Abstract WMP75: A Modified, Recombinant Tissue Plasminogen Activator-Responding Photothrombotic Stroke Model

Abstract

Introduction: Photodynamical dye-based stroke models (e.g. Rose Bengal RB/photothrombosis) have several attractive features, including simple surgical procedures, consistent infarct size, and low mortality rate. Yet, because RB/photothrombosis mainly induces platelet-rich but fibrin-poor clots, it responds to recombinant tissue plasminogen activator (rtPA)-based lytic therapy poorly making it less suitable for studying reperfusion injury or improving lytic therapy. Hypothesis: We hypothesize the addition of thrombin to Rose Bengal dye in photothrombosis will increase the fibrin component in clots, making it more susceptible to rtPA-thrombolysis. Methods: Based on the standard RB/photothrombotic stroke procedures (intravenous injection of 50 mg/kg RB, we mixed a subthreshold level of bovine thrombin with RB to create a thrombin-Rose Bengal (TRB)/photothrombotic stroke model. We compared the clot composition, effects of rtPA treatment on cerebral blood flow reperfusion, and the infarct size following rtTA treatment initiated at 30, 60, or 120 min after photothrombosis. Results: We report four sets of results. First, the addition of thrombin did not increase mortality rate or infarct size, when compared to standard RB/photothrombosis. Second, immunostaining showed a scant amount of fibrin at the border of RB/photothrombosis-induced clots. In contrast, fibrin was mixed with erythrocytes throughout TRB/photothrombosis-induced clots. Third, laser speckle contrast imaging indicated that intravenous infusion of rtPA (10 mg/kg) at 30 min after TRB/photothrombosis improved blood flow in the MCA-supplied area, but the same treatment had little effect in RB/photothrombosis. Finally, the therapeutic window of rtPA-treatment is at least 2 h in TRB/photothrombosis (n>10 in each group). Conclusions: Our results confirm the notion by Dr. Brant Watson et al. that rtPA lytic treatment confers little benefits in RB/photothrombosis, likely due to its fibrin-poor clots. Yet, the addition of subthreshold thrombin (TRB/photothrombosis) elevates the fibrin component in clot, making it far more responsive to rtPA-thrombolysis. The modified, rtPA-responding thrombotic stroke model may have broad applications in experimental stroke research.