Growing evidence supports the analgesic efficacy of electroacupuncture (EA) in managing chronic neuropathic pain (NP) in both patients and NP models induced by peripheral nerve injury. However, the underlying mechanisms remain incompletely understood. Ferroptosis, a novel form of programmed cell death, has been found to be activated during NP development, while EA has shown potential in promoting neurological recovery following acute cerebral injury by targeting ferroptosis. In this study, to investigate the detailed mechanism underlying EA intervention on NP, male Sprague‐Dawley rats with chronic constriction injury (CCI)‐induced NP model received EA treatment at acupoints ST36 and GV20 for 14 days. Results demonstrated that EA effectively attenuated CCI‐induced pain hypersensitivity and mitigated neuron damage and loss in the spinal cord of NP rats. Moreover, EA reversed the oxidative stress‐mediated spinal ferroptosis phenotype by upregulating reduced expression of xCT, glutathione peroxidase 4 (GPX4), ferritin heavy chain (FTH1) and superoxide dismutase (SOD) levels, and downregulating increased expression of acyl‐CoA synthetase long‐chain family member 4 (ACSL4), malondialdehyde levels and iron overload. Furthermore, EA increased the immunofluorescence co‐staining of GPX4 in neurons cells of the spinal cord of CCI rats. Mechanistic analysis unveiled that the inhibition of antioxidant pathway of Nrf2 signalling via its specific inhibitor, ML385, significantly countered EA's protective effect against neuronal ferroptosis in NP rats while marginally diminishing its analgesic effect. These findings suggest that EA treatment at acupoints ST36 and GV20 may protect against NP by inhibiting neuronal ferroptosis in the spinal cord, partially through the activation of Nrf2 signalling.

Background: Patients with neuropathic pain (NP), caused by injury or disease of the somatosensory nervous system, usually suffer from severe pain. Our previous studies revealed that electroacupuncture (EA) stimulation could effectively improve NP. However, the underlying mechanisms of EA have not been fully clarified. This study aimed to investigate the specific mechanisms of EA in alleviating NP, focusing on the pyroptosis. Materials and Methods: Chronic Constriction Injury (CCI) model was established on the male Sprague-Dawley rats. CCI rats were treated with EA at acupoints GV20 and ST36 or/with the NOD-like receptor protein 3 (NLRP3) antagonist MCC950. EA treatment was administered for successive 14 days 7 days after the CCI surgery. The mechanical withdrawal threshold (MWT) and paw withdrawal latency (PWL) were performed during the experiment. At the end of the experiment, spinal cord segments and serum of rats were collected, ELISA detected the expression of inflammatory factors, immunofluorescence detected the microglia and neuron cells with pyroptosis biomarkers, and Western blot detected the NLRP3 pathway. Results: EA treatment significantly alleviated pain hypersensitivity by increasing the MWT and PWL. Moreover, EA reduced levels of pro-inflammatory cytokines IL-1β and TNF-α in the spinal tissue. Mechanistically, the pyroptosis-related proteins, including NLRP3, N-GSDMD, Cleaved Caspase-1, IL-18 as well as IL-1β were downregulated by EA, indicating that EA attenuated the pyroptosis phenotype in NP rats. In particular, EA reduced the co-expression of NLRP3, Caspase-1 and N-GSDMD in microglia rather than in neuronal or astrocytic cells within the spinal cord of CCI rats. Pharmacological inhibition of NLRP3 inflammasome by MCC950 alleviates CCI-induced pain hypersensitivity while blocking EA’s effect on anti-pyroptosis in CCI rats. Conclusion: These findings demonstrate the EA ameliorates the neuroinflammation and pyroptosis to relieve chronic NP by suppressing NLRP3 inflammasome activation in microglia. EA may serve as a viable treatment therapy for chronic NP.

Https://doi.org/10.18632/oncotarget.28048 Shuyi Kong, Jing Wang, Hui Xu, Kaiqiang Wang