Huang Wenjiao, Chen Wenli, Zhao Zhong, Liu Lingchun, Zhao Yuanyuan, Chen Xinzhang, Li Rong
School of Medicine, Kunming University of Science and Technology, No. 727 Jingming South Road, Kunming, 650500, Yunnan, China.
Department of Neurology, The First People's Hospital of Yunnan Province, No. 157 Jinbi Road, Kunming, 650034, Yunnan, China.
Sci Rep. 2025 Jul 2;15(1):22958. doi: 10.1038/s41598-025-04914-4.
Epilepsy (EP) is a chronic nervous system disease characterized by recurrent attacks, and its causes are complicated. Inflammatory reaction mediated by microglia is an important factor in the progression of EP. Activating transcription factor 2 (ATF2) can be used as a transcription factor to regulate the microglia-mediated inflammatory response, but its role in EP is unclear. In this study, kainic acid (KA) was used to induce the EP cell and mouse model. Real-time polymerase chain reaction was used to detect ATF2, TNF-α, IL-6, TGF-β, and IL-10 mRNA expression. ATF2, INOS, ARG1, and TSC1 protein levels was examined by western blot. The fluorescence intensity of ATF2, IBA1, CD80, and CD206 was examined by immunofluorescence staining. The cell ratios of CD80, IL-1β, CD206, and CD63 were detected by flow cytometry. Dual-luciferase reporter and chromatin immunoprecipitation assays were conducted to verify the interaction between ATF2 and TSC1. Hematoxylin & eosin and Nissl staining were used to observe the structure of hippocampus and Nissl bodies. The results indicated that KA induced M1 polarization of HMC3 cells and increased the levels of TNF-α and IL-6 mRNA by activating KA receptors, and inhibiting KA receptors attenuated the M1 polarization of KA-induced HMC3 cells. ATF2 expression was increased in KA-induced HMC3 cells and hippocampal tissues of mouse, while TSC1 expression was repressed. ATF2 knockdown diminished the M1 polarization of KA-induced HMC3 cells, enhanced the M2 polarization, and relieved neuroinflammation in EP mouse. TSC1 overexpression inhibited M1 polarization in KA-induced HMC3 cells. Dual luciferase and chromatin immunoprecipitation results revealed that ATF2 bound to the promoter of TSC1 and negatively regulated the transcription of TSC1. In conclusion, inhibition of ATF2 and promotion of TSC1 transcription may be a new pathophysiological mechanism for the treatment of EP neuroinflammation.
癫痫（EP）是一种以反复发作为特征的慢性神经系统疾病，其病因复杂。小胶质细胞介导的炎症反应是EP进展的一个重要因素。激活转录因子2（ATF2）可作为一种转录因子来调节小胶质细胞介导的炎症反应，但其在EP中的作用尚不清楚。在本研究中，使用 kainic 酸（KA）诱导 EP 细胞和小鼠模型。采用实时聚合酶链反应检测 ATF2、TNF-α、IL-6、TGF-β 和 IL-10 mRNA 的表达。通过蛋白质免疫印迹法检测 ATF2、诱导型一氧化氮合酶（INOS）、精氨酸酶1（ARG1）和结节性硬化复合物1（TSC1）蛋白水平。通过免疫荧光染色检测 ATF2、离子钙结合衔接分子1（IBA1）、CD80 和 CD206 的荧光强度。通过流式细胞术检测 CD80、IL-1β、CD206 和 CD63 的细胞比例。进行双荧光素酶报告基因和染色质免疫沉淀试验以验证 ATF2 与 TSC1 之间的相互作用。采用苏木精-伊红染色和尼氏染色观察海马结构和尼氏体。结果表明，KA 通过激活 KA 受体诱导 HMC3 细胞发生 M1 极化，并增加 TNF-α 和 IL-6 mRNA 水平，而抑制 KA 受体可减弱 KA 诱导的 HMC3 细胞的 M1 极化。在 KA 诱导的 HMC3 细胞和小鼠海马组织中，ATF2 表达增加，而 TSC1 表达受到抑制。敲低 ATF2 可减弱 KA 诱导的 HMC3 细胞的 M1 极化，增强 M2 极化，并减轻 EP 小鼠的神经炎症。TSC1 过表达抑制 KA 诱导的 HMC3 细胞的 M1 极化。双荧光素酶和染色质免疫沉淀结果显示，ATF2 与 TSC1 的启动子结合并负向调节 TSC1 的转录。总之，抑制 ATF2 和促进 TSC1 转录可能是治疗 EP 神经炎症的一种新的病理生理机制。
