中国组织工程研究

中国组织工程研究 ›› 2026, Vol. 30 ›› Issue (11): 2712-2726.doi: 10.12307/2026.119

• 脊柱组织构建 spinal tissue construction • 上一篇    下一篇

IRF9基因在脊髓损伤后神经保护中的调控作用：生信分析与实验验证

田明昊，廖烨晖，周文洋，何宝强，冷叶波，徐世财，周佳俊，李  洋，唐  超，唐  强，钟德君   

  1. 西南医科大学附属医院骨科，四川省泸州市   646000

  • 收稿日期:2025-02-07 接受日期:2025-06-19 出版日期:2026-04-18 发布日期:2025-09-02
  • 通讯作者: 钟德君，教授，硕士生导师，西南医科大学附属医院骨科，四川省泸州市 646000
  • 作者简介:田明昊，男，1997年生，四川省德阳市人，汉族，硕士在读，主要从事脊柱外科相关疾病的基础与临床研究。
  • 基金资助:
    四川省医学会项目(Q22008)，项目负责人：廖烨晖；泸州市人民政府与西南医科大学科技战略合作项目(2023LZXNYDJ038)，项目负责人：廖烨晖；西南医科大学附属医院博士科研启动基金项目(22155)，项目负责人：廖烨晖；西南医科大学应用基础研究项目(2023QN070)，项目负责人：廖烨晖；四川省自然科学基金项目(2024NSFSC0682)，项目负责人：钟德君；西南医科大学科研基金项目(2024LCYXZX38)，项目负责人：钟德君

Neuroprotective regulation of the IRF9 gene after spinal cord injury: bioinformatics analysis combined with experimental validation

Tian Minghao, Liao Yehui, Zhou Wenyang, He Baoqiang, Leng Yebo, Xu Shicai, Zhou Jiajun, Li Yang, Tang Chao, Tang Qiang, Zhong Dejun   

  1. Department of Orthopedics, Affiliated Hospital of Southwest Medical University, Luzhou 646000, Sichuan Province, China 
  • Received:2025-02-07 Accepted:2025-06-19 Online:2026-04-18 Published:2025-09-02
  • Contact: Zhong Dejun, Professor, Master’s supervisor, Department of Orthopedics, Affiliated Hospital of Southwest Medical University, Luzhou 646000, Sichuan Province, China
  • About author:Tian Minghao, MS candidate, Department of Orthopedics, Affiliated Hospital of Southwest Medical University, Luzhou 646000, Sichuan Province, China
  • Supported by:
    Sichuan Medical Association Project, No. Q22008 (to LYH); Luzhou Municipal People’s Government and Southwest Medical University Science and Technology Strategic Cooperation Project, No. 2023LZXNYDJ038 (to LYH); Doctoral Research Initiation Fund Project of the Affiliated Hospital of Southwest Medical University, No. 22155 (to LYH); Applied Basic Research Program of Southwest Medical University, No. 2023QN070 (to LYH); Natural Science Foundation of Sichuan Province, No. 2024NSFSC0682 (to ZDJ); Southwest Medical University Scientific Research Fund Program, No. 2024LCYXZX38 (to ZDJ)

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摘要：


文题释义：
IRF9：是干扰素调控因子家族成员之一，主要参与抗病毒免疫和细胞抗病原体感染的信号传导过程。 
生物信息分析：是利用计算机工具、方法和技术对生命科学数据进行分析和研究的方法，是对大量生物信息数据进行有效处理、挖掘和分析的重要手段。

背景：脊髓损伤后炎症的具体调控机制尚不清楚。研究表明干扰素调节因子9(IRF9)在多种炎症相关疾病中发挥关键作用，但IRF9在脊髓损伤中的具体作用尚未被系统性研究。
目的：结合生信分析与体外实验系统探讨IRF9在脊髓损伤后炎症反应中的作用及其潜在的分子机制。
方法：①通过整合GEO数据库的基因芯片数据，采用差异基因分析、GO和KEGG功能富集分析、基因集富集分析及蛋白互作网络分析探讨脊髓损伤后差异基因的功能及通路，筛选与炎症相关的关键基因。②将第3代PC12细胞分4组培养：空白对照组不进行任何处理，损伤对照组加入脂多糖干预24 h，IRF9过表达组转染IRF9过表达慢病毒72 h后加入脂多糖干预24 h，IRF9敲低组转染IRF9敲低表达慢病毒72 h后加入脂多糖干预24 h。脂多糖干预结束后，CCK-8法检测细胞增殖，Elisa检测细胞培养基中脑源性神经生长因子和神经营养因子3水平，Western blot检测细胞内Bcl-2、Bax、Cleaved-Caspase3蛋白表达，划痕实验检测细胞迁移能力。
结果与结论：①生物信息学分析显示，脊髓损伤后显著激活了炎症和免疫相关通路，筛选出IRF9为潜在核心调控因子，IRF9高表达可能上调Toll样受体和核因子κB信号通路，同时抑制环磷酸腺苷和钙信号通路及谷氨酸能突触功能，参与炎症反应和神经功能障碍的调控；②细胞实验结果显示，与空白对照组比较，损伤对照组细胞增殖、迁移能力及Bcl-2蛋白表达降低，脑源性神经生长因子和神经营养因子3水平降低，Bax与Cleaved-Caspase-3蛋白表达升高；与损伤对照组比较，IRF9敲低组细胞增殖、迁移能力及Bcl-2蛋白表达升高，脑源性神经生长因子和神经营养因子3水平升高，Bax与Cleaved-Caspase-3蛋白表达降低；③结果表明，IRF9可能是脊髓损伤后继发性损伤的重要调控因子和潜在治疗靶点。

关键词: IRF9, 脊髓损伤, 生物信息学, PC12细胞, 炎症反应, 免疫调控, 细胞凋亡, 继发性损伤, 工程化组织构建

Abstract: BACKGROUND: The precise regulatory mechanisms of inflammation following spinal cord injury remain unclear. In recent years, studies have shown that interferon regulatory factor 9 (IRF9) plays a key role in various inflammation-related diseases, but its specific role in spinal cord injury has not been systematically studied.
OBJECTIVE: To systematically investigate the role of IRF9 in the inflammatory response after spinal cord injury and its potential molecular mechanisms by integrating bioinformatics analysis and in vitro experiments.  
METHODS: (1) Gene microarray data from the GEO database were integrated and analyzed using differential gene expression analysis, GO and KEGG functional enrichment analyses, gene set enrichment analysis, and protein-protein interaction network analysis to explore the functions and pathways of differentially expressed genes after spinal cord injury and identify the key genes related to inflammation. (2) The 3rd generation PC12 cells were cultured in four groups: blank control group without any treatment, injury control group with lipopolysaccharide intervention for 24 hours, IRF9 overexpression group transfected with IRF9 overexpression lentivirus for 72 hours followed by lipopolysaccharide intervention for 24 hours, and IRF9 knockdown group transfected with IRF9 knockdown expression lentivirus for 72 hours followed by lipopolysaccharide intervention for 24 hours. After the end of lipopolysaccharide intervention, cell proliferation was detected by cell counting kit-8 assay, brain-derived nerve growth factor and neurotrophic factor 3 levels in cell culture medium were detected by Elisa, intracellular Bcl-2, Bax, Cleaved-Caspase3 protein expression was detected by western blot, and cell migration ability was detected by cell scratch assay. 
RESULTS AND CONCLUSION: (1) Bioinformatics analysis demonstrated that spinal cord injury significantly activated inflammation- and immune-related pathways, identifying IRF9 as a potential core regulator. High IRF9 expression was found to potentially upregulate Toll-like receptor and nuclear factor-κB signaling pathways while suppressing cyclic adenosine monophosphate and calcium signaling pathways and glutamatergic synaptic function, contributing to the regulation of inflammation and neurological dysfunction. (2) The results of cellular experiments showed that compared with the blank control group, the injury control group had lower cell proliferation, migration ability and Bcl-2 protein expression, lower levels of brain-derived nerve growth factor and neurotrophic factor 3, and higher expression of Bax and Cleaved-Caspase-3 proteins. Compared with the injury control group, the IRF9 knockdown group showed higher cell proliferation, migration ability and Bcl-2 protein expression, higher levels of brain-derived nerve growth factor and neurotrophic factor 3, and lower expression of Bax and Cleaved-Caspase-3 proteins. To conclude, IRF9 may serve as an important regulator of secondary injury and a potential therapeutic target for spinal cord injury.

Key words: IRF9, spinal cord injury, bioinformatics, PC12 cells, inflammatory response, immune regulation, apoptosis, secondary injury, engineered tissue engineering

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