Abstract WP133: Prevention of the Post-stroke Induction of Endogenous tPA Mitigates Brain Damage and Facilitates Neurological Recovery

Abstract

Introduction: Multiple pieces of evidence suggest that the elevated endogenous tPA (endo-tPA) levels in the brain after ischemic stroke are neurotoxic and contribute to the ongoing brain damage. The neurotoxicity of endo-tPA could be due to activation of apoptotic cell signaling processes, extracellular matrix degradation, or increase in permeability of the neurovascular unit. The purpose of this study is to investigate the effect of specific endo-tPA suppression on post-stroke brain injury and neurological recovery. Methods: Male Sprague-Dawley rats were subjected to a suture model middle cerebral artery occlusion (MCAO) procedure followed by reperfusion. To achieve the specific knockdown of endo-tPA, plasmids expressing shRNAs specific to rat tPA (tPAsh) formulated as nanoparticles were administered immediately after reperfusion to rats intravenously via tail vein at a dose of 1 mg/kg body weight. Untreated, and vehicle/tPAsh-treated stroke-induced rats along with sham-operated rats were sacrificed at various post-reperfusion time points. We performed various techniques such as TTC staining, real-time PCR, and immunoblot analysis to determine both the efficiency of tPAsh plasmids and efficacy of tPAsh treatment on post-stroke brain injury. To assess the effect of tPAsh treatment on post-stroke neurological recovery, we performed modified neurological severity scores, modified adhesive removal, and beam walking tests at regular intervals until 14 days reperfusion. Results: Endo-tPA protein expression in the ischemic brain of rats was increased until 5 days reperfusion. Treatment with tPAsh formulation reduced the protein expression of tPA in the ischemic brain of rats and thereby demonstrated the in vivo efficiency of plasmids expressing tPA shRNAs. The MCAO procedure-induced mortality is comparable across the cohorts. Specific knockdown of endo-tPA reduced the infarct volume and facilitated the post-stroke neurological recovery. Conclusions: Preventing the induction of post-stroke endo-tPA levels in rat brain attenuates ischemic brain damage and improves post-stroke neurological recovery.