自噬-NRF2信号在神经病理性疼痛发生发展中的作用及调控机制

Neuropathic pain has high morbidity and low cure rates, while the pathogenesis of neuropathic pain remains obscure. Transcription factor NRF2 has antinociceptive effects against inflammatory pain due to its role of anti-inflammation and anti-oxidative stress. However, little is known about whether NRF2 regulates neuropathic pain. Some studies have found that autophagy-adaptor protein P62 promotes nuclear translocation of NRF2 and induces expression of NRF2 targets. These studies prompt that autophagy might regulate activation of NRF2 signaling. In our preliminary study, we found that the paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) were significantly decreased after spinal nerve ligation (SNL), and these change were accompanied by inhibited autophagy and increased protein level and nuclear translocation of NRF2 in the ipsilateral spinal cord. Inhibition of autophagy with 3-MA, a widely used autophagy inhibitor, could further increase the expression and nuclear translocation of NRF2 and reduce the production of inflammatory cytokines. Moreover, both PWT and PWL of NRF2 knockout mice were less than the WT mice’s after SNL, while the expression of inflammatory cytokines was increased. Therefore, we hypothesized that autophagy may regulate neuropathic pain by affecting the activation of NRF2 pathway. In this project, we will study the role and the molecular mechanism of autophagy-NRF2 pathway in the development of neuropathic pain with the NRF2 knockout mice, which may provide a new mechanism hypothesis and a potential therapeutic target of neuropathic pain.

神经病理性疼痛发病率高，疗效不佳，发病机制不清。转录因子NRF2具有抗炎、抗氧化应激和神经保护作用，参与炎性疼痛调控，但NRF2是否调控神经病理性疼痛尚不明确。研究发现自噬底物蛋白P62调控NRF2活化，发挥抗氧化应激作用，鉴于P62蛋白与自噬过程密切相关，提示自噬可能调控NRF2活化。我们预实验结果发现脊神经结扎后，小鼠疼痛阈值降低，脊髓自噬水平减弱，NRF2总蛋白表达及核转移增加；鞘内注射自噬抑制剂抑制自噬，NRF2总蛋白表达及核转移较对照组显著增加，炎性因子表达降低；利用NRF2敲除小鼠行脊神经结扎，其疼痛阈值较野生型小鼠降低而炎性因子表达增加。因此，我们推测自噬可能通过影响NRF2通路的活化来调控神经病理性疼痛。本项目中，我们将利用NRF2基因敲除小鼠来系统研究自噬-NRF2通路调控神经病理性疼痛的作用及具体分子信号机制，为神经病理性疼痛的发生发展提供新的机制解说和潜在治疗靶点。

自噬是一个进化保守的过程，在调节免疫炎症反应和神经系统稳态中发挥重要作用。然而，自噬在疼痛中的确切作用和机制尚不清楚。本课题集中研究自噬-NRF2通路调节神经病理性疼痛的相关机制。我们发现在神经病理性疼痛的维持过程中，自噬通量受损主要发生在星形胶质细胞中。无论是神经性疼痛的诱导阶段还是维持阶段，激活自噬可以减轻疼痛水平，而抑制自噬则加重了疼痛程度。此外，抑制或者激活自噬可分别导致神经炎症和活性氧水平升高或降低。进一步研究表明，抑制自噬导致神经炎症反应启动减缓但维持时间延长，这可能是通过促进肿瘤坏死因子受体相关因子6(TRAF6)与K63 泛素化蛋白结合，并增加有丝分裂原活化蛋白激酶-8(p-MAPK8/JNK)和NFKB/NF-κB磷酸化水平实现的。自噬受损也降低了星形胶质细胞对神经元活性氧应激的保护作用，这主要是由抑制自噬后星形胶质细胞释放谷胱甘肽水平下降介导的。通过激活抗氧化经典通路NFE2L2/NRF2通路可以改善自噬受损的星形胶质细胞对神经元的保护作用。我们还证明，与单独激活自噬相比，同时激活自噬和NFE2L2通路可进一步缓解疼痛。本研究提供了自噬参与神经病理性疼痛调控的潜在机制，结合NFE2L2激活可能成为神经病理性疼痛治疗的新途径。