Guangdong Provincial Key Laboratory of Brain Function and Disease, Pain Research Center, Department of Physiology, Zhongshan Medical School, Sun Yat-Sen University, 74 Zhongshan Road 2, Guangzhou 510080, China.
State Key Laboratory of Oncology in South China, Department of Anesthesiology, Sun Yat-Sen University Cancer Center, Collaborative Innovation Center for Cancer Medicine, 651 Dongfeng Road East, Guangzhou 510060, China.
Int J Mol Sci. 2023 Oct 22;24(20):15454. doi: 10.3390/ijms242015454.
The hyperexcitability of the anterior cingulate cortex (ACC) has been implicated in the development of chronic pain. As one of the key causes of ACC hyperexcitation, disinhibition of the ACC may be closely related to the dysfunction of inhibitory parvalbumin (PV)-expressing interneurons (PV-INs). However, the molecular mechanism underlying the ACC PV-INs injury remains unclear. The present study demonstrates that spared sciatic nerve injury (SNI) induces an imbalance in the excitation and inhibition (E/I) of the ACC. To test whether tumor necrosis factor-α (TNF-α) upregulation in the ACC after SNI activates necroptosis and participates in PV-INs damage, we performed a differential analysis of transcriptome sequencing using data from neuropathic pain models and found that the expression of genes key to the TNF-α-necroptosis pathway were upregulated. TNF-α immunoreactivity (IR) signals in the ACCs of SNI rats were co-located with p-RIP3- and PV-IR, or p-MLKL- and PV-IR signals. We then systematically detected the expression and cell localization of necroptosis-related proteins, including kinase RIP1, RIP3, MLKL, and their phosphorylated states, in the ACC of SNI rats. Except for RIP1 and MLKL, the levels of these proteins were significantly elevated in the contralateral ACC and mainly expressed in PV-INs. Blocking the ACC TNF-α-necroptosis pathway by microinjecting TNF-α neutralizing antibody or using an siRNA knockdown to block expression of MLKL in the ACC alleviated SNI-induced pain hypersensitivity and inhibited the upregulation of TNF-α and p-MLKL. Targeting TNF-α-triggered necroptosis within ACC PV-INs may help to correct PV-INs injury and E/I imbalance in the ACC in neuropathic pain.
前扣带皮层（ACC）的过度兴奋与慢性疼痛的发展有关。作为 ACC 过度兴奋的主要原因之一，ACC 的去抑制可能与抑制性 Parvalbumin（PV）表达中间神经元（PV-INs）的功能障碍密切相关。然而，ACC PV-INs 损伤的分子机制尚不清楚。本研究表明， spared sciatic nerve injury（SNI）导致 ACC 的兴奋和抑制（E/I）失衡。为了验证 SNI 后 ACC 中肿瘤坏死因子-α（TNF-α）的上调是否激活坏死性凋亡并参与 PV-INs 损伤，我们使用神经病理性疼痛模型的转录组测序数据进行了差异分析，发现 TNF-α-坏死性凋亡途径相关基因的表达上调。SNI 大鼠 ACC 中 TNF-α 免疫反应性（IR）信号与 p-RIP3 和 PV-IR 或 p-MLKL 和 PV-IR 信号共定位。然后，我们系统地检测了 SNI 大鼠 ACC 中坏死性凋亡相关蛋白的表达和细胞定位，包括激酶 RIP1、RIP3、MLKL 及其磷酸化状态。除了 RIP1 和 MLKL 之外，这些蛋白的水平在对侧 ACC 中显著升高，主要在 PV-INs 中表达。通过在 ACC 中微注射 TNF-α 中和抗体或使用 siRNA 敲低来阻断 MLKL 的表达，阻断 ACC TNF-α-坏死性凋亡途径减轻了 SNI 诱导的痛觉过敏，并抑制了 TNF-α 和 p-MLKL 的上调。靶向 ACC PV-INs 内 TNF-α 触发的坏死性凋亡可能有助于纠正神经病理性疼痛中 ACC 内的 PV-INs 损伤和 E/I 失衡。
