Migration of brain capillary endothelial cells inside poly (lactic acid) 3D scaffolds

The brain capillary endothelial cells (BCECs) form the blood brain barrier (BBB) under the effects of the brain microenvironment. BCECs are sealed together by tight junctions (TJs) that are responsible for the barrier phenotype. In these junctions, molecules such as JAM (junctional adhesion molecules), occludin and claudins are present. Threedimensional scaffolds are used to grow cells in order to obtain in vitro engineered tissues. On the base of these considerations, the aim of this work was to understand whether the endothelial cells were able to grow and survive on a new three-dimensional structure. If yes, indeed, this system could be further enriched and used to set a three-dimensional in vitro model of BBB containing also neurons and astrocytes. The starting work has been focusing on BCEC growth on poly-L-lactate (PLLA) 3D scaffold. BCECs were seeded (50,000 cells/scaffold) on scaffolds prepared at 0°C and precoated with Collagen IV. TJ distribution was analyzed by immunofluorescence with zonula occludens 1 (ZO1) antibodies and the number of alive cells was evaluated by staining cells with acridine orange (AO)/Ethidium bromide (Etbr). Moreover, scaffolds were fixed, sectioned and observed by transmission electron microscopy (TEM). Our data demonstrate that BCECs adapt well to this kind of scaffolds and are also able to migrate inside the pores present in them.