Spinal Cord Stimulation Alleviates Spinal Cord Injury-Induced Neuropathic Pain Via m

Spinal cord injury (SCI) often results in neuropathic pain (NP), which remains a major clinical challenge. Spinal cord stimulation (SCS) has emerged as a promising therapy for SCI-induced NP; however, its underlying mechanisms are not fully elucidated. 5-Methylcytosine (m5C) modification plays a critical role in various pathological processes, yet its involvement in SCI-induced NP has not been explored. This study aimed to investigate whether the therapeutic effects of SCS on SCI-induced NP are associated with m5C modification. An SCI mouse model was induced. Behavioral tests (evaluating mechanical allodynia and thermal hyperalgesia), enzyme-linked immunosorbent assays (measuring inflammatory cytokines), and histopathological analyses (hematoxylin and eosin staining and TUNEL staining) were performed. Bioinformatics analysis of the GSE256472 dataset was conducted to identify m5C-related genes involved in NP. Lentiviral-mediated overexpression or knockdown of NOP2/Sun RNA methyltransferase 2 (NSUN2) or TNF receptor-associated factor 2 (TRAF2) was employed to validate their functional roles. In vitro, BV2 microglial cells were transfected to modulate NSUN2 expression, followed by m⁵C-RNA immunoprecipitation, dual-luciferase reporter assays, and RNA stability assays to dissect the molecular mechanism. SCS significantly reversed mechanical allodynia and thermal hyperalgesia, suppressed neuroinflammation, and attenuated neuronal apoptosis in SCI mice. Bioinformatics and experimental validation revealed that NSUN2—a core m⁵C methyltransferase—was markedly upregulated in both SCI-induced NP patients and mice, and this elevation was normalized by SCS. Functional studies demonstrated that NSUN2 overexpression abolished the analgesic and neuroprotective benefits of SCS, whereas NSUN2 knockdown mimicked SCS effects. Mechanistically, NSUN2 directly deposited m⁵C modifications at position 929 of TRAF2 mRNA, enhancing its stability and expression. TRAF2 overexpression counteracted the therapeutic effects of NSUN2 inhibition, reinstating inflammation, apoptosis, and pain hypersensitivity. SCS alleviated SCI-induced NP by downregulating NSUN2, thereby reducing m⁵C-mediated stabilization of TRAF2 mRNA and subsequent activation of pro-inflammatory and pro-apoptotic pathways. The NSUN2/TRAF2 axis represents a novel epitranscriptomic mechanism in NP and a potential target for precision interventions in SCI-related pain management.