MicroRNA-146a在神经病理性疼痛中的作用和调控机制

Neuropathic pain is difficult to be treated in clinic. The current drug therapies offer very limited pain relief. Recent studies have implicated that microRNAs could induce the post-transcriptional gene silencing and are involved in several biological processes and disorders including chronic pain. Although several microRNAs have been revealed to be involved in pain regulation, they are quite limited and the undying mechanisms are not clear. MiR-146a was reported to be important in peripheral immune and inflammation regulation. Accumulating evidence support that glial cells mediated-neuroinflammation are important in chronic pain. Our previous studies showed that the expression of miR-146a is significantly upregulated in spinal cord astrocytes in spinal nerve ligation (SNL)-induced neuropathic pain model. In this project, we will investigate the role and the regulation mechanism of miR-146a in neuropathic pain, which includes: (1) The expression and cellular localization of miR-146a in the spinal cord in SNL mice; (2) The upstream and downstream signaling mechanisms of miR-146a; (3) The effect of regulation of miR-146a expression on the development and maintenance of neuropathic pain. This project will reveal a novel mechanism of miR-146a in neuropathic pain and provide new strategy for the development of novel analgesics.

神经病理性疼痛是临床镇痛的难点，目前药物治疗效果不佳。研究显示，MicroRNA诱导转录后基因沉默，参与多种生理和病理过程，包括慢性疼痛。但目前已知的参与疼痛调节的microRNA很有限且机制不清楚。miR-146a是参与外周免疫和炎症调节的重要microRNA；同时证据支持胶质细胞介导的神经炎症反应在慢性疼痛中起重要作用。我们前期结果表明，脊神经结扎(SNL )诱导的神经病理性疼痛小鼠脊髓星形胶质细胞中miR-146a表达上调。本项目将对miR-146a在神经病理性疼痛中的作用及机制进行深入研究，包括（1）miR-146a在SNL小鼠脊髓的表达时程和细胞定位；（2）调控miR-146a表达的上游分子和miR-146a作用的下游信号通路；（3）调节miR-146a的表达对神经病理性疼痛产生和维持的作用。本研究不但能阐明miR-146a参与神经病理性疼痛的新机制，也为开发新的镇痛药提供思路。

神经损伤引起的神经病理性疼痛是当今世界一个重要的临床问题，目前尚缺乏有效的镇痛药。其病理生理机制尚不十分清楚。胶质介导的神经炎症在神经性疼痛的发病机制中起着重要的作用。已经证实，MicroRNA (miR)-146a在受到IL-1β和LPS刺激后表达，并且下调其靶基因TNF受体相关因子 6 (TRAF6)和IL-1受体相关的激酶1 (IRAK1)，从而抑制单核细胞、巨噬细胞和星形胶质细胞中的炎症反应。在本研究中，我们研究了miR-146a-5p (miR-146a的成熟形式)及其靶基因TRAF6在脊神经结扎(SNL)诱导的神经性疼痛中的作用。我们发现，TRAF6通过脊髓星形胶质细胞中JNK/CCL2通路调节SNL诱导的机械性痛觉过敏。用TNF-α、IL-1β或LPS孵育培养的星形胶质细胞后，TRAF6 mRNA表达水平的下降与miR-146a-5p表达水平的增加有关。MiR-146a-5p通过与TRAF6 3 ' -UTR结合来抑制TRAF6的表达。此外，在星形胶质细胞中miR-146a-5p模拟物降低LPS诱导的TRAF6/pJNK/CCL2表达。重要的是，LPS增强了c-Jun与miR-146a-5p和ccl2启动子反应元件的结合，从而增加了miR-146a-5p和ccl2的转录活性。此外，TRAF6 siRNA或JNK抑制剂SP600125显著降低了星形胶质细胞中LPS诱导的miR-146a-5p增加。在小鼠鞘内给予miR-146a-5p模拟物能缓解TNF-α或LPS起的机械性痛觉过敏，并减少TRAF6的表达。最后，在SNL早期阶段(3天) miR-146a-5p的表达没有发生改变，但是在维持阶段(10和21天)表达增加。在脊髓中过表达的miR-146a-5p通过减少脊髓TRAF6的表达，对SNL诱导的机械性痛觉过敏产生镇痛作用。综上所述，我们的结果表明SNL增加TNF-α、IL - 1β、TLR4的表达，随后激活TRAF6/JNK/CCL2通路，诱导神经病理性疼痛的发生。同时，TRAF6/JNK通路的激活引起脊髓miR-146a-5p表达的上调。升高的MiR-146a-5p对TRAF6表达有靶向负调控作用。外源性增加miR-146a-5p可通过抑制TRAF6信号传递减弱。因此，miR-146a-5p可作为神经病理性疼痛干预治疗的新靶点。