CX3CR1 deficiency suppresses activation and neurotoxicity of microglia/macrophage in experimental ischemic stroke

Abstract

BackgroundChemokine (C-X3-C motif) ligand 1 (CX3CL1)/ CX3C chemokine receptor 1 (CX3CR1) signaling is important in modulating the communication between neurons and resident microglia/migrated macrophages in the central nervous system (CNS). Although CX3CR1 deficiency is associated with an improved outcome following ischemic brain injury, the mechanism of this observation is largely unknown. The aim of this study was to investigate how CX3CR1 deficiency influences microglia/macrophage functions in the context of its protection following brain ischemia.MethodsWild-type (WT) and CX3CR1-deficient (CX3CR1-/-) mice were subjected to transient middle cerebral artery occlusion (MCAO) and reperfusion. The ischemic brain damage was monitored by rodent high-field magnetic resonance imaging. Neurological deficit was assessed daily. Neuronal apoptotic death and reactive oxygen species (ROS) production were analyzed by immunostaining and live imaging. Activation/inflammatory response of microglia/macrophage were assessed using immunohistochemistry, flow cytometry, 5-bromo-2-deoxyuridine labeling, cytokine ELISA, and real-time PCR.ResultsCX3CR1-/- mice displayed significantly smaller infarcts and less severe neurological deficits compared to WT controls, following MCAO. In addition, CX3CR1-/- MCAO mice displayed fewer apoptotic neurons and reduced ROS levels. Impaired CX3CR1 signaling abrogated the recruitment of monocyte-derived macrophages from the periphery, suppressed the proliferation of CNS microglia and infiltrated macrophage, facilitated the alternative activation (M2 state) of microglia/macrophages, and attenuated their ability to synthesize and release inflammatory cytokines.ConclusionOur results suggest that inhibition of CX3CR1 signaling could function as a therapeutic modality in ischemic brain injury, by reducing recruitment of peripheral macrophages and expansion/activation of CNS microglia and macrophages, resulting in protection of neurological function.

Neuroimmune Response in Ischemic Preconditioning

“…Neurons are protected from OGD after short-term exposure to hypoxia, and this protection lasts for up to 48 h after the preconditioning stimulus [128]. Pathways implicated as protective included the inhibition of caspase-12 after preconditioning, but not prolonged ischemia, and activation of multiple unfolded protein response pathways [128]. Finally, recent studies suggest that 14-3-3γ, a multifunctional scaffolding protein expressed in astrocytes in response to IPC, is also upregulated in neuronal cultures in response to OGD [103].…”

“…There is some evidence that endogenous cannabinoids, acting through the CB 1 receptor and G proteins, may protect neurons against glutamatemediated injury [126], as well as other injury mechanisms [127], suggesting CB 1 receptors may be a potential therapeutic target for preconditioning. Neurons are protected from OGD after short-term exposure to hypoxia, and this protection lasts for up to 48 h after the preconditioning stimulus [128]. Pathways implicated as protective included the inhibition of caspase-12 after preconditioning, but not prolonged ischemia, and activation of multiple unfolded protein response pathways [128].…”

Leukocyte Invasion of the Brain After Experimental Intracerebral Hemorrhage in Mice

“…According to previous reports, [27][28][29][30][31][32] low /CD11b high cells were defined as microglia and were excluded from the calculation of infiltrating leukocyte numbers. Although the hematoma volume was reduced by day 5, the number of brain-infiltrating leukocytes rose significantly between day 1 and day 5 in both ICH models ( Figure 1D).…”

“…Similar to several previous studies including 2 in experimental stroke using 2-photon imaging and immunohistochemistry, we distinguished microglial cells from infiltrating leukocytes based on their lower CD45 expression. [27][28][29][30][31][32]39,40 A limitation of this approach is that CD45 can be upregulated on microglia as demonstrated by conversion toward the CD45 high phenotype in experimental autoimmune encephalitis. 41 We found a peak of all infiltrating leukocyte subsets 5 days after blood injection.…”

Correction to: Neuroimmune Response in Ischemic Preconditioning

“…After neuronal injury, including injury caused by ischemia, the loss of the fractalkine ligand expression on the surface of neurons results in enhanced microglial activation in several models of inflammation [19,129]. In the early stages following ischemia, however, deficiency in CX 3 CR1 signaling suppresses activation of microglia/macrophages, reduces neurotoxicity, and leads to a reduction in poststroke infarct volume [130,131]. The microglial/macrophage response to CX 3 CL1/CX 3 CR1 signaling in ischemia likely evolves over time and the net effect of CX 3 CL1/ CX 3 CR1 signaling in IPC remains to be determined.…”