Effect of Fractalkine Signaling on Neuroinflammation and Neuropathic Pain-like Behavior in Mice

Neuropathic pain is the most prevalent and incapacitating form of chronic pain. Understanding its pathogenesis at the cellular and molecular levels is crucial to guide the development of new therapeutics. A major controversy has been on the roles of Fractalkine (CX3CL1) signaling between neurons and immune cells in the pathogenesis of neuropathic pain. This study investigated two distinct pathways of CX3CL1 signaling in the pathogenesis of neuropathic pain, namely the CX3CR1-dependent pathway and the transforming growth factor β (TGF-β)/Smad2/3-dependent pathway.

Neuropathic pain-like behavior was induced in mice using the sciatic nerve chronic constriction injury (CCI) model. Experiments were performed in wild-type, CX3CR1 knockout, and CX3CL1-transgenic mice to examine distinct CX3CL1 signaling pathways. Microglia and macrophage activation, CX3CL1 processing, TGF-β/Smad2/3 signaling, and inflammatory cytokine expression were assessed using molecular and immunohistochemical techniques. Behavioral assays quantified mechanical allodynia and thermal hyperalgesia, and transcriptomic analyses evaluated neuroimmune gene expression changes after CCI injury and CX3CL1 signaling manipulations.

CCI of the sciatic nerve led to activation of microglia and macrophages, cleavage and consumption of full-length CX3CL1, and a decrease in the expression of its downstream TGF-β family proteins. Deletion of CX3CR1 in CX3CR1-knockout mice exacerbated CCI-induced neuroinflammation. Conversely, overexpression of CX3CL1 in CX3CL1-Tg mice significantly enhanced TGF-β/Smad2/3 signaling, reduced proinflammatory cytokines, inhibited microglia activation, and alleviated CCI-induced hyperalgesia. Moreover, CX3CL1 overexpression effectively reversed the CCI-induced differentially expressed genes and neuroimmune responses.

These findings demonstrate that enhancing CX3CL1 signaling effectively mitigates neuroinflammation and alleviates neuropathic pain-like behavior, likely through CX3CL1 signaling between neuronal and immune cells via two distinct pathways: the CX3CR1-dependent pathway mediated through the CX3CL1 N terminus and the TGF-β-dependent pathway mediated through the CX3CL1 C terminus.