Hypoxia triggers responses in endothelial cells that play roles in many conditions including high-altitude
pulmonary edema and
tumor angiogenesis. Signaling pathways activated by
hypoxia modify cytoskeletal and
contractile proteins and alter the biomechanical properties of endothelial cells. Intermediate filaments are major components of the cytoskeleton whose contribution to endothelial physiology is not well understood. We have previously shown that
hypoxia-activated signaling in endothelial cells alters their contractility and adhesiveness. We have also linked p38-MAP
kinase signaling pathway leading to HSP27 phosphorylation and increased actin stress fiber formation to endothelial barrier augmentation. We now show that
vimentin, a major intermediate filament
protein in endothelial cells, is regulated by
hypoxia. Our results indicate that exposure of endothelial cells to
hypoxia causes
vimentin filament networks to initially redistribute perinuclearly. However, by 1 hour
hypoxia these networks reform and appear more continuous across cells than under normoxia.
Hypoxia also causes transient changes in
vimentin phosphorylation, and activation of PAK1, a
kinase that regulates
vimentin filament assembly. In addition, exposure to 1 hour
hypoxia increases the ratio of insoluble/soluble
vimentin. Overexpression of phosphomimicking mutant HSP27 (pmHSP27) causes changes in
vimentin distribution that are similar to those observed in hypoxic cells. Knocking-down HSP27 destroys the
vimentin filamentous network, and disrupting
vimentin filaments with acrylamide increases endothelial permeability. Both
hypoxia- and pmHSP27 overexpression-induced changes are reversed by inhibition of
phosphatase activity. In conclusion
hypoxia causes redistribution of
vimentin to a more insoluble and extensive filamentous network that could play a role in endothelial barrier stabilization.
Vimentin redistribution appears to be mediated through altering the phosphorylation of the
protein and its interaction with HSP27.