IMM-H004 therapy for permanent focal ischemic cerebral injury via CKLF1/CCR4-mediated NLRP3 inflammasome activation
IMM-H004 therapy for permanent focal ischemic cerebral injury via CKLF1/CCR4-mediated NLRP3 inflammasome activation
复制标题
IMM-H004 通过 CKLF1/CCR4 介导的 NLRP3 炎症小体激活治疗永久性局灶性缺血性脑损伤
DOI：
10.1016/j.trsl.2019.05.007
复制
发表时间：
2019-10-01
影响因子：
7.8
通讯作者：
Chen, Naihong
中科院分区：
文献类型：
--
作者：
Ai, Qd;Chen, Chen;Chen, Naihong
趋化因子样因子1（CKLF1）是缺血性脑卒中治疗的一个潜在靶点。核苷酸结合寡聚化结构域样受体蛋白3（NLRP3）炎症小体被认为在缺血/再灌注（I/R）损伤过程中介导炎症反应。化合物IMM - H004是一种新型香豆素衍生物，能够改善脑I/R损伤。本研究旨在探究IMM - H004对缺血性脑卒中损伤的影响，并进一步阐明其分子机制。本文采用标准的大脑中动脉永久闭塞（pMCAO）局灶性缺血模型。在缺血6小时后给药，于缺血9小时后进行行为学评估、实施安乐死并评估各项结果指标。通过2,3,5 - 三苯基氯化四氮唑（TTC）染色、行为学测试、酶联免疫吸附试验（ELISA）和尼氏染色来确定IMM - H004对缺血性脑卒中损伤的作用。采用免疫组织化学染色、免疫荧光染色、定量逆转录聚合酶链反应（qPCR）、蛋白质免疫印迹法（western blotting）以及免疫共沉淀（CO - IP）试验来阐明潜在机制。在大鼠中，IMM - H004治疗通过依赖CKLF1的抗炎途径对缺血性脑卒中提供了显著的保护作用。IMM - H004下调了CKLF1与C - C趋化因子受体4型的结合量，进一步抑制了NLRP3炎症小体的激活及后续的炎症反应，最终对缺血脑组织起到保护作用。这项临床前研究证实了IMM - H004作为永久性脑缺血潜在治疗药物的有效性。这些结果为进一步研发IMM - H004用于急性脑缺血的人体临床试验提供了支持，尤其适用于那些不适合再灌注治疗的患者。
Chemokine-like factor 1 (CKLF1) is a potential target for ischemic stroke therapy. The NOD-like receptor protein 3 (NLRP3) inflammasome has been postulated to mediate inflammatory responses during ischemic/reperfusion (I/R) injury. The compound IMM-H004 is a novel coumarin derivative that can improve cerebral I/R injury. This study aims to investigate the effects of IMM-H004 on ischemia stroke injury and further elucidate the molecular mechanisms. The standard pMCAO model of focal ischemia was used in this paper. Drugs were administered at 6 hours after ischemia, and behavioral assessment, euthanasia, and outcome measures were evaluated at 9 hours after ischemia. The effects of IMM-H004 on ischemic stroke injury were determined using 2,3,5-triphenyltetrazolium chloride (TIC) staining, behavioral tests, enzyme-linked immunosorbent assay (ELISA), and Nissl staining. Immunohistologic staining, immunofluorescence staining, quantitative RT-PCR (qPCR), western blotting, and coimmunoprecipitation (CO-IP) assays were used to elucidate the underlying mechanisms. IMM-H004 treatment provided significant protection against ischemia stroke through a CKLF1-dependent anti-inflammatory pathway in rats. IMM-H004 downregulated the amount of CKLF1 binding with C-C chemokine receptor type 4, further suppressing the activation of NLRP3 inflammasome and the following inflammatory response, ultimately protecting the ischemic brain. This preclinical study established the efficacy of IMM-H004 as a potential therapeutic medicine for permanent cerebral ischemia. These results support further efforts to develop IMM-H004 for human clinical trials in acute cerebral ischemia, particularly for patients who are not suitable for reperfusion therapy.