TLR7通过AMPA受体调节树突棘密度促进脓毒症相关性脑病的作用及机制研究

Sepsis-associated encephalopathy (SAE) has a high incidence and decreases the survival rate and quality of life of patients with sepsis. Neuron dysfunction is critical for behavioral changes in SAE. Furthermore, dendritic spines density reflects the storage and transfer capabilities of neuron. Toll-like receptor 7, one of the damage-associated molecular pattern recognition receptor, is consistently expressed in neuron. We found that the depletion of TLR7 gene can improve the survival rate of septic mice and their acute neurobehavioral score. Activation of TLR7 promotes neuroinflammation. Moreover, other studies have shown that NF-κB has an impact on AMPAR function. Therefore, we speculate that TLR7 may affect the dendritic spines density through NF-κB and AMPAR in SAE. In this study, we will activate TLR7 and sacrifice TLR7 gene depletion mice to discover: (1) whether TLR7 contributes to the regulation of dendritic spines density in polymicrobial sepsis;(2) whether TLR7 affects the expression of AMPAR through NF-κB. This study will provide new theoretical foundation for targeting TLR7 in the treatment of sepsis-associated encephalopathy.

脓毒症相关性脑病（SAE）发病率高，降低脓毒症患者生存率及生活质量。神经元功能受损是SAE行为学改变核心，而树突棘密度反映神经元存储和信息传递能力。Toll样受体7是损伤相关分子模式识别受体，且持续性地表达于神经元。我们发现TLR7基因缺失可改善脓毒症小鼠生存率及急性期神经行为学评分，激活TLR7加剧神经元炎症反应。另有研究表明疾病状态下NF-κB参与调节树突棘膜受体AMPAR活性。由此我们推测：机体发生脓毒症时，神经元TLR7激活后可能通过下游NF-κB通路调节AMPAR表达，影响树突棘密度，参与SAE进程。本课题拟通过TLR7受体激活或缺失，（1）从整体、器官和细胞水平探讨TLR7是否调节树突棘密度改善SAE；（2）探索TLR7是否通过下游NF-κB通路影响AMPAR表达，调节树突棘密度。本研究将为TLR7作为SAE干预靶点提供新的理论依据。

脓毒症相关性脑病（SAE）影响患者近远期死亡率及预后，其特征为弥散性脑功能紊乱，诸多方法疗效欠佳。神经元形态及功能改变是SAE行为学改变的核心，树突棘是认知功能的结构基础。Toll样受体7是损伤相关分子模式识别受体，TLR7基因缺失可改善脓毒症小鼠生存率。但是SAE否通过TLR7影响神经元形态及功能尚未明确。本研究从在体研究、RNA测序及分析以及体外细胞水平三个层面进行研究，观察到脓毒症小鼠总体运动距离和平均速度显著下降，停滞时间显著延长，提示小鼠脑功能紊乱。SAE对小鼠神经元树突棘新生率没有显著影响，但树突棘消失率明显增加。SAE小鼠神经元自发性钙信号显著减少，影响运动皮层神经元钙信号发放次数和神经网络，但不影响神经细胞单次发放的动力过程。RNA-Seq检测及分析表明小鼠运动皮层区域与免疫/炎症反应相关的信号通路被显著富集，Toll样受体信号通路相关基因明显上调。此外，我们体外实验发现激活/阻断miR-146a-TLR7通路可影响神经元细胞HT-22炎症因子表达、细胞活力和Cleaved caspase-3表达水平。上述结果揭示脓毒症相关性脑病对大脑皮层神经元树突棘结构、钙信号的影响和机制，并在体外实验证实抑制TLR7上下游通路对对脓毒症相关性脑病神经元的影响，为临床治疗提供新的思路，具有一定应用价值。本研究2篇论文在投，数篇论著待审待发表，协助培养硕士研究生1名，实验技术员1名。