Original article Regulation by intracellular glutathione of TNF-a-induced p38 MAP kinase activation and RANTES production by human pulmonary vascular endothelial cells Background: We have previously shown that p38 mitogen-activated protein (MAP) kinase regulates tumor necrosis factor-alpha (TNF-a)-induced RANTES production by human pulmonary vascular endothelial cells, and that sensitivity to TNF-a is inversely correlated with cellular reduction and oxidation (redox) state. However, a regulatory role of intracellular glutathione (GSH) in TNF-a- induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production has not been determined. In the present study, therefore, we extended our previous studies and focused on redox regulation on p38 MAP kinase activation. Methods: Human pulmonary vascular endothelial cells were exposed to N- acetylcysteine (NAC) or buthionine sulfoximine (BSO), and then TNF-a- induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production were determined. Results: The results showed that 1) NAC attenuated TNF-a-induced p38MAP kinase activation and RANTES production 2) SB 203580 as the speci®c inhibitor of p38 MAP kinase activity attenuated TNF-a-induced RANTES production 3) BSO facilitated TNF-a-induced p38 MAP kinase activation and RANTES production 4) SB 203580 attenuated BSO-mediated facilitation of TNF-a-induced RANTES production. Conclusions: These results indicated that TNF-a-induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production by human pulmonary vascular endothelial cells are inversely regulated by intracellular GSH levels. S. Hashimoto, Y. Gon, K. Matsumoto, I. Takeshita, Y. Asai, Y. Asai, T. Machino, T. Horie First Department of Internal Medicine, Nihon University School of Medicine, Tokyo, Japan Key words: GSH; p38 MAP kinase; pulmonary vascular endothelium; RANTES. Dr Shu Hashimoto First Department of Internal Medicine Nihon University School of Medicine 30-1 Oyaguchikamimachi Itabashi-ku Tokyo 173-8610 Japan Accepted for publication 20 December 1999 Introduction In the allergic in¯ammatory responses of asthmatic airways, a variety of cells and their products have the potential to contribute to the feature of allergic in¯ammation (1, 2). Of these cells, eosinophils and their products have been demonstrated to play a pivotal role in the production of allergic in¯ammation (3). The extravasation and accumulation of eosinophils at the sites of allergic in¯ammation depend upon adhesion to and migration through endothelial linings. A variety of cytokines and in¯ammatory mediators which display a chemotactic activity for eosinophils participate in this process (4±6). Among these, RANTES, a member of the C-C chemokine family, has been shown to play a role in the production of allergic in¯ammation through its chemotactic activity (4±6). Vascular endothelial cells have been shown to express RANTES (7, 8); therefore, RANTES produced by pulmonary vascular endothelial cells plays a role in the recruitment of eosinophils into the sites of allergic in¯ammation. We have previously shown that p38 mitogen- activated protein (MAP) kinase regulates tumor necrosis factor-alpha (TNF-a)-induced RANTES pro- duction by human pulmonary vascular endothelial cells (9). Recent evidence has suggested that p38 MAP kinase activation is regulated by cellular reduction/ oxidation (redox) state (10, 11); however, the role of cellular redox as regulated by intracellular glutathione (GSH) in TNF-a-induced p38 MAP kinase activation and p38 MAP kinase-mediated RANTES production Allergy 2000: 55: 463±469 Printed in UK. All rights reserved Copyright # Munksgaard 2000 ALLERGY ISSN 0105-4538 463