Co-culture of rat brain cells as a tool for studying cell-cell interactions

Brain capillary endothelial cells (BCECs) form the blood-brain barrier (BBB) in response to interaction with other brain cells (astrocytes, pericytes and neurons). BCECs are characterized by tight junctions (TJ), maturation and stabilization of which require different proteins, such as occludin. When co-cultured with astrocytes and neurons, BCECs were found to form a monolayer resembling the natural BBB: paracellular flux of dopamine and sucrose (i.e. compounds which are unable to cross the BBB in vivo) significantly decreased (1), while the transendothelial electrical resistance (TEER) increased. In these conditions, BCECs produced a larger amount of occludin and tended to localize it at the cell periphery, thus suggesting formation of TJs (1). Since we also discovered that oligodendroglioma cells shed extracellular membrane vesicles (MVs; 2), we investigated whether also neurons and/or astrocytes can release MVs and whether these vesicles contained angiogenic factors. The results of these analyses demonstrated that all kinds of brain cells actually shed MVs containing FGF-2 and VEGF (3-4). On the basis of these findings, we investigated the possibility that the BBB model could be used to study the molecular events that result in BBB damage, in some pathological conditions, such as, for example, multiple sclerosis (5). We are now investigating whether cultured astrocytes shed vesicles containing aquaporin 4 (AQP4), a protein which has been involved in brain edema. Our results suggest that production of AQP4 increases in stressed astrocytes.