From the College of Bioengineering, Chongqing University, China (X.C., D.H.).
Research Institute of Surgery/Daping Hospital (X.W., Y.T., L.Y., Y.P., H.T., R.L., W.B., L.L.), Third Military Medical University, Chongqing, China.
Stroke. 2018 Oct;49(10):2473-2482. doi: 10.1161/STROKEAHA.117.019253.
Background and Purpose- Accumulated evidence suggests that hemin-a breakdown product of hemoglobin-plays a pivotal role in the inflammatory injuries that result after hemorrhagic stroke through the Toll Like Receptor 2-Toll Like Receptor 4 signal pathway. However, the mechanism of how hemin triggers neuronal necroptosis directly after intracranial hemorrhage (ICH) is still an area of active research. As animal model and preclinical studies have shown, the recombinant interleukin-1 receptor antagonist (IL-1RA) improves clinical outcomes after stroke. As such, we have chosen to investigate the mechanism of how IL-1RA exerts protective effect in hemin-induced neuronal necroptosis after ICH. Methods- Our ICH model was induced by hemin injection in C57BL/6 mice and IL-1R1 mice. In addition, we used primary cultured neurons to assess hemin-induced cell death. Co-immunoprecipitation, immunoblot, immunofluorescent staining, neurological deficit scores, and brain water content were used to study the mechanisms of IL-1R1 modulation in neuronal necroptosis both in vitro and in vivo. Results- Free hemin could mediate neuronal necroptosis directly by assembling necrosome complex and then to trigger cell death. This phenomenon was driven by IL-1R1 as IL-1R1 can form a complex with necrosome. After treatment with IL-1RA, both the expression and translocation of the necrosome decreased while disruption of the interaction between IL-1R1 and RIP1/RIP3 (receptor interacting protein 1/3) increased neuron survival. In addition, the IL-1R1-deficient mice demonstrated lower levels of necrosome components, including RIP1, RIP3, and MLKL (mixed lineage kinase domain-like protein), compared with control groups after hemin treatment. In addition, the neurological deficit scores, brain water content, and inflammatory response were all also reduced in the IL-1R1-deficient mice. Conclusions- Functional inhibition of the interaction between IL-1R1 and the necrosome complex improves neuron survival and promotes the recovery of neurological function in experimental ICH. Targeting IL-1R1/RIP1/RIP3 assembly could be a promising therapeutic strategy for patients with ICH.
背景与目的-有证据表明，血红素（血红蛋白的降解产物）通过 Toll 样受体 2-4 信号通路在出血性中风后的炎症损伤中发挥关键作用。然而，血红素在颅内出血（ICH）后直接引发神经元坏死性凋亡的机制仍然是一个活跃的研究领域。动物模型和临床前研究表明，重组白细胞介素-1 受体拮抗剂（IL-1RA）改善中风后的临床结果。因此，我们选择研究 IL-1RA 在血红素诱导的 ICH 后神经元坏死性凋亡中发挥保护作用的机制。方法-我们通过血红素注射诱导 C57BL/6 小鼠和 IL-1R1 小鼠的 ICH 模型。此外，我们使用原代培养神经元来评估血红素诱导的细胞死亡。共免疫沉淀、免疫印迹、免疫荧光染色、神经功能缺损评分和脑水含量用于研究 IL-1R1 在体外和体内神经元坏死性凋亡中的调节机制。结果-游离血红素可以通过组装坏死体复合物直接介导神经元坏死性凋亡，然后引发细胞死亡。这种现象是由 IL-1R1 驱动的，因为 IL-1R1 可以与坏死体形成复合物。用 IL-1RA 处理后，坏死体的表达和易位减少，而 IL-1R1 与 RIP1/RIP3（受体相互作用蛋白 1/3）的相互作用中断则增加神经元的存活。此外，与对照组相比，血红素处理后，IL-1R1 缺陷型小鼠的坏死体成分（包括 RIP1、RIP3 和 MLKL（混合谱系激酶结构域样蛋白））水平较低。此外，IL-1R1 缺陷型小鼠的神经功能缺损评分、脑水含量和炎症反应也均降低。结论-抑制 IL-1R1 与坏死体复合物的相互作用可改善神经元存活并促进实验性 ICH 后神经功能的恢复。靶向 IL-1R1/RIP1/RIP3 组装可能是 ICH 患者有希望的治疗策略。
