Zhang Xingyuan, Hou Yuxuan, Cai Rixin, Zhou Yue, He Bingqiang, Cao Zhilong, Li Aicheng, Song Honghua, Wang Yingjie, Jiang Haiyan, Qi Lei, Wang Yongjun
Key Laboratory of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, Nantong University, Nantong, 226001, People's Republic of China.
Department of Intensive Care Unit, Nantong Third People's Hospital, Nantong, 226001, People's Republic of China.
J Inflamm Res. 2025 Jun 11;18:7651-7669. doi: 10.2147/JIR.S505433. eCollection 2025.
D-dopachrome tautomerase (D-DT), a homolog of macrophage migration inhibitory factor (MIF), has been revealed to promote astrocytic inflammation and worsen neuropathology following spinal cord injury (SCI). So far, the mechanism about D-DT-activated astrocytic inflammation remains elusive. Ubiquitin-specific peptidase 18 (Usp18) is an active player in regulating innate immunity through ISG15-deconjugated dependent or independent manner in multiple cell types. Whether D-DT activates astrocytic inflammation regulation of Usp18 deserves further study.
SCI model was prepared by cord contusion at T8-T10 of rats. The expression changes of D-DT and Usp18 were examined by ELISA, Western blot, RT-PCR or immunohistochemistry. Primary astrocytes were treated by different concentration of D-DT, either for transcriptome sequencing or for analysis of D-DT-mediated expression of Usp18. Knockdown of CD74 or Usp18 expression was performed by siRNA transfection of astrocytes. The locomotor functions were assessed using the Basso, Beattie, and Bresnahan (BBB) locomotor scale.
Usp18 was significantly upregulated in the astrocytes at lesion sites following SCI, in parallel with the elevation of D-DT protein levels. D-DT inhibitor 4-CPPC remarkably decreased the astrocytic expression of USP18. Transcriptome sequencing of D-DT-stimulated primary astrocytes identified that Usp18 was the hub modulator of D-DT-mediated astrocytic inflammation. D-DT-mediated expression of Usp18 was able to activate MAPKs, contributing to the production of proinflammatory cytokines and chemokines. Specifically, phosphorylation of P38 kinase was shown to promote the expression of Usp18 by formation of a positive feedback loop. Administration of D-DT inhibitor 4-CPPC at lesion sites following SCI significantly reduced the protein levels of Usp18 and ameliorated functional deficits of rat hindlimb locomotion.
SCI-induced elevation of D-DT at lesion sites activates astrocytic inflammation upregulating the expression of Usp18. Identification of this novel regulator associated with astrocytic inflammation will provide an alternative target for clinical therapy of neuroinflammation.
D - 多巴色素互变异构酶（D - DT）是巨噬细胞迁移抑制因子（MIF）的同源物，已被证实可促进脊髓损伤（SCI）后的星形胶质细胞炎症反应并加重神经病理学改变。到目前为止，D - DT激活星形胶质细胞炎症反应的机制仍不清楚。泛素特异性肽酶18（Usp18）是多种细胞类型中通过ISG15去共轭依赖性或非依赖性方式调节先天免疫的活跃参与者。D - DT是否通过调节Usp18激活星形胶质细胞炎症反应值得进一步研究。
通过大鼠T8 - T10脊髓挫伤制备SCI模型。采用酶联免疫吸附测定（ELISA）、蛋白质印迹法、逆转录 - 聚合酶链反应（RT - PCR）或免疫组织化学检测D - DT和Usp18的表达变化。用不同浓度的D - DT处理原代星形胶质细胞，用于转录组测序或分析D - DT介导的Usp18表达。通过对星形胶质细胞进行小干扰RNA（siRNA）转染来敲低CD74或Usp18的表达。使用Basso、Beattie和Bresnahan（BBB）运动评分量表评估运动功能。
SCI后损伤部位的星形胶质细胞中Usp18显著上调，与D - DT蛋白水平升高同步。D - DT抑制剂4 - CPPC显著降低了USP18的星形胶质细胞表达。对D - DT刺激的原代星形胶质细胞进行转录组测序发现，Usp18是D - DT介导的星形胶质细胞炎症反应的核心调节因子。D - DT介导的Usp18表达能够激活丝裂原活化蛋白激酶（MAPKs），促进促炎细胞因子和趋化因子的产生。具体而言，P38激酶的磷酸化通过形成正反馈环促进Usp18的表达。SCI后在损伤部位给予D - DT抑制剂4 - CPPC可显著降低Usp18的蛋白水平，并改善大鼠后肢运动功能缺陷。
SCI诱导损伤部位D - DT升高，通过上调Usp18的表达激活星形胶质细胞炎症反应。鉴定这种与星形胶质细胞炎症反应相关的新型调节因子将为神经炎症的临床治疗提供一个替代靶点。
