Brain endothelial CXCL12 attracts protective natural killer cells during ischemic stroke.

BACKGROUND: The innate lymphoid cell (ILC) family consists of NK cells, ILC type 1, 2, 3 and lymphoid tissue inducer cells. They have been shown to play important roles in homeostasis and immune responses and are generally considered tissue resident. Not much is known about the presence of ILC members within the central nervous system and whether they are tissue resident in this organ too. Therefore, we studied the presence of all ILC members within the central nervous system and after ischemic brain insult. METHODS: We used the photothrombotic ischemic lesion method to induce ischemic lesions within the mouse brain. Using whole-mount immunofluorescence imaging, we established that the ILCs were present at the rim of the lesion. We quantified the increase of all ILC members at different time-points after the ischemic lesion induction by flow cytometry. Their migration route via chemokine CXCL12 was studied by using different genetic mouse models, in which we induced deletion of Cxcl12 within the blood-brain barrier endothelium, or its receptor, Cxcr4, in the ILCs. The functional role of the ILCs was subsequently established using the beam-walk sensorimotor test. RESULTS: Here, we report that ILCs are not resident within the mouse brain parenchyma during steady-state conditions, but are attracted towards the ischemic stroke. Specifically, we identify NK cells, ILC1s, ILC2s and ILC3s within the lesion, the highest influx being observed for NK cells and ILC1s. We further show that CXCL12 expressed at the blood-brain barrier is essential for NK cells and NKp46+ ILC3s to migrate toward the lesion. Complementary, Cxcr4-deficiency in NK cells prevents NK cells from entering the infarct area. Lack of NK cell migration results in a higher neurological deficit in the beam-walk sensorimotor test. CONCLUSIONS: This study establishes the lack of ILCs in the mouse central nervous system at steady-state and their migration towards an ischemic brain lesion. Our data show a role for blood-brain barrier-derived CXCL12 in attracting protective NK cells to ischemic brain lesions and identifies a new CXCL12/CXCR4-mediated component of the innate immune response to stroke.

Stroke induces disease-specific myeloid cells in the brain parenchyma and pia.

Inflammation triggers secondary brain damage after stroke. The meninges and other CNS border compartments serve as invasion sites for leukocyte influx into the brain thus promoting tissue damage after stroke. However, the post-ischemic immune response of border compartments compared to brain parenchyma remains poorly characterized. Here, we deeply characterize tissue-resident leukocytes in meninges and brain parenchyma and discover that leukocytes respond differently to stroke depending on their site of residence. We thereby discover a unique phenotype of myeloid cells exclusive to the brain after stroke. These stroke-associated myeloid cells partially resemble neurodegenerative disease-associated microglia. They are mainly of resident microglial origin, partially conserved in humans and exhibit a lipid-phagocytosing phenotype. Blocking markers specific for these cells partially ameliorates stroke outcome thus providing a potential therapeutic target. The injury-response of myeloid cells in the CNS is thus compartmentalized, adjusted to the type of injury and may represent a therapeutic target.

Cxcr4 distinguishes HSC-derived monocytes from microglia and reveals monocyte immune responses to experimental stroke.

Monocyte-derived and tissue-resident macrophages are ontogenetically distinct components of the innate immune system. Assessment of their respective functions in pathology is complicated by changes to the macrophage phenotype during inflammation. Here we find that Cxcr4-CreER enables permanent genetic labeling of hematopoietic stem cells (HSCs) and distinguishes HSC-derived monocytes from microglia and other tissue-resident macrophages. By combining Cxcr4-CreER-mediated lineage tracing with Cxcr4 inhibition or conditional Cxcr4 ablation in photothrombotic stroke, we find that Cxcr4 promotes initial monocyte infiltration and subsequent territorial restriction of monocyte-derived macrophages to infarct tissue. After transient focal ischemia, Cxcr4 deficiency reduces monocyte infiltration and blunts the expression of pattern recognition and defense response genes in monocyte-derived macrophages. This is associated with an altered microglial response and deteriorated outcomes. Thus, Cxcr4 is essential for an innate-immune-system-mediated defense response after cerebral ischemia. We further propose Cxcr4-CreER as a universal tool to study functions of HSC-derived cells.