In pathological situations such as cerebral ischemia, excessive increases in vascular permeability lead to opening of the blood brain barrier (BBB) and vasogenic edema, which is a major cause of death in acute stroke patients. Any therapeutic strategy aimed at attenuating the disruption of the BBB and therefore the severity of vasogenic edema will have a significant impact in the mortality of patients with ischemic stroke. Tissue-type plasminogen activator (tPA) is predominantly found in the blood where its primary function is as a thrombolytic enzyme. tPA is also expressed within the central nervous system (CNS) where it is thought to have a different function. In ischemic stroke, endogenous tPA activity increases within the ischemic hemisphere and either genetic deficiency or pharmacological inhibition of tPA have been associated with decrease in infarct volume. Preliminary studies demonstrate that both endogenous tPA released from intracellular stores in response to the ischemic insult and exogenous tPA administered as a thrombolytic therapy for embolic stroke, directly cause disruption of the BBB. Since tPA is the only FDA-approved medication for the treatment of patients with acute stroke, it is of vital importance to establish whether this proteinase has a deleterious effect on the BBB during cerebral ischemia. Accordingly, this application will focus on understanding the role of tPA as a regulator of cerebrovascular permeability during cerebral ischemia. By using a combination of biochemical, molecular, pharmacological and genetic approaches, both in vitro and in vivo, and building on previous studies of the effect of tPA and the Lipoprotein Receptor Related Protein (LRP) on cerebrovascular permeability, we will test the following hypotheses: 1) following the onset of cerebral ischemia there is an early increase in the release of tPA, possibly from the glial cells; 2) the association of this tPA with LRP in the BBB induces a specific cell signaling event, likely through phosphorylation of a member of the Mitogen-Activated Protein Kinase (MAPK) family; and 3) MAPK phosphorylation results in changes in cytoskeletal and tight junction (TJ) proteins with subsequent increases in cerebrovascular permeability. ? ? ?
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