Reversal of gene dysregulation in cultured cytotrophoblasts reveals possible causes of preeclampsia
Yan Zhou, … , Michael T. McMaster, Susan J. Fisher
Yan Zhou, … , Michael T. McMaster, Susan J. Fisher
Published June 24, 2013
Citation Information: J Clin Invest. 2013;123(7):2862-2872. https://doi.org/10.1172/JCI66966.
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Research Article

Reversal of gene dysregulation in cultured cytotrophoblasts reveals possible causes of preeclampsia

Abstract

During human pregnancy, a subset of placental cytotrophoblasts (CTBs) differentiates into cells that aggressively invade the uterus and its vasculature, anchoring the progeny and rerouting maternal blood to the placenta. In preeclampsia (PE), CTB invasion is limited, reducing placental perfusion and/or creating intermittent flow. This syndrome, affecting 4%–8% of pregnancies, entails maternal vascular alterations (e.g., high blood pressure, proteinuria, and edema) and, in some patients, fetal growth restriction. The only cure is removal of the faulty placenta, i.e., delivery. Previously, we showed that defective CTB differentiation contributes to the placental component of PE, but the causes were unknown. Here, we cultured CTBs isolated from PE and control placentas for 48 hours, enabling differentiation and invasion. In various severe forms of PE, transcriptomics revealed common aberrations in CTB gene expression immediately after isolation, including upregulation of SEMA3B, which resolved in culture. The addition of SEMA3B to normal CTBs inhibited invasion and recreated aspects of the PE phenotype. Additionally, SEMA3B downregulated VEGF signaling through the PI3K/AKT and GSK3 pathways, effects that were observed in PE CTBs. We propose that, in severe PE, the in vivo environment dysregulates CTB gene expression; the autocrine actions of the upregulated molecules (including SEMA3B) impair CTB differentiation, invasion and signaling; and patient-specific factors determine the signs.

Authors

Yan Zhou, Matthew J. Gormley, Nathan M. Hunkapiller, Mirhan Kapidzic, Yana Stolyarov, Victoria Feng, Masakazu Nishida, Penelope M. Drake, Katherine Bianco, Fei Wang, Michael T. McMaster, Susan J. Fisher

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Figure 5

SEMA3B inhibited PI3K/AKT and GSK3β signaling in CTBs and the same effects were observed in sPE.

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SEMA3B inhibited PI3K/AKT and GSK3β signaling in CTBs and the same effec...
(A) SEMA3B and wortmannin (WM) inhibited PI3K activity, which was stimulated by VEGF. DMSO was used as a vehicle control. Mean ± SEM, 2-tailed Student’s t test. *P < 0.05, **P < 0.01. (B) The addition of SEMA3B to UtMVECs resulted in the dissociation of the p85 and the p110α subunits of PI3K, which was rescued by the addition of VEGF. (C) In COS-1 cells, SEMA3B inhibited AKT Ser473 phosphorylation (activation), which increased during CTB differentiation/invasion (0–12 hours). The addition of SEMA3B inhibited AKT phosphorylation, which was enhanced by exogenous VEGF. (D) In COS-1 cells, SEMA3B inhibited GSK3β Ser9 phosphorylation (inactivation), which increased during CTB differentiation/invasion (0–12 hours). Exogenous SEMA3B inhibited GSK3β phosphorylation, which was enhanced by VEGF. GSK3α Ser21 phosphorylation was variable. (E) In CTBs, sPE correlated with dissociation of the p85 and p110α (and γ) subunits of PI3K relative to control cells isolated from normal third trimester placentas. (F) In freshly isolated CTBs, sPE was associated with decreased phosphorylation of AKT Ser473 and GSK3β Ser9. α-Actin was used as a loading control. (G) In chorionic villi, sPE was associated with phosphorylation (inactivation) of β-catenin. (A–D) The same results were obtained in 3 separate experiments that used different preparations of cells. (F and G) The results shown are representative of analyses of a total of 6 CTB isolates from different placentas of women diagnosed with sPE.