During early pregnancy, placentation occurs in a relatively hypoxic environment that is essential for appropriate embryonic development. Intervillous blood flow increases around 10 to 12 weeks of gestation and results in exposure of trophoblast cells to increased oxygen tension. Before this time, low oxygen appears to prevent trophoblast differentiation toward an invasive phenotype. Using human villous explants of 5–8 weeks’ gestation, we found that low oxygen tension triggered trophoblast proliferation, fibronectin synthesis, α₅ integrin expression, and gelatinase A activity. These biochemical markers were barely detectable under oxic conditions. We therefore examined the placental expression of hypoxia-inducible factor-1 (HIF-1), a master regulator of oxygen homeostasis, and determined that expression of HIF-1α subunit during the first trimester of gestation parallels that of TGFβ₃, an inhibitor of extravillous trophoblast differentiation. Expression of both molecules is high in early pregnancy and falls around 9 weeks of gestation, when placental pO₂ levels are believed to increase. Increasing oxygen tension induced a similar decrease in expression in cultured explants. Moreover, antisense inhibition of HIF-1α expression in hypoxic explants inhibited expression of TGFβ₃, arrested cell proliferation, decreased α₅ expression and gelatinase A activity, and triggered biochemical markers of an invasive trophoblast phenotype such as α₁ integrin and gelatinase B expression. These data suggest that the oxygen-regulated early events of trophoblast differentiation are in part mediated by TGFβ₃ through HIF-1 transcription factors.

Background: Oxygen plays a central role in human placental pathologies including preeclampsia, a leading cause of fetal and maternal death and morbidity. Insufficient uteroplacental oxygenation in preeclampsia is believed to be responsible for the molecular events leading to the clinical manifestations of this disease. Design: Using high-throughput functional genomics, we determined the global gene expression profiles of placentae from high altitude pregnancies, a natural in vivo model of chronic hypoxia, as well as that of first-trimester explants under 3 and 20% oxygen, an in vitro organ culture model. We next compared the genomic profile from these two models with that obtained from pregnancies complicated by preeclampsia. Microarray data were analyzed using the binary tree-structured vector quantization algorithm, which generates global gene expression maps. Results: Our results highlight a striking global gene expression similarity between 3% O₂-treated explants, high-altitude placentae, and importantly placentae from preeclamptic pregnancies. We demonstrate herein the utility of explant culture and high-altitude placenta as biologically relevant and powerful models for studying the oxygen-mediated events in preeclampsia. Conclusion: Our results provide molecular evidence that aberrant global placental gene expression changes in preeclampsia may be due to reduced oxygenation and that these events can successfully be mimicked by in vivo and in vitro models of placental hypoxia.

Elevated expression of soluble vascular endothelial growth factor receptor-1 (sFlt-1) in preeclampsia plays a major role in the pathogenesis of this serious disorder of human pregnancy. Although reduced placental oxygenation is thought to be involved in the pathogenesis of preeclampsia, it is unclear how oxygen regulates placental sFlt-1 expression. The aims herein were to investigate sFlt-1 expression in in vivo and in vitro physiological and pathological models of human placental hypoxia and to understand the role of hypoxia inducible factor-1 (HIF-1) in regulating the expression of this molecule. sFlt-1 expression in placental villi was significantly increased under physiological low oxygen conditions in early first-trimester and in high-altitude placentae, as well as in pathological low oxygen conditions, such as preeclampsia. In high-altitude and in preeclamptic tissue, sFlt-1 localized within villi to perivascular regions, the syncytiotrophoblast layer, and syncytial knots. In first-trimester villous explants, low oxygen, but not hypoxia-reoxygenation (HR), increased sFlt-1 expression. Moreover, exposure of villous explants to dimethyloxalyl-glycin, a pharmacological inhibitor of prolyl-hydroxylases, which mimics hypoxia by increasing HIF-1α stability, increased sFlt-1 expression. Conversely, HIF-1α knockdown using antisense oligonucleotides, decreased sFlt-1 expression. In conclusion, placental sFlt-1 expression is increased by both physiologically and pathologically low levels of oxygen. This oxygen-induced effect is mediated via the transcription factor HIF-1. Low oxygen levels, as opposed to intermittent oxygen tension (HR) changes, play an important role in regulating sFlt-1 expression in the developing human placenta and hence may contribute to the development of preeclampsia.

Preeclampsia, the major cause of maternal morbidity and mortality in developed countries, is associated with abnormalities of placenta function due to shallow invasion of the maternal decidua by trophoblasts. Data suggest that TGF-β may play a role in inhibiting trophoblast outgrowth or invasion, or both. We report that placental TGF-β₃ expression is high in early pregnancy but falls at around 9 weeks’ gestation. This pattern is inversely correlated with trophoblast outgrowth and fibronectin synthesis, markers of early trophoblast differentiation toward an invasive phenotype. We demonstrate that TGF-β₃ is overexpressed in preeclamptic placentae. In contrast to control placentae, explants from preeclamptic pregnancies fail to exhibit spontaneous invasion in vitro. Significantly, antisense-induced inhibition of TGF-β₃ expression, and inhibition of TGF-β₃ activity with antibodies, induces the formation of columns of trophoblast cells, which migrate out of the explant into the underlying Matrigel. To our knowledge, this is the first demonstration that the hypoinvasive placental phenotype characteristic of preeclampsia can be essentially normalized in vitro by biochemical manipulation. We speculate that a failure to downregulate expression of TGF-β₃ at around 9 weeks’ gestation results in shallow trophoblast invasion and predisposes the pregnancy to preeclampsia.