Expression of a dominant-negative mutant of p21(ras) inhibits induction of nitric oxide synthase and activation of nuclear factor-κB in primary astrocytes

The present study underlines the importance of p21(ras) in regulating the inducible nitric oxide synthase (iNOS) in primary astrocytes. Bacterial lipopolysaccharides induced the GTP loading of p21(ras), and the expression of a dominant-negative mutant of p21(ras) (Δp21(ras)) inhibited lipopolysaccharide-induced GTP loading in rat primary astrocytes. To delineate the role of p21(ras) in the induction of iNOS, we examined the effect of Δp21(ras) on the expression of iNOS and the production of nitric oxide. It is interesting that expression of Δp21(ras) markedly inhibited the production of nitric oxide and the expression of iNOS in lipopolysaccharide- and proinflammatory cytokine (tumor necrosis factor-α, interleukin-1β; interferon-γ)-stimulated rat and human primary astrocytes. Inhibition of iNOS promoter-derived chloramphenicol acetyltransferase activity by Δp21(ras) suggests that p21(ras) is involved in the transcription of iNOS. As activation of nuclear factor-κB (NF-κB) is necessary for the transcription of iNOS, we examined the effect of Δp21(ras) on the activation of NF-κB. Expression of Δp21(ras) inhibited the DNA binding as well as the transcriptional activity of NF-κB in activated astrocytes, suggesting that Δp21(ras) inhibits the expression of iNOS by inhibiting the activation of NF-κB. These studies also suggest that inhibitors of p21(ras) may be used as therapeutics in nitric oxide- and cytokine-mediated neuroinflammatory diseases.