kinase are involved in silica-induced nuclear factor-kappa B (NF-κB) activation. Protein tyrosine phosphatase (PTPase) acts to limit protein tyrosine phosphorylation. In the present study, we investigated the role of PTPase in NF-κB activation and tyrosine phosphorylation in silica-stimulated macrophages, and the involvement of ROS in these responses. Treatment of mouse peritoneal macrophages (RAW264.7 cells) with a PTPase inhibitor, pervanadate, markedly enhanced the DNA-binding activity of NF-κB in the presence or absence of silica. The stimulatory effect of pervanadate on NF-κB activation was also demonstrated in LPS-stimulated macrophages. A specific inhibitor of protein tyrosine kinase (PTK), genistein, prevented the NF-κB activation induced by pervanadate in the presence of silica while inhibitors of protein kinase A or C, such as staurosporine or H7, had no inhibitory effect on NF-κB activation. A variety of antioxidants, such as catalase, superoxide dismutase, N-acetyl cysteine (NAC), and pyrrolidine dithiocarbamate, inhibited NF-κB activation induced by pervanadate in the presence of silica. Furthermore, pervanadate markedly enhanced silica- or LPS-induced protein tyrosine phosphorylation in cells. Treatment of macrophages with NAC abolished the increase in tyrosine phosphorylation in cells stimulated with the combination of pervanadate and either silica or LPS or with silica alone. The results suggest that PTPase may play a crucial role in the negative regulation of silica-signaling pathways leading to NF-κB activation in macrophages. Furthermore, ROS appear to be involved in downstream signaling between PTPase inhibition and NF-κB activation. Copyright (C) 2000 Elsevier Science Ireland Ltd.