SUR1-TRPM4通道介导的小胶质细胞焦亡在心肺复苏后脑损伤的作用及其机制研究

Post-cardiopulmonary resuscitation (post-CPR) brain injury is the leading cause of death and disability in patients successfully resuscitated from cardiac arrest, but the mechanism remains largely unknown. In recent years, cell pyroptosis and its subsequent inflammatory reaction has been found to play important roles in various types of brain injury. Recent researches and our preliminary study strongly suggested that microglia underwent caspase-1-dependent pyroptosis after CPR, and inhibition of microglial pyroptosis alleviated neuronal injury. We previously have shown that the sulfonylurea receptor 1-transient potential receptor cation channel subfamily M member 4 (SUR1-TRPM4) channel, which may participate in post-CPR brain injury, was expressed in microglia. The opening of SUR1-TRPM4 channel could induce cell lysis and microvesicle shedding, two morphological features similar to cell pyroptosis. Moreover, the influx of sodium caused by opening of SUR1-TRPM4 channel might activate cellular caspase-1, presumably by increasing the generation of reactive oxygen species and inducing the formation of inflammasome. Thus, we speculate that the SUR1-TRPM4 channel may cause post-CPR brain injury through mediation of caspase-1-dependent microglial pyroptosis. By using experimental methods including gene editing and pharmacological intervention, in this study, we will further verify the role of microglial pyroptosis in post-CPR brain injury and the mechanism under SUR1-TRPM4 channel-mediated microglial pyroptosis. This study will provide experimental evidence for the exploration of neuroprotective strategies targeting microglial pyroptosis in treating post-CPR brain injury.

脑损伤是心脏骤停后心肺复苏成功患者的重要致死、致残原因，但其发生机制尚不完全明确。近年来发现细胞焦亡及其引起的炎性反应在多种脑损伤中起重要作用，国外研究及我们的预实验提示心肺复苏后小胶质细胞可能发生caspase-1依赖性焦亡，且抑制后可减轻神经元损伤。我们前期发现与心肺复苏后脑损伤有关的SUR1-TRPM4通道可在小胶质细胞上表达。该通道开放不仅可使细胞发生类似焦亡的形态改变（裂解和出泡），其导致的钠离子内流还可能通过增加活性氧产生等途径诱导炎症小体形成而激活细胞内的caspase-1。由此，我们推测SUR1-TRPM4通道介导了caspase-1依赖性小胶质细胞焦亡，进而参与心肺复苏后脑损伤。本项目将利用基因编辑及药物干预等方法，进一步验证小胶质细胞焦亡在心肺复苏后脑损伤的作用及SUR1-TRPM4通道介导小胶质细胞焦亡的分子机制，为靶向小胶质细胞焦亡治疗心肺复苏后脑损伤提供实验依据。

细胞焦亡及其引起的炎性反应在多种脑损伤中起重要作用，国外研究及我们的预实验提示心肺复苏后小胶质细胞可能发生caspase-1依赖性焦亡，且抑制后可减轻神经元损伤。我们前期发现与心肺复苏后脑损伤有关的SUR1-TRPM4通道可在小胶质细胞上表达。该通道开放不仅可使细胞发生类似焦亡的形态改变（裂解和出泡），其导致的钠离子内流还可能通过增加活性氧产生等途径诱导炎症小体形成而激活细胞内的caspase-1。本项目研究SUR1-TRPM4通道是否介导了caspase-1依赖性小胶质细胞焦亡，进而参与心肺复苏后脑损伤。本项目发现：（1）SUR1-TRPM4通道选择性抑制剂格列本脲减轻心脏骤停/心肺复苏造模大鼠脑水肿；（2）SUR1-TRPM4通道选择性抑制剂格列美脲与格列本脲在短暂性脑缺血后具有相似的保护作用，而且其保护作用在Trpm4基因敲除小鼠中消失，说明格列本脲等磺脲类药物主要通过阻滞SUR1-TRPM4通道起作用；（3）NLRP3/Caspase-1依赖的小胶质细胞焦亡参与介导心肺复苏后脑损伤，选择性抑制NLRP3或Caspase-1均可通过减少小胶质细胞焦亡而减轻心肺复苏后脑损伤；（4）SUR1-TRPM4通道介导心肺复苏后小胶质细胞焦亡，使用通道阻滞剂格列本脲或敲除Trpm4基因均可减少小胶质细胞焦亡而减轻心肺复苏后脑损伤；（5）SUR1-TRPM4通道主要通过改变膜电位、打开下游的K+通道和放大K+外流信号激活NLRP3/Caspase-1信号通路，进而引发小胶质细胞焦亡。这些结果提示：SUR1-TRPM4通过激活NLRP3/caspase-1信号通路介导小胶质细胞焦亡，进而加剧心肺复苏后脑损伤。