Spinal cord injury (SCI) causes sensation and motion dysfunction. Activation of microglial cells (MCs) in the central nervous system (CNS) is heterogeneous. Heterogeneous types of MCs can produce cytotoxic or neuroprotective effects, secrete proinflammatory or anti-inflammatory factors. The cytotoxic effect of MCs is one of the reasons for secondary damage after SCI. The NLR family pyrin domain containing 3 (NLRP3) inflammasome is a protein that can recognize pathogen-related molecular patterns or host-derived danger signal molecules, responses to microbial infection, and sterile stressors. SCI triggers activation of the NLRP3 inflammasome in the CNS. We investigated the interaction between miR-423-5p and NLRP3 in MCs polarization after SCI. A rat model of SCI was established by a modified version of Allen's method. Spinal samples were adopted for preparation and sequencing of RNA. We screenedapromising microRNA (miR-423-5p) according to the results. Then, we found that NLRP3 was one of the prediction targets of miR-423-5p. By intervening in expression of miR-423-5p and NLRP3, we observed the different polarization of MCs. We employeda dual-luciferase reporter study, proteomics, and transcriptomicsto ascertain the direct targeting relationship between miR-423-5p and NLRP3. MiR-423-5p expression was decreased significantly after SCI in vivo and in vitro. Upregulation of miR-423-5p expression could prevent MCs from lipopolysaccharide-induced M1 polarization. Knockdown of NLRP3 expression could prevent MCs from lipopolysaccharide-induced M1 polarization. MiR-423-5p inhibited MCs polarization to the M1 phenotype by targeting NLRP3.