Diagnosing and treating chronic orofacial pain is challenging due to its complex structure and limited understanding of its causes and mechanisms. In this study, we used RNA sequencing to identify differentially expressed genes (DEGs) in the rostral ventral medulla (RVM) and thalamus of rats with persistent orofacial pain, aiming to explore its development. DEGs were functionally analyzed using gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Results showed a significant association between immune response and pain in this model. Key DEG mRNA expression trends were further validated using real-time quantitative polymerase chain reaction (RT-PCR), confirming their crucial roles in chronic orofacial pain. After injecting complete Freund’s adjuvant (CFA) into the bilateral temporomandibular joint cavity for 14 days, we observed 293 upregulated genes and 14 downregulated genes in the RVM, and 1086 upregulated genes and 37 downregulated genes in the thalamus. Furthermore, we identified 27 common DEGs with altered expression (upregulation) in both the thalamus and RVM, including Cd74, C3, Cxcl13, C1qb, Itgal, Fcgr2b, C5ar1, and Tlr2, which are pain-associated genes. Protein-protein interaction (PPI) analysis using Cytoscape revealed the involvement of Toll-like receptors, complement system, differentiation clusters, and antigen presentation-related proteins in the interaction between the thalamus and RVM. The results of this study show that the immune system seems to have a more significant influence on chronic orofacial pain. There may be direct or indirect influence between the thalamus and RVM, which may participate in the regulation of chronic orofacial pain.

由于其复杂的结构以及对其原因和机制的了解有限，诊断和治疗慢性口面部疼痛具有挑战性。在本研究中，我们利用RNA测序来鉴定持续性口面部疼痛大鼠的头端腹侧延髓（RVM）和丘脑中的差异表达基因（DEG），旨在探讨其发展过程。使用基因本体论（GO）和京都基因和基因组百科全书（KEGG）通路分析对DEG进行功能分析。结果显示该模型中免疫反应与疼痛之间存在显着关联。使用实时定量聚合酶链反应 (RT-PCR) 进一步验证了关键 DEG mRNA 表达趋势，证实了它们在慢性口面部疼痛中的关键作用。在双侧颞下颌关节腔注射完全弗氏佐剂（CFA）14天后，我们在RVM中观察到293个上调基因和14个下调基因，在丘脑中观察到1086个上调基因和37个下调基因。此外，我们还鉴定了 27 个在丘脑和 RVM 中表达改变（上调）的常见 DEG，包括Cd74、C3、Cxcl13、C1qb、Itgal、Fcgr2b、C5ar1和Tlr2，这些都是疼痛相关基因。使用 Cytoscape 进行的蛋白质-蛋白质相互作用 (PPI) 分析揭示了 Toll 样受体、补体系统、分化簇和抗原呈递相关蛋白质参与丘脑和 RVM 之间的相互作用。这项研究的结果表明，免疫系统似乎对慢性口面部疼痛有更显着的影响。丘脑和RVM之间可能存在直接或间接的影响，可能参与慢性口面部疼痛的调节。