Hypoxia Promotes Extravillous Trophoblast Cell Invasion through the Hypoxia-Inducible Factor Urokinase-Type Plasminogen Activator Receptor Pathway

Abstract

<b><i>Objectives:</i></b> Hypoxia is a common feature of extravillous trophoblast (EVT) cells that promote invasion during the early stages of human placentation. This study aimed to examine whether hypoxia-induced an invasive phenotype in EVT cells in vitro and explore the underlying molecular mechanisms. <b><i>Design:</i></b> The invasiveness of primary EVT cells isolated from the first trimester placental tissues during weeks 5–8 of gestation was examined under hypoxic (5% O₂) and normoxic (20% O₂) conditions. <b><i>Methods:</i></b> Invasiveness was determined by transwell and wound-healing invasion assays using the IncuCyte ZOOM™ Live-Cell Imaging System. Protein expression of the urokinase plasminogen activator (uPA), uPA receptor (uPAR), and plasminogen activator inhibitor of hypoxia or normoxia-treated cells was measured using Western blot analysis. Knockdown of hypoxia-inducible factor-1 alpha (HIF-1α) was assessed using small interfering RNA (siRNA). <b><i>Results:</i></b> Hypoxia enhanced EVT cell invasion but did not affect apoptosis. The stimulatory effect of hypoxia on EVT cell invasiveness was associated with induction of the uPA-uPAR pathway. The synthetic inhibitor of uPAR significantly inhibited hypoxia-induced EVT cell invasion. Silencing of HIF-1α by siRNA abolished the stimulatory effect of hypoxia and inhibited the upregulation of uPAR expression, suggesting that the HIF-1α-uPAR signal is the key mediator for hypoxia-induced EVT cell invasion. Further experiments need to be performed to elucidate the HIF-1α-uPAR signal pathways. <b><i>Conclusions:</i></b> The low oxygen-regulated early events of EVT invasion may be mediated by the HIF-1α-uPAR pathway.