gw-4869 and Peripheral-Nerve-Injuries

Our study is aimed at investigating the mechanism by which electroacupuncture (EA) promoted nerve regeneration by regulating the release of exosomes and exosome-mediated miRNA-21 (miR-21) transmission. Furthermore, the effects of Schwann cells- (SC-) derived exosomes on the overexpression of miR-21 for the treatment of PNI were investigated.. A sciatic nerve injury model of rat was constructed, and the expression of miR-21 in serum exosomes and damaged local nerves was detected using RT-qPCR after EA treatment. The exosomes were identified under a transmission electron microscope and using western blotting analysis. Then, the exosome release inhibitor, GW4869, and the miR-21-5p-sponge used for the knockdown of miR-21 were used to clarify the effects of exosomal miR-21 on nerve regeneration promoted by EA. The nerve conduction velocity recovery rate, sciatic nerve function index, and wet weight ratio of gastrocnemius muscle were determined to evaluate sciatic nerve function recovery. SC proliferation and the level of neurotrophic factors were assessed using immunofluorescence staining, and the expression levels of SPRY2 and miR-21 were detected using RT-qPCR analysis. Subsequently, the transmission of exosomal miR-21 from SC to the axon was verified. We found that EA inhibited the release of serum exosomal miR-21 in a PNI model of rats during the early stage of PNI, while it promoted its release during later stages. EA enhanced the accumulation of miR-21 in the injured nerve and effectively promoted the recovery of nerve function after PNI. The treatment effect of EA was attenuated when the release of circulating exosomes was inhibited or when miR-21 was downregulated in local injury tissue via the miR-21-5p-sponge. Normal exosomes secreted by SC exhibited the ability to promote the recovery of nerve function, while the overexpression of miR-21 enhanced the effects of the exosomes. In addition, exosomal miR-21 secreted by SC could promote neurite outgrowth. Our results demonstrated the mechanism of EA on PNI from the perspective of exosome-mediated miR-21 transport and provided a theoretical basis for the use of exosomal miR-21 as a novel strategy for the treatment of PNI.

Topics: Aniline Compounds; Animals; Benzylidene Compounds; Cell Line, Transformed; Disease Models, Animal; Electroacupuncture; Exosomes; Gene Expression; Gene Expression Regulation; Gene Knockdown Techniques; Male; MicroRNAs; Nerve Regeneration; Nerve Tissue Proteins; Peripheral Nerve Injuries; Rats; Rats, Wistar; Recovery of Function; Schwann Cells; Sciatic Nerve; Signal Transduction; Transfection

Recent observations indicate that peripheral nerve injury induces central sensitization through microglial activation and the release of inflammatory cytokines, resulting in the development of neuropathic pain. However, the underlying mechanisms of this phenomenon remain to be fully elucidated. In this study, we examined the involvement of spinal ceramide, a bioactive lipid, in the development of neuropathic pain induced by partial sciatic nerve ligation (PSL). We found that the mRNA expression levels for ceramide synthase and neutral sphingomyelinase, which are enzymes of ceramide biosynthesis, were up-regulated in the spinal cord from 3h to 1 day after PSL. The mRNA expressions of cytokines (interleukin-1β and tumor necrosis factor-α) and the microglial specific molecules (Iba-1 and CD11b) were also increased in the spinal cord after PSL. In the von Frey test, intrathecal injection of the ceramide biosynthesis inhibitors Fumonisin B1 and GW4869 at 3h and day 3 after PSL significantly attenuated PSL-induced tactile allodynia. By immunohistochemistry, microglial activation in the dorsal horn was suppressed by Fumonisin B1 and GW4869. Therefore, we conclude that spinal ceramide may play a crucial role in PSL-induced neuropathic pain through the activation of microglia.

Topics: Aniline Compounds; Animals; Benzylidene Compounds; Ceramides; Cytokines; Fumonisins; Hyperalgesia; Ligation; Male; Mice; Mice, Inbred ICR; Microglia; Neuralgia; Oxidoreductases; Peripheral Nerve Injuries; RNA, Messenger; Sciatic Nerve; Sphingomyelin Phosphodiesterase; Spinal Cord