Endothelial cells are known to participate in angiogenesis, adaptation of vascular tonus and maintenance of blood fluidity in the microcirculation. To investigate these functions in the placenta, we devised a method of isolation and culture of endothelial cells from villous microvessels. In primary culture, these intraplacental endothelial cells exhibited many features observed in microvascular endothelium from other organs: spindle-shape, rosette associations, circular arrangements and confluence. In contrast to the confluent endothelial cells derived from the umbilical vein, cells from microvessels did not form a cobblestone network. After trypsin digestion of microvessels, magnetic microbeads coated with S-Endol immunoglobulin, antithrombomodulin and Ulex europaeus-I lectins were tested for sorting endothelial cells. Only the microbeads coated with antithrombomodulin allowed a suitable magnetic cell separation after trypsinization. By contrast, the microbeads coated with each of these antibodies or with lectins attached to confluent cells from the second passage. The microbeads detached from the cells at different rates. Their examination by scanning microscopy indicates that a portion of these microbeads was phagocytosed. Microvascular endothelial cells from the second passage were intensively stained by the anti-von Willebrand reaction and only weakly by the anti-smooth muscle alpha-actin reaction. They incorporated acetylated-low density lipoproteins coupled to a fluorescent probe. The positive reactions against the anti-von Willebrand factor and the uptake of the fluorescent acetylated-low density lipoproteins were modified after eight passages.

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The cellular localization of GLUT3 messenger ribonucleic acid (mRNA) and protein was examined in human term placenta using a combination of methodologies. In situ hybridization indicated that GLUT3 mRNA was present in the trophoblast cell layer and in vascular endothelium with a heterogeneous distribution pattern. GLUT3 protein migrating at an apparent molecular mass of 49 kDa was detected by immunoblotting in membranes from whole placenta and endothelial cells derived from intraplacental microvessels, but not in isolated trophoblast cells. This cell-specific pattern of expression was confirmed by immunocytochemical studies showing a prominent localization of GLUT3 protein in vascular endothelium. These findings indicate a differential distribution of GLUT3 mRNA and protein in the human placenta. Based on its cell-specific distribution at the fetal interface, GLUT3 protein could be of cardinal importance in the transport of glucose from the placenta to the fetal circulation.

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