tPA Variant tPA-A296-299 Prevents Impairment of Cerebral Autoregulation After Stroke Through LRP Dependent Increase in cAMP and p38 MAPK

Introduction: The sole FDA approved treatment for acute stroke is tissue type plasminogen activator (tPA). However, its brief therapeutic window and post-treatment complications markedly constrain its use. The limited efficacy of tPA may be explained in part by our finding that it potentiates impairment of cerebral autoregulation after stroke in pigs. Upregulation of JNK MAPK after thrombotic stroke contributes to tPA mediated impairment of autoregulation whereas p38 is protective. Vasodilation is mediated by elevation of cAMP. However, cAMP falls after stroke due to over activation of NMDA receptors (NMDA-Rs) by toxic levels of glutamate, which is exacerbated by tPA. Hypothesis: Wild type (wt) tPA can bind to either the lipoprotein-related receptor (LRP), which mediates vasodilation, or NMDA-Rs. We propose that wt tPA given after stroke primarily binds to NMDA-Rs, reduces cAMP, which impairs ability of cerebral vessels to vasodilate. tPA-A296-299, a variant that is fibrinolytic but cannot bind to NMDA-Rs, preferentially binds LRP, which increases cAMP and p38 and limits impairment of autoregulation after stroke. Methods: Anesthetized pigs equipped with a closed cranial window were used. Stroke was induced by photothrombosis, CBF determined by radiolabeled microspheres, and CSF cAMP and p38 determined by ELISA. Results: CBF was unchanged during hypotension (mean arterial pressure decreased by 45%) in sham animals. CBF was reduced in brain tissue surrounding the infarct and was reduced further during hypotension, indicating impairment of autoregulation. Autoregulation was further impaired by wt tPA which was prevented by MK801 and tPA-A296-299. Protection by tPA-A296-299 was blocked by anti LRP Ab, the LRP antagonist RAP, the p38 inhibitor SB 203580, but not by IgG. Stroke reduced CSF cAMP, which was reduced further by wt tPA, but augmented by tPA-A296-299. CSF p38 was unchanged by wt tPA, robustly increased by tPA-A296-299, and decreased by co-administered anti LRP Ab and RAP but not by IgG. Conclusions: tPA-A296-299 prevents impairment of cerebral autoregulation after stroke through LRP dependent increase in cAMP and p38.