Department of Translational Neuroscience, Jing'an District Centre Hospital of Shanghai, State Key Laboratory of Medical Neurobiology-Ministry of Education (MOE) Frontiers Center for Brain Science, Institutes of Brain Science, Fudan University, Shanghai, China.
Blood. 2021 Jul 8;138(1):91-103. doi: 10.1182/blood.2020008913.
Intracerebral hemorrhage associated with thrombolytic therapy with tissue plasminogen activator (tPA) in acute ischemic stroke continues to present a major clinical problem. Here, we report that infusion of tPA resulted in a significant increase in markers of neutrophil extracellular traps (NETs) in the ischemic cortex and plasma of mice subjected to photothrombotic middle cerebral artery occlusion. Peptidylarginine deiminase 4 (PAD4), a critical enzyme for NET formation, is also significantly upregulated in the ischemic brains of tPA-treated mice. Blood-brain barrier (BBB) disruption after ischemic challenge in an in vitro model of BBB was exacerbated after exposure to NETs. Importantly, disruption of NETs by DNase I or inhibition of NET production by PAD4 deficiency restored tPA-induced loss of BBB integrity and consequently decreased tPA-associated brain hemorrhage after ischemic stroke. Furthermore, either DNase I or PAD4 deficiency reversed tPA-mediated upregulation of the DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS). Administration of cGAMP after stroke abolished DNase I-mediated downregulation of the STING pathway and type 1 interferon production and blocked the antihemorrhagic effect of DNase I in tPA-treated mice. We also show that tPA-associated brain hemorrhage after ischemic stroke was significantly reduced in cGas-/- mice. Collectively, these findings demonstrate that NETs significantly contribute to tPA-induced BBB breakdown in the ischemic brain and suggest that targeting NETs or cGAS may ameliorate thrombolytic therapy for ischemic stroke by reducing tPA-associated hemorrhage.
与急性缺血性脑卒中溶栓治疗组织型纤溶酶原激活剂（tPA）相关的脑出血仍然是一个主要的临床问题。在这里，我们报告称，tPA 输注会导致光血栓性大脑中动脉闭塞小鼠缺血皮质和血浆中中性粒细胞胞外诱捕网（NETs）的标志物显著增加。NET 形成的关键酶肽基精氨酸脱亚氨酶 4（PAD4）在 tPA 治疗小鼠的缺血脑中也显著上调。在体外血脑屏障（BBB）模型中，缺血性挑战后 BBB 破坏在暴露于 NETs 后加剧。重要的是，DNase I 破坏 NETs 或 PAD4 缺乏抑制 NET 产生可恢复 tPA 诱导的 BBB 完整性丧失，并因此减少缺血性脑卒中后 tPA 相关脑出血。此外，DNase I 或 PAD4 缺乏均可逆转 tPA 介导的 DNA 传感器环鸟苷酸-腺苷酸合酶（cGAS）的上调。中风后施用 cGAMP 可消除 DNase I 介导的 STING 途径和 I 型干扰素产生的下调，并阻止 DNase I 在 tPA 治疗小鼠中的抗出血作用。我们还表明，cGas-/- 小鼠缺血性脑卒中后 tPA 相关脑出血显著减少。总之，这些发现表明 NETs 可显著促进缺血性大脑中 tPA 诱导的 BBB 破裂，并表明靶向 NETs 或 cGAS 可能通过减少 tPA 相关出血来改善缺血性脑卒中的溶栓治疗。
