Applied Neurovascular Research, Neurosurgery, Department of Clinical Sciences, Lund University, Sweden (K.A., H.A., A.A., S.A.).
Department of Basic Medical Sciences, University of Arizona College of Medicine, Phoenix, AZ (T.S.W., R.J.G.).
Circ Res. 2024 Apr 12;134(8):954-969. doi: 10.1161/CIRCRESAHA.123.323371. Epub 2024 Mar 19.
Acute ischemic stroke triggers endothelial activation that disrupts vascular integrity and increases hemorrhagic transformation leading to worsened stroke outcomes. rt-PA (recombinant tissue-type plasminogen activator) is an effective treatment; however, its use is limited due to a restricted time window and hemorrhagic transformation risk, which in part may involve activation of MMPs (matrix metalloproteinases) mediated through LOX-1 (lectin-like oxLDL [oxidized low-density lipoprotein] receptor 1). This study's overall aim was to evaluate the therapeutic potential of novel MMP-9 (matrix metalloproteinase 9) ± LOX-1 inhibitors in combination with rt-PA to improve stroke outcomes.
A rat thromboembolic stroke model was utilized to investigate the impact of rt-PA delivered 4 hours poststroke onset as well as selective MMP-9 (JNJ0966) ±LOX-1 (BI-0115) inhibitors given before rt-PA administration. Infarct size, perfusion, and hemorrhagic transformation were evaluated by 9.4-T magnetic resonance imaging, vascular and parenchymal MMP-9 activity via zymography, and neurological function was assessed using sensorimotor function testing. Human brain microvascular endothelial cells were exposed to hypoxia plus glucose deprivation/reperfusion (hypoxia plus glucose deprivation 3 hours/R 24 hours) and treated with ±tPA and ±MMP-9 ±LOX-1 inhibitors. Barrier function was assessed via transendothelial electrical resistance, MMP-9 activity was determined with zymography, and LOX-1 and barrier gene expression/levels were measured using qRT-PCR (quantitative reverse transcription PCR) and Western blot.
Stroke and subsequent rt-PA treatment increased edema, hemorrhage, MMP-9 activity, LOX-1 expression, and worsened neurological outcomes. LOX-1 inhibition improved neurological function, reduced edema, and improved endothelial barrier integrity. Elevated MMP-9 activity correlated with increased edema, infarct volume, and decreased neurological function. MMP-9 inhibition reduced MMP-9 activity and LOX-1 expression. In human brain microvascular endothelial cells, LOX-1/MMP-9 inhibition differentially attenuated MMP-9 levels, inflammation, and activation following hypoxia plus glucose deprivation/R.
Our findings indicate that LOX-1 inhibition and ± MMP-9 inhibition attenuate negative aspects of ischemic stroke with rt-PA therapy, thus resulting in improved neurological function. While no synergistic effect was observed with simultaneous LOX-1 and MMP-9 inhibition, a distinct interaction is evident.
急性缺血性中风会引发内皮细胞激活，破坏血管完整性并增加出血转化，导致中风预后恶化。重组组织型纤溶酶原激活剂（rt-PA）是一种有效的治疗方法；然而，由于时间窗有限和出血转化风险，其应用受到限制，部分原因可能涉及通过 LOX-1（凝集素样 oxLDL[氧化低密度脂蛋白]受体 1）介导的 MMPs（基质金属蛋白酶）的激活。本研究的总体目标是评估新型 MMP-9（基质金属蛋白酶 9）±LOX-1（BI-0115）抑制剂与 rt-PA 联合使用以改善中风预后的治疗潜力。
利用大鼠血栓栓塞性中风模型，研究 rt-PA 在中风发作后 4 小时给药以及在 rt-PA 给药前给予选择性 MMP-9（JNJ0966）±LOX-1（BI-0115）抑制剂的影响。通过 9.4-T 磁共振成像评估梗死面积、灌注和出血转化，通过酶谱法评估血管和实质 MMP-9 活性，通过感觉运动功能测试评估神经功能。将人脑微血管内皮细胞暴露于缺氧加葡萄糖剥夺/再灌注（缺氧加葡萄糖剥夺 3 小时/R 24 小时）中，并使用±tPA 和±MMP-9 ±LOX-1 抑制剂进行处理。通过跨内皮电电阻评估屏障功能，通过酶谱法测定 MMP-9 活性，通过 qRT-PCR（定量逆转录 PCR）和 Western blot 测定 LOX-1 和屏障基因表达/水平。
中风和随后的 rt-PA 治疗增加了水肿、出血、MMP-9 活性、LOX-1 表达，并使神经功能恶化。LOX-1 抑制改善了神经功能，减少了水肿，并改善了内皮屏障完整性。升高的 MMP-9 活性与水肿、梗死体积增加和神经功能下降相关。MMP-9 抑制降低了 MMP-9 活性和 LOX-1 表达。在人脑微血管内皮细胞中，LOX-1/MMP-9 抑制可降低缺氧加葡萄糖剥夺/R 后的 MMP-9 水平、炎症和激活。
我们的研究结果表明，LOX-1 抑制和±MMP-9 抑制减轻了 rt-PA 治疗缺血性中风的负面影响，从而改善了神经功能。尽管同时抑制 LOX-1 和 MMP-9 没有观察到协同作用，但明显存在相互作用。
