X-linked inhibitor of apoptosis protein is an important regulator of vascular endothelial growth factor-dependent bovine aortic endothelial cell survival.

Vascular endothelial growth factor (VEGF) is a critical regulator of endothelial cell biology and vascular function. Chronic VEGF treatment has been shown to inhibit tumor necrosis factor-induced apoptosis in endothelial cells. However, the mechanism for this cell survival is unclear. Interestingly, VEGF also enhances the expression of X-linked inhibitor of apoptosis (XIAP), a well-established antiapoptotic factor. XIAP has been shown to suppress apoptosis by blocking caspase activity in cancer cells, but it remains under studied in the endothelium. Therefore, we hypothesized that VEGF affects important endothelial functions, such as apoptosis and cell migration, by regulating XIAP expression and downstream caspase activity. To test this hypothesis, caspase activity, apoptosis, and cell migration were assessed following XIAP overexpression or depletion in bovine aortic endothelial cells. Much like VEGF treatment, ectopic expression of XIAP blocked tumor necrosis factor-induced apoptosis. Surprisingly, the mechanism was caspase-independent. In addition, XIAP-associated cell survival was the result of enhanced nitric oxide (NO) production, and XIAP was partially localized in caveolae. In these lipid rafts, XIAP interacted with a regulator of NO production, caveolin-1, via a binding motif (FtFgtwiY, where the bold letters represent aromatic amino acids) in the baculoviral IAP repeat-3 domain. Endothelial NO synthase binding to caveolin-1 was competitively inhibited by XIAP, suggesting that XIAP acts as a modulator of NO production by releasing endothelial NO synthase from caveolin-1. Further studies showed that endothelial cell migration was also controlled by XIAP-dependent NO. Taken together, these results suggest that XIAP plays a novel role in endothelial cells, interacting with caveolin-1 and acting as a regulator of vascular antiatherogenic function.

Laminar shear stress up-regulates peroxiredoxins (PRX) in endothelial cells: PRX 1 as a mechanosensitive antioxidant.

Shear stress plays a significant role in endothelial cell biology and atherosclerosis development. Previous work by our group has shown that fluid flow stimulates important functional changes in cells through protein expression regulation. Peroxiredoxins (PRX) are a family of antioxidant enzymes but have yet to be investigated in response to shear stress. Studies have shown that oscillatory shear stress (OS) increases reactive oxygen species (ROS) levels in endothelial cells, whereas laminar shear stress (LS) blocks this response. We hypothesized that PRX are responsible for the anti-oxidative effect of LS. To test this hypothesis, bovine aortic endothelial cells (BAEC) were subjected to LS (15 dyn/cm(2)), OS (+/-5 dyn/cm(2), 1 Hz), or static conditions for 24 h. Using Western blot and immunofluorescence staining, all six isoforms of PRX were identified in BAEC. When compared with OS and static, exposure to chronic LS up-regulated PRX 1 levels intracellularly. LS also increased expression of PRX 5 relative to static controls, but not OS. PRX exhibited broad subcellular localization, with distribution in the cytoplasm, Golgi, mitochondria, and intermediate filaments. In addition, PRX 1 knock down, using specific small interference RNA, attenuated LS-dependent reactive oxygen species reduction in BAEC. However, PRX 5 depletion did not. Together, these results suggest that PRX 1 is a novel mechanosensitive antioxidant, playing an important role in shear-dependent regulation of endothelial biology and atherosclerosis.