Abstract

The blood-brain barrier (BBB) is a network formed mainly by brain microvascular endothelial cells. The integrity of the BBB is critical for brain function. Breakdown of the BBB is commonly seen in AIDS patients with HIV-1-associated dementia (HAD), despite the lack of productive HIV-infection of the brain endothelium. The processes by which HIV causes these pathological conditions are not well understood. Here, we characterized the molecular mechanisms by which Tat mediates its pathogenic effects in-vitro on primary human brain microvascular endothelial cells (HBMECs). Tat treatment of HBMECs stimulated cytoskeletal organization and increased focal adhesion sites as compared to control cells or cells treated with heat-inactivated Tat. Pretreatment with Tat antibodies or with the specific inhibitor SU-1498, which interferes with VEGFR-2 (Flk-1/KDR) receptor phosphorylation, blocked the ability of Tat to stimulate focal adhesion assembly and the migration of HBMECs. Focal adhesion kinase (FAK) was tyrosine-phosphorylated by Tat and found to be an important component of focal adhesion sites. Inhibition of FAK by the dominant-interfering mutant form FRNK (FAK-related non-kinase) significantly blocked HBMEC migration and disrupted focal adhesions upon Tat activation. Furthermore, HIV-Tat induced permeability changes in HBMECs in a time dependent manner. Tat also impaired BBB permeability as observed in HIV-1 Tat transgenic mice. These studies define a mechanism for HIV-1 Tat in focal adhesion complex assembly in HBMECs, via activation of FAK, leading to cytoskeletal reorganization and permeability changes.

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