N‐acetylcysteine attenuates TNF‐α‐induced p38 MAP kinase activation and p38 MAP kinase‐mediated IL‐8 production by human pulmonary vascular endothelial cells

We have previously shown that tumour necrosis factor‐α (TNF‐α) activates p38 mitogen‐activated protein (MAP) kinase to produce interleukin‐8 (IL‐8) by human pulmonary vascular endothelial cells. Reactive oxygen species (ROS) including H 2 O 2 generated by TNF‐α can act as signalling intermediates for cytokine induction; therefore, scavenging ROS by anti‐oxidants is important for the regulation of cytokine production. However, the effect of N‐acetylcysteine (NAC), which acts as a precursor of glutathione (GSH) synthesis, on TNF‐α‐induced activation of p38 MAP kinase pathway and p38 MAP kinase‐mediated IL‐8 production by human pulmonary vascular endothelial cells has not been determined. To clarify these issues, we examined the effect of NAC on TNF‐α‐induced activation of p38 MAP kinase, MAP kinase kinase (MKK) 3 and MKK6 which are upstream regulators of p38 MAP kinase, and p38 MAP kinase‐mediated IL‐8 production. Human pulmonary vascular endothelial cells that had been preincubated with NAC were stimulated with TNF‐α and then the activation of p38 MAP kinase and MKK3/MKK6 in the cells and IL‐8 concentrations in the culture supernatants were determined. Intracellular GSH levels increased in NAC‐treated cells. NAC attenuated TNF‐α‐induced activation of p38 MAP kinase and MKK3/MKK6. NAC attenuated p38 MAP kinase‐mediated IL‐8 production by TNF‐α‐stimulated cells. These results indicate that the cellular reduction and oxidation (redox) regulated by intracellular GSH is critical for TNF‐α‐induced activation of p38 MAP kinase pathway and p38 MAP kinase‐mediated IL‐8 production by human pulmonary vascular endothelial cells, and we emphasize that anti‐oxidant therapy is an important strategy for the treatment of acute lung injury.British Journal of Pharmacology (2001) 132 , 270–276; doi: 10.1038/sj.bjp.0703787