miR‐26a‐5p alleviates CFA‐induced chronic inflammatory hyperalgesia through Wnt5a/CaMKII/NFAT signaling in mice

BACKGROUND: Inflammation often leads to the occurrence of chronic pain, and many miRNAs have been shown to play a key role in the development of inflammatory pain. However, whether miR‐26a‐5p relieves pain induced by inflammation and its possible mechanism are still unclear. METHODS: The complete Freund's adjuvant (CFA)‐induced inflammatory pain mouse model was employed. Intrathecal or subcutaneous injection of miR‐26a‐5p agomir was performed after modeling to study its antinociceptive effect and the comparison of different administration methods. Bioinformatics analysis of miRNAs was performed to study the downstream mechanisms of miR‐26a‐5p. HE staining, RT‐qPCR, Western blotting, and immunofluorescence were used for further validation. RESULTS: A single intrathecal and subcutaneous injection of miR‐26a‐5p both reversed mechanical hypersensitivity and thermal latency in the left hind paw of mice with CFA‐induced inflammatory pain. HE staining and immunofluorescence studies found that both administrations of miR‐26a‐5p alleviated inflammation in the periphery and spinal cord. Bioinformatics analysis and dual‐luciferase reporter gene analysis identified Wnt5a as a direct downstream target gene of miR‐26a‐5p. Wnt5a was mainly expressed in neurons and microglia in the spinal cord of mice with inflammatory pain. Intrathecal injection of miR‐26a‐5p could significantly reduce the expression level of Wnt5a and inhibit the downstream molecules of noncanonical Wnt signaling Camk2/NFAT, inhibiting the release of spinal cord inflammatory factors and alleviating the activation of microglia. In addition, miR‐26a‐5p could also inhibit lipopolysaccharide (LPS)‐stimulated BV2 cell inflammation in vitro through a noncanonical Wnt signaling pathway. CONCLUSIONS: miR‐26a‐5p is a promising therapy for CFA‐induced inflammatory pain. Both intrathecal and subcutaneous injections provide relief for inflammatory pain. miR‐26a‐5p regulated noncanonical Wnt signaling to be involved in analgesia partly through antineuroinflammation, suggesting a pain‐alleviating effect via noncanonical Wnt signaling pathway in the CFA‐induced inflammatory pain model in vivo.