Altered Liver Secretion of Vascular Regulatory Proteins in Hypoxic Pregnancies Stimulate Angiogenesis in vitro

Placental vascular malformations result in fetal hypoxia, a serious pregnancy complication. Recent studies have linked liver-secreted and hemostatic proteins with angiogenesis. We therefore evaluated liver protein secretion changes following hypoxia, and their effect on angiogenesis, to identify potential angiogenic protein changes in the plasma of hypoxic newborns. Human vascular endothelial cells exhibited 10-fold increased tube formation with secretions from HepG2 cells cultured in 1% O₂ and 3-fold in 4% O₂ (p < 0.0001, p < 0.05) compared to 20% O₂. 2-DGE profiling of the secretions revealed significant density changes (p < 0.05) in spots identified as angiogenic proteins by LC−MS/MS. Clusterin decreased (−1.6-fold), whereas two spots of plasminogen activator inhibitor-1 (PAI-1) (2.4, and 3.6-fold), and three spots of transferrin (1.3, 1.5, and 2.6-fold) increased with 1% O₂. The levels of these proteins, subsequently determined in fetal plasma by immunoassays, strongly correlate with the fetal blood oxygen level at birth; PAI-1 and transferrin increase with low venous pO₂ (r = −0.70, p = 0.02, and r = −0.66, p = 0.04), clusterin and fibrinogen decrease (r = 0.82, p = 0.002, and r = 0.70, p = 0.02). These findings demonstrate that low oxygen levels in utero lead to pro-angiogenic changes in liver secreted plasma proteins. The pro-vascular plasma environment in hypoxic pregnancies may be acting to mitigate the compromised vasculature.