Astrocyte regulation of endothelial tissue plasminogen activator in a blood-brain barrier model

Expression of tissue plasminogen activator (tPA) substantially determines e ndothelial-dependent fibrinolysis. We used a blood-brain barrier (BBB) mode l to analyze regulation of brain capillary endothelial tPA and its inhibito r, plasminogen activator inhibitor-1 (PAI-1). This model consists of cocult ure of murine astrocytes with bovine brain capillary endothelial cells grow n as capillary-like structures (CS); after 1 week. astrocytes become extens ively associated with CS, and the BBB-associated enzyme gamma-glutamyl tran speptidase is present. We measured tPA and PAI-I mRNA and tPA activity in t his model. Reverse transcription-polymerase chain reaction (RT-PCR) studies showed similar tPA and PAI-I mRNA levels after 1 day mono-culture (endothe lial cells only) versus astrocyte-endothelial coculture preparations. After 7 days (i.e., when elements of the BBB are present), astrocyte-endothelial cocultures (compared with endothelial mono-cultures) showed a 50.7% +/- 27 .1% (mean +/- SD) reduction in tPA mRNA (P < 0.03) and a 183.3% +/- 86.98 i ncrease in PAI-I mRNA expression (P < 0.02). Moreover, 7-day cocultures dem onstrated reduced tPA activity compared with mono-cultures (14.6 +/- 2.9 IU /mL versus 30.2 +/- 7.7 IU/mL, P < 0.01); 1-day cocultures and mono-culture s had similar tPA activity. These findings demonstrate that astrocytes regu late brain capillary endothelial expression of tPA when elements of the BBB phenotype are present in this model. These data suggest an important role for astrocytes in the regulation of brain capillary endothelial fibrinolysi s.