Tissue type plasminogen activator (tPA) induces neuronal apoptosis, disrupt the blood-brain-barrier (BBB), and promotes dilation of the cerebral vasculature. The timing, sequence and contributions of these and other deleterious effects of tPA and their contribution to post-ischemic brain damage after stroke, have not been fully elucidated. To dissociate the effects of tPA on BBB permeability, cerebral vasodilation and protease-dependent pathways, we developed several tPA mutants and PAI-1 derived peptides constructed by computerized homology modeling of tPA. Our data show that intravenous administration of human tPA to rats increases BBB permeability through a non-catalytic process, which is associated with reversible neurotoxicity, brain damage, edema, mortality and contributes significantly to its brief therapeutic window. Furthermore, our data show that inhibiting the effect of tPA on BBB function without affecting its catalytic activity, improves outcome and significantly extends its therapeutic window in mechanical as well as thromboembolic models of stroke.

Urokinase plasminogen activator (uPA) plays a major role in fibrinolytic processes and also can potentiate LPS-induced neutrophil activation through interactions with its kringle domain (KD). To investigate the role of the uPA KD in modulating acute inflammatory processes in vivo, we cloned and then developed Abs to the murine uPA KD. Increased pulmonary expression of uPA and the uPA KD was present in the lungs after LPS exposure. Administration of anti-kringle Abs diminished LPS-induced up-regulation of uPA and uPA KD in the lungs, and also decreased the severity of LPS-induced acute lung injury, as determined by development of lung edema, pulmonary neutrophil accumulation, histology, and lung IL-6, MIP-2, and TNF-α cytokine levels. These proinflammatory effects of the uPA KD appeared to be mediated through activation of Akt and NF-κB. The present studies indicate that the uPA KD plays a major role in the development of TLR4-mediated acute inflammatory processes, including lung injury. Blockade of the uPA KD may prevent the development or ameliorate the severity of acute lung injury induced through TLR4-dependent mechanisms, such as would occur in the setting of Gram-negative pulmonary or systemic infection.