MIXED CULTURE OF PERICYTES AND ENDOTHELIAL-CELLS FROM FETAL MICROVESSELS OF THE HUMAN PLACENTA

To explore the role of perivascular cells in angiogenesis and vasomotr icity, placental cultures of perivascular cells were performed from ca librated villi excised from term placentas. Microvessels were isolated using repeated digestion of villi by collagenase-dispase and purifica tion by Percoll gradients. Plated on Petri dishes, the microvessels be came adherent to the gelatin matrix permitting to cells to proliferate . Cells were harvested and subcultured. Endothelial and pericyte cell lines were identified by phase contrast microscopy. Pericyte number pr edominated rapidly, the endothelial cells remaining visible. After sev en days, cells started to cluster, thus piled up and built numerous no dules. Medium-size oval endothelial cells were stained by anti-von Wil lebrand factor and anti-IgG coupled to fluorescein. Large cells with i rregular border reacted to smooth muscle anti-a-actin and anti-IgG cou pled to fluorescein. There was no cross-reaction of these two cell typ es with the antibodies. In contrast, nodules were stained by both immu nostainings. Endothelial cells reacting to von Willebrand factor antib ody were frequently associated to the nodule. The isolation of microve ssels from the human placenta described in this study allowed the esta blishment of cultures of endothelial cells and pericytes that show: i) rapid predominance of pericytes over endothelial cells, ii) formation of nodules, iii) participation of endothelial cells and pericytes to nodules formation.