Research Grants 2002-2003

Development of an in vitro Cell Model of the Human Blood Brain Barrier

Senén Vilaró, PhD
School of Biology, University of Barcelona, Barcelona, Spain

The main obstacle for efficient drug targeting to the brain lies on the blood-brain barrier (BBB). Separating brain from blood at the level of the endothelial cells from cerebral capillaries, this cell barrier strictly controls the entry of solutes into the central nervous system. To assess the toxicity or bioavailability of drugs in development, academic groups and pharmaceutical companies would benefit from an in vitro model of BBB, based on brain endothelial cells in culture. In response to the need of such in vitro models, the main objective of our proposal is to develop easy-to-use stable immortalized endothelial cell lines from human brain microvessels, which will retain their functional properties and allow to develop an in vitro model of the human BBB as a highly reliable and reproducible alternative method to animal experimentation for pharmacological and toxicological testing of CNS-directed compounds.

Using the isolation procedure developed during the first year of CAAT funding, primary cultures of microvascular endothelial cells will be obtained from total brain samples from post-mortem multidonor patients. As BBB properties are quickly lost in culture, immortalization will be performed on early primary cultures using viral constructs expressing the human telomerase, as immortalizing gene. Selection will be based on expressing the human telomerase, as immortalizing gene. Selection will be based on growth properties and expression of specific brain endothelial markers. Positive immortalized subpopulations will be characterized, in solo or in co-culture with astrocytes, for transendothelial electrical resistance and permeability to standard molecules with known BBB profiles. A correlation will be established between in vitro and in vivo permeability values. Once validated, this in vitro cell model of human BBB will constitute a valuable alternative to animal use for product screening and safety tests. Its application in large high through-put screening will accelerate the CNS-oriented drug discovery, and decrease the R&D costs for pharmaceutical industry.