HCG in the regulation of placental angiogenesis. Results of an in vitro study

Abstract

Placental vascular development is essential for fetal growth and development. Inadequate placental vascular development is associated with early pregnancy losses and other pregnancy related pathologies. In addition to the ubiquitous, well-characterized angiogenic factors like vascular endothelial growth factor (VEGF) or basic fibroblast growth factor (bFGF), some pregnancy-specific factors (e.g. human chorionic gonadotropin (hCG), insulin-like growth factor-II (IGF-II) or alpha fetoprotein (AFP) were recently described to play a possible regulatory role in this process. In the present study we described an improved separation method for human placental microvascular endothelial cells (HPMVEC) and their functional characterization. Using the combination of enzymatic digestion and multistep immunomagnetic sorting with CD31 antibodies a model for villous vascularization was established. Isolated cells took up ac-dil-LDL, spontaneously formed capillary-like structures, and expressed common endothelial markers such as vascular endothelial growth factor receptor-2 (VEGFR-2), angiopoetin-1 and -2, Tie-2, CD144, thrombomodulin, and von Willebrand factor (vWF) as shown by RT-PCR, flow cytometry and Western blot analysis. The expression of the hCG/LH receptor in the placental vascular tree was verified both in vitro and in vivo. hCG stimulated proliferation of HPMVEC in a dose specific manner. While hCG alone had no significant effect on endothelial cell apoptosis, the combination of VEGF-A and hCG protected HPMVEC from staurosporine-induced apoptosis. hCG significantly stimulated sprout formation when compared to controls in a spheroid angiogenesis assay. Our results demonstrate a modified and reproducible method allowing studies of placental vascular development and provide new insights into the possible role of trophoblastic factors in this process.