The role of poly(ADP-ribose) polymerase (PARP) in the autonomous proliferative response of endothelial cells to hypoxia

Objective: The autonomous proliferative response of endothelial cells to hypoxia has been shown to be dependent on activation of NAD(P)H oxidase, on the cytosolic Ca2+ load, and, consequently, on nuclear translocation of extracellular signal-regulated kinase (ERK)1/2 during transient hypoxia. The aim of the present study was to investigate whether poly(ADP-ribose) polymerase (PARP) is a downstream signal of NAD(P)H oxidase, mediating cytosolic Ca2+ load and hence nuclear translocation of ERK1/2 and endothelial cell proliferation. Methods: Porcine aortic endothelial cells were incubated under hypoxic conditions for 40 min. Cytosolic [Ca2+] and reactive oxygen species (ROS) formation were measured in fura-2- and DCF-loaded cells, respectively. PARP activation was detected by immunocytochemistry, and endothelial cell proliferation was determined 24 h after 60 min of transient hypoxia. Results: Inhibition of NAD(P)H oxidase with antisense oligonucleotide against the p22(phox) subunit, MEK/ERK signalling with UO 126 (30 mu M), or PARP with PJ 34 (10 mu M) leads to a marked reduction in hypoxia-induced cytosolic Ca2+ load and activation of PARP. Hypoxia-induced translocation of ERKI/2 and endothelial cell proliferation were also prevented when NAD(P)H oxidase or PARP were inhibited; however, hypoxic ROS formation was not affected in the presence of PARP inhibitor. Conclusion: PARP represents a downstream effector of NADP(H) oxidase and acts as a necessary intermediate step for the hypoxic proliferative response of endothelial cells. (c) 2006 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.