CX3CL1 attenuates neurological deficit and neuroinflammation through CX3CR1/p38 MAPK/ERK1/2 signaling pathway in traumatic brain injury.

BACKGROUND

Traumatic brain injury (TBI) induces a complex cascade of secondary pathological events, including neuroinflammation and oxidative stress, which substantially contribute to neuronal damage and long-term neurological deficits. The chemokine CX3CL1 (fractalkine) and its unique receptor CX3CR1constitute a critical signaling axis mediating neuron-microglia communication. However, the specific mechanisms by which the CX3CL1/CX3CR1 axis modulates TBI-induced neuroinflammatory responses remain incompletely understood.

METHODS

We established an in vitro oxidative stress model using H2O2-treated human microglial HMC3 cells and an in vivo mouse TBI model to study the CX3CL1/CX3CR1 axis. RNA sequencing identified differentially expressed genes. Protein expression and localization were analyzed by Western blot and immunofluorescence. Functional roles were investigated using CX3CR1 overexpression, knockdown, and pharmacological modulation of mitogen-activated protein kinase (MAPK). Microglial polarization and cytokine release were measured by flow cytometry and ELISA. Recombinant CX3CL1 (r-CX3CL1) was administered intranasally to validate therapeutic effects.

RESULTS

CX3CR1 expression significantly increased after oxidative stress in vitro and after TBI in vivo, peaking at 12 h and day 3, respectively. CX3CL1 localized primarily to neurons and astrocytes, while CX3CR1 was mainly found on microglia. CX3CR1 overexpression promoted M2 microglial polarization, suppressed inflammation, inhibited p38 MAPK, and activated ERK1/2 signaling. Conversely, CX3CR1 knockdown enhanced M1 polarization and inflammation, partially reversible by MAPK inhibition. In TBI mice, intranasal r-CX3CL1 improved microglial polarization and neurological recovery through MAPK signaling, effects abolished in CX3CR1 knockout mice.

CONCLUSIONS

The CX3CL1/CX3CR1 axis alleviates neuroinflammation and neurological deficits post-TBI by modulating microglial polarization via the MAPK pathway, highlighting its therapeutic potential.