Angiogenin Expression during Early Human Placental Development; Association with Blood Vessel Formation
Characterisation by double labelling of chorionic villi undergoing intense morphological changes. Frozen cross-sections of human chorionic villi at week 8 were reacted with a mix of two primary antibodies. Angiogenin was detected with a specific polyclonal antibody (ANG, in red). The other antibody (in green) was directed against either the early endothelial markers: VE-cadherin (a, b) and VEGF-R2 (c, d), or the proliferation marker Ki-67 (e, f). Cells positive for the endothelial markers were stained for angiogenin. Note the angiogenin- and VE-cadherin-positive single cell (a, arrowhead) beneath the trophoblastic layer establishing a bridge between the layer and a vascular cell aggregate (VCA) stained for VE-cadherin; VEGF-R2-labelled VCA in close contact with the trophoblast layer (c); and angiogenin-labelled cytoplasmic processes (d). Proliferative cells labelled for Ki-67 and angiogenin were observed in aggregates (e, green arrow). Coordinated sites of proliferation (Ki-67+) composed of villous trophoblasts and facing nontrophoblastic cell were positive for angiogenin (f, green arrow). White and yellow arrows point to VE-cadherin and VEGF-R2 staining of trophoblast layer, respectively. Nuclei were counterstained with Dapi, in blue. Bar, 20 μm. IS: intervillous space; TB: trophoblast layer; VCA: vascular cell aggregate; and VM: villous mesenchyme.