The pathogenesis of preeclampsia is directly related to vascular endothelial dysfunction and vasoconstriction. Soluble endoglin (sEng), an anti-angiogenic protein, has been implicated in the disruption of vascular homeostasis. Overexpression of sEng has been identified in preeclamptic placentas and serum of preeclamptic patients. sEng is thought to inhibit TGF-1 mediated nitric oxide (NO) vasodilatation. Herein we identified the signaling pathway of sEng that leads to inhibition of nitric oxide.
HTR-8/Svneo trophoblast cells were cultured and treated for 24 hours. Cell treatment groups included: no treatment (control), TGF-1 alone (20 ng/l), sEng alone (100 ng/l) and TGF-1 and sEng together (20 ng/l + 100 ng/l). Protein lysates were prepared for the Kinetworks KPSS 1.3 screen to determine phosphorylation targets and also for Western blotting. Quantitative densitometer readings indicating the intensity of phosphorylation at regulated peptide sites was analyzed using the Student t-test. By convention, phosphorylation sites demonstrating a change in response to sEng of at least 25% (up or down) were considered to be biologically meaningful.
The Kinetworks KPSS 1.3 screen followed by confirmatory immunoblotting identified a key phosphorylation site, on Akt, Thr 308, which was induced with TGF-1 and inhibited with the addition of sEng. Downstream of Akt, eNOS phosphorylation at Ser 1177 was similarly induced by TGF-1 but inhibited by adding sEng, leading to reduction in the activity of eNOS and decreased NO synthesis.
These results indicate that a mechanism for vasoconstriction in preeclampsia includes the de-phosphorylation of phospho-Akt (Thr 308) and phospho-eNOS (Ser 1177) in the presence of sEng. Identification of this signaling pathway elucidates some of the molecular mechanisms underlying preeclampsia. In conclusion, elevation of sEng, as in preeclampsia, initiates a cascade that leads to impaired NO production with potential vasoconstriction, thrombosis, and inflammation.
© 2008 Mosby, Inc. Published by Elsevier Inc. All rights reserved.