Abstract：
BACKGROUND: We previously reported that the HMGB1/TLR4 axis promoted inflammation during the acute phase of intracerebral hemorrhage. Given that this phase is known to involve neuronal pyroptosis and neuroinflammation, here we explore whether HMGB1/TLR signaling activate inflammasome and pyroptosis after intracerebral hemorrhage.
METHODS: Autologous blood was injected into Sprague-Dawley rats to induce intracerebral hemorrhage. Neurological deficits were assessed using a modified neurological severity score. These expression and localization of NLRP1 and NLRP3 inflammasomes, as well as the levels of pyroptosis and pyroptosis-associated proteins were assessed using Western blot or immunocytochemistry. These experiments were repeated in animals that received treatment with short interfering RNAs against NLRP1 or NLRP3, with HMGB1 inhibitor ethyl pyruvate or TLR4 inhibitor TAK-242.
RESULTS: Intracerebral hemorrhage upregulated NLRP1 and NLRP3 in the ipsilateral striatum and increased the proportions of these cells that were pyroptosis-positive. Additionally, the levels of caspase protein family (e.g., pro-caspase-1 and caspase-1), apoptosis-associated speck-like protein (ASC), pro-interleukin-1β (IL-1β), and IL-1β were also elevated. These effects on pyroptosis and associated neurological deficit, were partially reversed by knockdown of NLRP1 or NLRP3, or by inhibition of HMGB1 or TLR4. Inhibition of HMGB1 or TLR4 resulted in the downregulation NLRP3 but not NLRP1.
CONCLUSIONS: The HMGB1/TLR4 signaling may activate the NLRP3 inflammasome during the acute phase of intracerebral hemorrhage, resulting in the inflammatory process known as pyroptosis. These insights suggest potential therapeutic targets for the mitigation tissue injury and associated neurological deficits following hemorrhagic stroke.
METHODS: Autologous blood was injected into Sprague-Dawley rats to induce intracerebral hemorrhage. Neurological deficits were assessed using a modified neurological severity score. These expression and localization of NLRP1 and NLRP3 inflammasomes, as well as the levels of pyroptosis and pyroptosis-associated proteins were assessed using Western blot or immunocytochemistry. These experiments were repeated in animals that received treatment with short interfering RNAs against NLRP1 or NLRP3, with HMGB1 inhibitor ethyl pyruvate or TLR4 inhibitor TAK-242.
RESULTS: Intracerebral hemorrhage upregulated NLRP1 and NLRP3 in the ipsilateral striatum and increased the proportions of these cells that were pyroptosis-positive. Additionally, the levels of caspase protein family (e.g., pro-caspase-1 and caspase-1), apoptosis-associated speck-like protein (ASC), pro-interleukin-1β (IL-1β), and IL-1β were also elevated. These effects on pyroptosis and associated neurological deficit, were partially reversed by knockdown of NLRP1 or NLRP3, or by inhibition of HMGB1 or TLR4. Inhibition of HMGB1 or TLR4 resulted in the downregulation NLRP3 but not NLRP1.
CONCLUSIONS: The HMGB1/TLR4 signaling may activate the NLRP3 inflammasome during the acute phase of intracerebral hemorrhage, resulting in the inflammatory process known as pyroptosis. These insights suggest potential therapeutic targets for the mitigation tissue injury and associated neurological deficits following hemorrhagic stroke.
摘要：
背景：我们先前报道了HMGB1/TLR4轴在脑出血急性期促进炎症。鉴于这一阶段已知涉及神经元焦亡和神经炎症，在此,我们探讨HMGB1/TLR信号是否激活脑出血后的炎性小体和焦亡.
方法：Sprague-Dawley大鼠注射自体血诱发脑出血。使用改良的神经系统严重程度评分评估神经系统缺陷。这些NLRP1和NLRP3炎性体的表达和定位，以及使用Westernblot或免疫细胞化学评估焦亡和焦亡相关蛋白的水平.在接受针对NLRP1或NLRP3的短干扰RNA、HMGB1抑制剂丙酮酸乙酯或TLR4抑制剂TAK-242治疗的动物中重复这些实验。
结果：脑出血上调同侧纹状体中的NLRP1和NLRP3，并增加这些细胞的比例。此外,caspase蛋白家族的水平(例如，pro-caspase-1和caspase-1），凋亡相关斑点样蛋白(ASC)，前白细胞介素-1β（IL-1β），IL-1β也升高。这些对焦亡和相关神经功能缺损的影响，通过敲低NLRP1或NLRP3或通过抑制HMGB1或TLR4而部分逆转。HMGB1或TLR4的抑制导致NLRP3而非NLRP1的下调。
结论：HMGB1/TLR4信号可能在脑出血急性期激活NLRP3炎性体，导致称为焦亡的炎症过程。这些见解提出了减轻出血性中风后组织损伤和相关神经功能缺损的潜在治疗目标。
方法：Sprague-Dawley大鼠注射自体血诱发脑出血。使用改良的神经系统严重程度评分评估神经系统缺陷。这些NLRP1和NLRP3炎性体的表达和定位，以及使用Westernblot或免疫细胞化学评估焦亡和焦亡相关蛋白的水平.在接受针对NLRP1或NLRP3的短干扰RNA、HMGB1抑制剂丙酮酸乙酯或TLR4抑制剂TAK-242治疗的动物中重复这些实验。
结果：脑出血上调同侧纹状体中的NLRP1和NLRP3，并增加这些细胞的比例。此外,caspase蛋白家族的水平(例如，pro-caspase-1和caspase-1），凋亡相关斑点样蛋白(ASC)，前白细胞介素-1β（IL-1β），IL-1β也升高。这些对焦亡和相关神经功能缺损的影响，通过敲低NLRP1或NLRP3或通过抑制HMGB1或TLR4而部分逆转。HMGB1或TLR4的抑制导致NLRP3而非NLRP1的下调。
结论：HMGB1/TLR4信号可能在脑出血急性期激活NLRP3炎性体，导致称为焦亡的炎症过程。这些见解提出了减轻出血性中风后组织损伤和相关神经功能缺损的潜在治疗目标。
