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

Abstract

BackgroundAcute 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 studys 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. MethodsThromboembolic 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. ResultsRt-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. ConclusionHere, we conclude that MMP-9/LOX-1 inhibition attenuates negative aspects of delayed rt-PA therapy leading to improved neurological function.