Apomycin relieves neuropathic pain by attenuating NRLP 3-medicated inflammasome activation via inhibiting NADPH induced ROS in microglia

Neuroinflammation in the peripheral and central nervous system plays an important role in the induction and maintenance of chronic pain. However, the potential mechanism of ROS production from inflammasome activation in microglia during neuroinflammation remains elusive. We performed spinal nerve ligation (SNL) model for chronic pain and implanted intrathecal catheter for intrathecal application of apomycin. Behavioral tests including von Frey hairs and paw withdrawal tests were constructed on postoperative days 1, 3, 5, 7, 14 and after apomycin delivery to assess mechanical threshold and heat hyperalgesia. ELISA of IL-1β production and real-time PCR assay of IL-1β, IL-6 and TNF-α in supernatant from microglia cells pretreatment of apomycin were performed. Besides, caspase-1 content in spinal lysis from different groups including sham-operated rats with vehicle injection, shamoperated rats with apomycin injection, SNL-injured rats with vehicle injection, SNL-injured rats with apomycin and 12 h after injection were detected by western blot analysis and also CD4 and CD8 mRNA expression were measured by Q-PCR assay. Here, we first found that apomycin significantly decreased the IL-1β production which was induced by NRLP3 activation in microglia cells, while the TLR4-ligation induced mRNA transcription of IL-1β, and other proinflammatory cytokines, such as IL-6 and TNF-α were not significantly inhibited by apomycin pretreatment. However, the activation of NF-κB and MAPK signaling pathway was only slightly decreased by apomycin. Furthermore, supplement of ROS in culture abolished the inhibition of IL-1β by apomycin. Our data suggest that apomycin ameliorates neuropathic pain by attenuating NRLP3 activation induced IL-1β production through inhibition of NADPH induced ROS in microglia, which outlines the novel application of apomycin in treatment of neuropathic pain and the novel potential signal pathway of apomycin.