DNMT3a介导的DRG Kcna2甲基化修饰在神经病理性疼痛中的作用及其机制

Nerve injury-induced neuropathic pain is a major public health problem. Its treatment is highly unsatisfactory in part due to uncompleted understanding the mechanisms underlying neuropathic pain gensis. Nerve injury-induced downregulation of voltage-dependent ion channels Kv1.2 mRNA and protein in dorsal root ganglion (DRG) contributes to the induction and maintenanace of neuropathic pain, but how this downregulation occurs is still elusive. Our pilot study showed that peripheral nerve injury increased the levels of total DNA methylation and DNMT3a protein expression (but not DNMT3b) in DRG. Additionally, DRG DNMT3a could bind to the promoter region of the Kv1.2 gene. These findings allow us to hypothesize that the increased DNMT3a binds to the CpG sites of Kv1.2 gene promoter region, resulting in an increase in its DNA methylation and a consequent silence of Kv1.2 expression in the DRG. This process may be involved in neuropathic pain development. This proposal will carry out both in vivo and in vitro preclinical models and investigate whether and how DNMT3a participates in nerve injury-induced DRG Kv1.2 downregulation. The proposed study may provide a new therapeutic target in neuropathic pain management.

神经病理性疼痛（NPP）是重要的公共卫生问题，发病机制尚不完全清楚，临床治疗效果差。外周神经损伤后，背根神经节（DRG）神经元钾离子通道Kv1.2表达下降，诱发其异常放电，是NPP外周敏化的重要机制之一，但其调控尚不清楚。DNA甲基化通过转录抑制，调控基因表达。我们在大鼠脊神经结扎（SNL）模型中证实，DRG中DNA甲基转移酶3a(DNMT3a)表达上调，DNA甲基化增强，并且发现DNMT3a和Kv1.2基因（Kcna2）启动子区结合。据此，我们提出假说：DNMT3a通过催化Kcna2 启动子区CpG岛（CG序列密集区）高度甲基化，抑制其转录，调控Kv1.2表达，参与NPP的发病过程。本研究通过构建离体、在体模型，从分子、细胞、整体动物水平，探索DNMT3a介导DNA甲基化在外周神经损伤引起的DRG神经元Kv1.2表达下降中的作用及其机制，为寻找治疗NPP的有效药物提供作用靶点和理论依据。

神经损伤诱导背根神经节（DRG）神经元中基因转录的变化，这可能导致神经损伤诱导的神经性疼痛，DNA甲基化可以抑制基因表达。我们研究指出周围神经损伤通过激活转录因子八聚体转录因子1增加受损DRG神经元中DNA甲基转移酶DNMT3a的表达。同时，阻断这种增加可防止神经损伤诱导的电压依赖性钾（Kv）通道亚基Kcna2启动子区域的甲基化，并挽救受损DRG中的Kcna2表达并减轻神经性疼痛。相反，在没有神经损伤的情况下，模仿这种增加会降低Kcna2启动子活性，减少Kcna2表达，降低Kv电流，增加DRG神经元的兴奋性并导致脊髓中枢致敏和神经性疼痛症状。这些发现表明，DNMT3a可能通过抑制DRG中的Kcna2表达而导致神经性疼痛。