Hypoxia induces trophoblast migration and invasion gene pathways, a subset of which are reduced in preeclampsia

Placentation requires the proper regulation of extravillous trophoblast (EVT) migration and invasion into the decidua and maternal vasculature, processes which are initiated in physiologic hypoxic conditions. Abnormal EVT migration and/or invasion have been suggested to lead to pregnancy complications, such as preeclampsia. The objectives of this study are to determine how exposure to hypoxia impacts gene expression and cellular motility of first trimester trophoblasts, and to assess if expression of migration-associated genes is dysregulated in 2nd trimester chorionic villous samples (CVS) from preeclampsia pregnancies relative to CVS from healthy pregnancies. The 1st trimester trophoblast cell line, HTR8/SVneo, was used to investigate the relationship between hypoxia and Notch signaling in trophoblast migration and invasion. RNA sequencing and quantitative RT-PCR analyses show that exposure to hypoxia (2.5% O2) activates Notch signaling in HTR-8/SVneo. We demonstrate that exposure of HTR-8/SVneo to hypoxia induces expression of genes associated with cellular migration and invasion and increases HTR-8/SVneo cellular migration and invasion, whereas inhibition of gamma-secretase decreases Notch signaling and decreases HTR-8/SVneo migration and invasion. Analysis of RNA sequencing data from CVS of preeclampsia and uncomplicated pregnancies identified significant differentially expressed genes that are involved in cellular migration and invasion. Decreased expression of migration and invasion genes in CVS from preeclampsia pregnancies, may impair trophoblast migration and invasion in the 2nd trimester of pregnancy, resulting in the development of preeclampsia. Overall design: 1) HTR-8/SVneo: 6 samples were analyzed; n=3 samples for the control group (FC1, FC2, FC3) were exposed to normoxic conditions (21% 02); n=3 samples for the experimental group (Hyp7, Hyp8, Hyp9) were exposed to hypoxic conditions (2.5% 02). For each condition, RNA was extracted from cells that were grown and processed independently to serve a replicates. 2) Chorionic villous sampling: n=6 samples from 6 unique human pregnancies were analyzed; n=4 samples were from uncomplicated, healthy pregnancies (CVS24, CVS25, CVS27, CVS33); n=2 samples were from preeclampsia pregnancies (CVS20, CVS32)