Blood-brain barrier characteristic enzymatic properties in cultured brain capillary endothelial cells

Isolated brain capillary endothelial cells contain high activity levels of the blood-brain barrier (BBB) marker enzymes gamma-glutamyltranspeptidase (gamma-GT) and alkaline phosphatase (ALP). In primary culture the activities of these specific enzymes decrease with increasing cell proliferation to a constant low value characteristic for a confluent monolayer. However, activities are retained in non-proliferating cells. After passage of cells from a confluent cell monolayer a further reduction of enzyme activity was observed which corresponds to the newly triggered cell proliferation. Culture of cerebral endothelial cells on structural components of the native vascular basement membrane-like type IV collagen, fibronectin, laminin or a commercially available basement membrane cannot prevent the activity decrease of both gamma-GT and ALP. Antiserum raised against a native renal dog gamma-GT binds to the cerebral endothelial gamma-GT and suppresses its activity. The relative activity decrease induced by a given amount of anti-gamma-GT-antiserum is constant at all times in culture. This result clearly shows that the observed decrease in gamma-GT activity in proliferating cells in culture correlates to a decreased number of enzyme molecules per cell and not to an inhibition of expressed enzymes. Possibly the de novo synthesis of this enzyme is prevented in vitro. In contrast to the loss of the activity of the BBB marker enzymes gamma-GT and ALP, the activity of angiotensin-converting enzyme (ACE), a marker for all vascular endothelial cells, is highly preserved in cultured cerebral endothelial cells.