Silica induces nuclear factor-κB activation through tyrosine phosphorylation of IκB-α in RAW264.7 macrophages

It was previously reported that protein tyrosine kinase (PTK) but not protein kinase C or A plays an important role in silica-induced activation of NF-kappaB in macrophages. The question is raised whether PTK stimulation and NF-kappaB activation in silica-stimulated macrophages are directly connected through tyrosine phosphorylation of I kappaB-alpha. Results indicate that stimulation of macrophages with silica led to NF-kappaB activation through tyrosine phosphorylation without serine phosphorylation. Specific inhibitors of protein tyrosine kinase, such as genistein and tyrophostin AG126, prevented tyrosine phosphorylation of I kappaB-alpha in response to silica. I kappaB-alpha protein levels remained relatively unchanged for up to 60 min after silica stimulation. Moreover, inhibition of proteasome proteolytic activity did not affect NF-kappaB activation by silica Antioxidants, such as superoxide dismutase (SOD), N-acetylcysteine (NAC), and pyrrolidine dithiocarbamate (PDTC), blocked tyrosine phosphorylation of I kappaB-alpha: induced by silica, suggesting reactive oxygen species (ROS) may be important regulatory molecules in NF-kappaB activation through tyrosine phosphorylation of I kappaB-alpha. The results suggest that tyrosine phosphorylation of I kappaB-alpha represents a proteasome proteolytic activity-independent mechanism for NF-kappaB activation that directly couples NF-kappaB to cellular tyrosine kinase in silica-stimulated macrophages. This proposed mechanism of NF-kappaB activation induced by silica could be used as a target for development of antiinflammatory and antifibrosis drugs, (C) 2000 Academic Press.