Electroacupuncture alleviates neuropathic pain caused by SNL by promoting M2 microglia polarization through PD-L1.

As a common clinical disease, neuropathic pain is difficult to be cured with drugs. The occurrence and progression of pain is closely related to the response of spinal microglia. Aspartof the regulation of microglialactivity,PD-L1 playsacriticalrole. Loss of PD-L1 promoted the polarization of M1-like microglia. Increased expression of PD-L1 promoted M2-like polarization. Electroacupuncture has a significant analgesic effect in clinical practice, but its specific mechanism remains to be further explored. In this study, we verified the role of PD-L1 in EA analgesia and the underlying molecular mechanism through spinal nerve ligation (SNL) in rats and lipopolysaccharide (LPS)-treated BV2 microglial cells. Forbehavioralstudiesofrats,mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were measured, and spinal cord neuros were examined under transmission electron microscopyto determine changes to their myelin structure. The expression levels of PD-L1 and M1/M2-specific markers in rat spinal cord and BV2 microglial cells were measured by enzyme-linked immunosorbent assay, flow cytometry, immunofluorescence staining and Western blot analysis. Our study showed that EA increased the pain threshold, reduced the destruction of myelin structure, promoted the expression of PD-L1 and PD-1, inhibited the MAPK signaling pathway, and promoted the conversion of microglial polarization from the M1 phenotype to the M2 phenotype in SNL rats. PD-L1 knockdown reversed these effects of EA. In addition, PD-L1 knockdown activated the MAPK signaling pathway, promoted microglial polarization to the M1 phenotype, decreased the expression of anti-inflammatory mediators and increased the expression of proinflammatory factors in LPS-stimulated BV2 microglial cells. Our results showed that EA may regulate the excitability of primary afferent neurons through PD-L1 and then inhibit the MAPK signaling pathway to promote the transformation of activated M1 microglia into M2 microglia, reduce inflammatory reactions, and finally achieve analgesic effects. A therapy targeting PD-L1 may be an effective strategy for treating neuropathic pain.

Electroacupuncture inhibits dendritic spine remodeling through the srGAP3-Rac1 signaling pathway in rats with SNL.

Previous studies have shown that peripheral nerve injury can lead to abnormal dendritic spine remodeling in spinal dorsal horn neurons. Inhibition of abnormal dendritic spine remodeling can relieve neuropathic pain. Electroacupuncture (EA) has a beneficial effect on the treatment of neuropathic pain, but the specific mechanism remains unclear. Evidence has shown that slit-robo GTPase activating protein 3 (srGAP3) and Rho GTPase (Rac1) play very important roles in dendritic spine remodeling. Here, we used srGAP3 siRNA and Rac1 activator CN04 to confirm the relationship between SrGAP3 and Rac1 and their roles in improving neuropathic pain with EA. Spinal nerve ligation (SNL) was used as the experimental model, and thermal withdrawal latency (TWL), mechanical withdrawal threshold (MWT), Western blotting, immunohistochemistry and Golgi-Cox staining were used to examine changes in behavioral performance, protein expression and dendritic spines. More dendritic spines and higher expression levels of srGAP3 were found in the initial phase of neuropathic pain. During the maintenance phase, dendritic spines were more mature, which was consistent with lower expression levels of srGAP3 and higher expression levels of Rac1-GTP. EA during the maintenance phase reduced the density and maturity of dendritic spines of rats with SNL, increased the levels of srGAP3 and reduced the levels of Rac1-GTP, while srGAP3 siRNA and CN04 reversed the therapeutic effects of EA. These results suggest that dendritic spines have different manifestations in different stages of neuropathic pain and that EA may inhibit the abnormal dendritic spine remodeling by regulating the srGAP3/Rac1 signaling pathway to alleviate neuropathic pain.

Electroacupuncture inhibits dendritic spine remodeling through the srGAP3-Rac1 signaling pathway in rats with SNL

Previous studies have shown that peripheral nerve injury can lead to abnormal dendritic spine remodeling in spinal dorsal horn neurons. Inhibition of abnormal dendritic spine remodeling can relieve neuropathic pain. Electroacupuncture (EA) has a beneficial effect on the treatment of neuropathic pain, but the specific mechanism remains unclear. Evidence has shown that slit-robo GTPase activating protein 3 (srGAP3) and Rho GTPase (Rac1) play very important roles in dendritic spine remodeling. Here, we used srGAP3 siRNA and Rac1 activator CN04 to confirm the relationship between SrGAP3 and Rac1 and their roles in improving neuropathic pain with EA. Spinal nerve ligation (SNL) was used as the experimental model, and thermal withdrawal latency (TWL), mechanical withdrawal threshold (MWT), Western blotting, immunohistochemistry and Golgi-Cox staining were used to examine changes in behavioral performance, protein expression and dendritic spines. More dendritic spines and higher expression levels of srGAP3 were found in the initial phase of neuropathic pain. During the maintenance phase, dendritic spines were more mature, which was consistent with lower expression levels of srGAP3 and higher expression levels of Rac1-GTP. EA during the maintenance phase reduced the density and maturity of dendritic spines of rats with SNL, increased the levels of srGAP3 and reduced the levels of Rac1-GTP, while srGAP3 siRNA and CN04 reversed the therapeutic effects of EA. These results suggest that dendritic spines have different manifestations in different stages of neuropathic pain and that EA may inhibit the abnormal dendritic spine remodeling by regulating the srGAP3/Rac1 signaling pathway to alleviate neuropathic pain.