Hypoxia and kinase activity regulate lung epithelial cell glutathione

The authors investigated the mechanisms by which hypoxia regulates glutathione (GSH) in lung epithelial cells, and specifically whether the mitogen-activated protein kinase (MAPK) system is involved in the response to hypoxia. Hypoxia decreased cellular GSH content and appeared to decrease the effect of N-acetylcysteine on repletion of GSH after hypoxia. Hypoxia decreased 2 key enzyme activities that regulate GSH synthesis, glutamate cysteine ligase (GCL) (E. C. 6.3.2.2) and glutathione synthase (GS) (E. C. 6.3.2.3). No hypoxia-dependent change occurred in GCL or GS protein expression on Western blots. When epithelial cells were transfected with an adenoviral vector that caused over expression of human catalase protein (Ad. Cat or Ad.mCat), GCL and GS activities did not decrease in hypoxia. Inhibition of p38(MAPK) (using SB203580) or extracellular signal-regulated kinase (ERK; PD98059) prevented the hypoxia-dependent decrease in GCL and GS activity. To seek in vivo correlation, the authors assayed total glutathione in lungs and livers from MK2(-/-)(homozygous knockout) mice. MK2(-/-) mice are presumably unable to phosphorylate heat shock protein 27 (Hsp27) normally, because of absent kinase (MK2) activity. Liver GSH content (expressed per mg protein) was 20% less in MK2(-/-) mice than in nontransgenic Black 6 controls. Down-regulation of lung GSH content in hypoxia depends on peroxide tone of the cell and the p38(MAPK) system.