Investigation of Nodal and Activin receptor like kinase 7 (ALK7) signaling pathway in human placenta from normal and compromised pregnancies

Trophoblast cell invasion is a critical process in placentation. Defective trophoblast invasion is linked to pregnancy complications, such as preeclampsia (PE), and intra-uterine growth restriction (IUGR). This aim of this study was to determine the role of Nodal, a member of the transforming growth factor-beta (TGF-beta) superfamily, and its type I receptor; Activin like kinase (ALK) 7, in the regulation of trophoblast cell invasion.;To examine the role of Nodal in trophoblast cell invasion, I used first trimester human trophoblast cell line and placental explants. I found that over-expression of Nodal or constitutively active ALK7, or treatment with recombinant Nodal resulted in decreased trophoblast cell migration/invasion and inhibited explants EVT outgrowth. On the other hand, siRNA mediated knock-down of Nodal reversed the effects of Nodal. Nodal up-regulated the secretion of tissue inhibitor of metalloproteinase-1 (TIMP-1) and inhibited matrix metalloproteinase (MMP) 2 and MMP9 activity, suggesting that TIMP-1/MMP2/MMP9 pathway is involved in mediating the effect of Nodal/ALK7 on trophoblast migration and invasion.;Further, I investigated the role of p27 in anti-proliferative and anti-migratory/invasive effects of Nodal. I demonstrated that Nodal stimulates p27 transcription and translation, enhances its mRNA and protein stability and regulates its phosphorylation and subcellular localization in trophoblast cells. Interestingly, Nodal induces simultaneous nuclear export of p27 and cdk2 resulting in inhibition of cell proliferation and migration/invasion. The displacement of cdk2 from its site of function inhibits cell proliferation while p27 binds to stathmin in the cytoplasm and inhibits its microtubule destabilizing effect resulting in immobility of the cell. Importantly, siRNA mediated knockdown of p27 abolished the inhibitory effect of Nodal on cell proliferation, migration and invasion, verifying that p27 is the key mediator of Nodal function.;Taken together, this study demonstrates that Nodal inhibits trophoblast cell invasion and that cytoplasmic translocation of p27 and cdk2 is the key molecular event by which Nodal regulates trophoblast cell proliferation, migration and invasion. This study also suggests that Nodal signaling plays an important role in the regulation of normal placental development and that aberration of Nodal/ALK7 signaling pathway may contribute to the development of PE.;Using immunohistochemistry, I determined the expression pattern of Nodal and ALK7 in human placentae from healthy and PE complicated pregnancies. In normal placenta, Nodal and ALK7 levels were high during first trimester, declined at the beginning of second trimester, and became undetectable at the end of pregnancy. Trophoblast cells, both villous and extravillous (EVT), and the apoptotic syncytial knots were found positive for Nodal and ALK7 expression. In addition, placenta from PE patients showed very strong immunoreactivity for both Nodal and ALK7, as compared to their age-matched controls.