LOX-1 and MMP-9 inhibition attenuates the detrimental effects of delayed rt-PA therapy and improves outcomes after acute ischemic stroke

Background Acute ischemic stroke triggers endothelial activation that disrupts vascular integrity and increases hemorrhagic transformation leading to worsened stroke outcomes. Recombinant-tissue plasminogen activator (rt-PA) is an effective treatment; however, its use is limited due to a restricted time window and high risk for hemorrhagic transformation, which in part may involve activation of metalloproteinases (MMPs) mediated through lectin-like oxidized LDL receptor 1 (LOX-1). This study’s overall aim was to evaluate the therapeutic potential of novel MMP-9 and LOX-1 inhibitors in combination with rt-PA to improve stroke outcomes. Methods Thromboembolic rat stroke model was utilized to investigate the impact of rt-PA delivered 4h post-stroke onset as well as selective LOX-1 (BI-0115) and/or MMP-9 (JNJ0966) inhibitors given prior to rt-PA administration. Infarct size, perfusion, and hemorrhagic transformation were evaluated by MRI. Neurological function was assessed using sensorimotor functioning testing. Using an in vitro , human brain microvascular endothelial cell (HBMEC) model, cells were exposed to hypoxia plus glucose deprivation (3h)/reperfusion (12h) (HGD/R) and treated with rt-PA ± an MMP-9 and LOX-1 inhibition cocktail. MMP-9 activity was determined with zymography, and endothelial barrier marker gene expression and LOX-1 levels were evaluated via qRT-PCR and western blot respectively. Results Rt-PA treatment increased edema, hemorrhage, and worsened neurological outcomes post stroke. LOX-1 inhibition significantly improved neurological function and reduced edema after delayed rt-PA treatment. Hemorrhagic transformation, edema, and increased MMP-9 activity were attenuated by the MMP-9 inhibitor. Stroke induced increases in cerebrovascular LOX-1 expression correlated with increased MMP-9 activity and elevated activity correlated with increased edema, infarct volume, and decreased neurological function. In cultured HBMECs, LOX-1/MMP-9 inhibition differentially attenuated rt-PA-mediated increases in endothelial derived MMP-9 levels and activity, inflammation, and activation following HGD/R. Conclusion Here, we conclude that MMP-9/LOX-1 inhibition attenuates negative aspects of delayed rt-PA therapy leading to improved neurological function. ### Competing Interest Statement The authors have declared no competing interest.