Abstract 13533: Thioredoxin Interacting Protein Prevents Endothelial Cell Apoptosis - Critical Mediator of VEGF Receptor 2 Activation via Poly-ADP Ribose Polymerase 1
Background. Oxidative stress and inflammation are major contributors to the development of cardiovascular diseases, such as atherosclerosis. Poly-ADP ribose polymerase 1 (PARP1) and Thioredoxin-interacting protein (TXNIP) are both nuclear proteins that are regulated by changes in the redox state of endothelial cells (EC). Previously we showed a novel mechanism for EC protection from stress-induced apoptosis via PARP1-inhibition and VEGFR2 transactivation. Furthermore, we previously showed a novel redox dependent mechanism for TXNIP localization to the plasma membrane (PM) and activation of VEGFR2. Because TXNIP is an established arrestin-like protein and is regulated by changes in redox signaling, we hypothesized that PARP1 regulates TXNIP localization and function that might have important physiologic consequences in EC inflammation and death.
Methods and Results. Immunofluorescence and western blot analysis were utilized to demonstrate that Human umbilical vein endothelial cells (HUVEC) treated with 10 umol/L PARP1 inhibitor (PJ34) were protected from TNF (10 ng/ml) or H2O2 (300 umol/L) mediated apoptosis. HUVEC transfected with TXNIP siRNA lost the protective effect of PARP1 inhibition, suggesting a protective role for TXNIP. Using Immunofluorescence, cell fractionation analysis and PM sheet assay we report TXNIP localization to the PM after 30 minutes stimulation of TNF (3.5-fold increase). TXNIP translocation to the PM was associated with increased PM tyrosine phosphorylation (20-fold increase). Importantly, activation of pro-survival signaling pathways including VEGFR2 and Akt in response to PARP1 inhibition was found to be TXNIP-dependent. Mechanistically, immunoprecipitation demonstrated that a complex of PARP1 and TXNIP was present under basal conditions, and disrupted after treatment with PARP1 inhibitor. Functionally, changes in TXNIP-PARP1 interaction suggest PARP1 as a novel regulator of TXNIP localization and function.
Conclusions. PARP1 regulates TXNIP subcellular localization and its ability to protect EC from stress-induced apoptosis via activation of pro-survival signaling at the PM.
- © 2011 by American Heart Association, Inc.