After SCI, a heterogeneous macrophage response, comprised of resident microglia and blood monocytes, dominates the lesion environment and exerts divergent effects on indices of neurodegeneration and tissue repair. These CNS macrophages vigorously respond to injury, relying on environmental cues for chemoattraction and activation. These heterogenous macrophage response may be regulated by spatiotemporally distinct microenvironments to which each cell is subjected. At the molecular level, this regulation may involve signaling through CX3CR1. Fractalkine (CX3CL1) is a unique chemokine and immunoregulator that binds to the receptor CX3CR1, present on microglia and monocytes. In the healthy CNS, neuronal expression of CX3CL1 promotes a resting microglial phenotype. In the context of injury however, CX3CL1 shed by damaged neurons and endothelia promotes the recruitment and activation of microglia and monocytes. What role CX3CR1 signaling plays in the microglial and monocyte response to SCI has to this point been unexplored.
Here we show that intraspinal CX3CL1 and CX3CR1 are dynamically regulated after SCI and that abolishing CX3CR1 signaling in SCI mice confers neuroprotection and promotes functional recovery. This is associated with a significant phenotypic alteration of responding macrophages. Deficient CX3CR1 signaling in CNS macrophages attenuates their ability to synthesize and release inflammatory cytokines and oxidative metabolites. CX3CR1-deficient mice had an increased accumulation of CCR2+Ly6Chi macrophages. Importantly, iNOS expression was virtually abolished on CX3CR1-deficient macrophages. In addition, MHC II expression was also decreased on CX3CR1-deficient macrophages, potentially decreasing harmful lymphocyte activation. These data suggest that CX3CR1 deficiency prevents or alters the recruitment and/or differentiation of harmful macrophages after SCI. Further experiments with chimeric mice showed that mice with wild-type microglia but CX3CR1-deficient monocytes had the highest degree of locomotor recovery after SCI. This work may potentially lead to development of cell-specific therapeutic strategies for tailoring the immune response towards neuroprotection after SCI.
What role the spleen, and in particular splenic monocytes, play in the immunopathology associated with SCI has to this point also been unexplored. If different peripheral sources of monocytes contribute different monocyte subsets, then discerning the source of these monocytes may allow future therapies to target certain populations. We found that in normal wild-type mice after SCI, that the circulating monocyte pool remains relatively constant, even during robust intraspinal accumulation. Splenic numbers decrease immediately after injury, suggesting that the spleen may be responsible for a large degree of the initial buffering of the circulation. In addition, there is a colossal increase in the bone marrow population after SCI, suggesting injury-induced myelopoiesis. The monocyte/macrophage population in the spinal cord decreases through 14 days post-injury in splenectomized mice. The phenotype of intraspinal macrophages is also altered with splenectomy. However, we found no effect on locomotor recovery or tissue sparing resulting from splenectomy. Thus, a more extensive characterization of the role of the spleen in post-SCI inflammation is warranted.