CCL2 activates NLRP3 inflammasomes to regulate microglial phenotype transformation to promote central sensitisation...

Central sensitisation is a significant pathogenic mechanism underlying chronic migraine (CM). Mounting evidence suggests that this process is driven by central and peripheral neuroinflammation. Although increased CC motif chemokine ligand 2 (CCL2) is present in the cerebrospinal fluid (CSF) of patients with CM, whether it is a contributing factor remain unclear. In our study, we simulated CM by injecting inflammatory soup (IS) intraventricularly into rats for 7 days. Microglia expressing CD68 and CD163 were labelled with immunofluorescence staining, indicating pro and anti-inflammatory phenotypes, respectively. Enzyme-linked immunosorbent assay (ELISA) detected the cytokines interleukin (IL)-1β and IL-10, and western blotting (WB) detected Fos proto-oncogene (c-Fos) and p-extracellular signal-regulated kinase (ERK), which are markers of central sensitisation. Mechanical and thermal nociceptives were used to assess pain associated with CM. Following IS-induced modelling, protein levels of CCL2 and its receptor CCR2 increased, and CCL2 was co-localised with astrocytes while CCR2 was associated with microglia. With CCL2 recombinant protein treatment, microglia shifted to a proinflammatory phenotype with increased IL-1β and decreased IL-10 levels. In addition, central sensitisation was enhanced and mechanical and thermal pain thresholds were reduced. These adverse effects were reversed by CCL2 inhibitor and NOD-like receptor protein 3 (NLRP3) inhibitor downstream of CCL2. Significantly, MCC950 reversed the microglial phenotypic changes induced by recombinant CCL2 and attenuated central sensitisation in CM rats. These data suggest that CCL2 in the trigeminal nucleus caudalis (TNC) regulates the microglial phenotype by activating NLRP3, promoting central sensitisation of CM.